The suppressor of cytokine signalling 2 (SOCS2) is a key repressor of insulin secretion.

AIMS/HYPOTHESIS: Suppressor of cytokine signalling (SOCS) proteins are powerful inhibitors of pathways involved in survival and function of pancreatic beta cells. Whereas SOCS1 and SOCS3 have been involved in immune and inflammatory processes, respectively, in beta cells, nothing is known about SOCS2 implication in the pancreas. METHODS: Transgenic (tg) mice were generated that constitutively produced SOCS2 in beta cells (betaSOCS2) to define whether this protein is implicated in beta cell functioning and/or survival. RESULTS: Constitutive production of SOCS2 in beta cells leads to hyperglycaemia and glucose intolerance. This phenotype is not a consequence of decreased beta cell mass or inhibition of insulin synthesis. However, insulin secretion to various secretagogues is profoundly altered in intact animals and isolated islets. Interestingly, constitutive SOCS2 production dampens the rise in cytosolic free calcium concentration induced by glucose, while glucose metabolism is unchanged. Moreover, tg islets have a depletion in endoplasmic reticulum Ca(2+) stores, suggesting that SOCS2 interferes with calcium fluxes. Finally, in betaSOCS2 mice proinsulin maturation is impaired, leading to an altered structure of insulin secretory granules and augmented levels of proinsulin. The latter is likely to be due to decreased production of prohormone convertase 1 (PC1/3), which plays a key role in proinsulin cleavage. CONCLUSIONS/INTERPRETATIONS: SOCS2 was shown to be a potent regulator of proinsulin processing and insulin secretion in beta cells. While its constitutive production is insufficient to induce overt diabetes in this mouse model, it causes glucose intolerance. Thus, increased SOCS2 production could be an important event predisposing to beta cell failure.

Comparative effects of Citrullus colocynthis, sunflower and olive oil-enriched diet in streptozotocin-induced diabetes in rats.

Citrullus colocynthis (colocynth) seeds are traditionally used as antidiabetic medication in Mediterranean countries. The present study evaluated the differential effects of diets enriched with C. colocynthis, sunflower or olive oils on the pancreatic beta-cell mass in streptozotocin (STZ)-induced diabetes in rats. STZ injection induced rapid hyperglycaemia in all animals. However, 2 months later, hyperglycaemia was significantly less pronounced in the rats fed a C. colocynthis oil-enriched diet compared with other rat groups (7.9mM versus 12mM and 16mM with colocynth versus olive and sunflower oils, respectively). Assessment of insulin sensitivity using the homoeostasis model assessment (HOMA) method also indicated less insulin resistance in the rats fed a C. colocynthis oil-enriched diet versus the other rats. Finally, 2 months after STZ injection, the pancreatic beta-cell mass was similar in both the STZ-treated rats fed the colocynth oil-enriched diet and their controls fed the same diet. In contrast, the pancreatic beta-cell mass remained lower in the STZ-induced diabetic rats fed with olive oil- and sunflower oil-enriched diets compared with the C. colocynthis group. We conclude that C. colocynthis oil supplementation may have a beneficial effect by partly preserving or restoring pancreatic beta-cell mass in the STZ-induced diabetes rat model.

Improvement of metabolic state in an animal model of nutrition-dependent type 2 diabetes following treatment with S 23521, a new glucagon-like peptide 1 (GLP-1) analogue.

Glucagon-like peptide 1 (GLP-1) analogues are considered potential drugs for type 2 diabetes. We studied the effect of a novel GLP-1 analogue, S 23521 ([a8-des R36] GLP-1-[7-37]-NH2), on the metabolic state and beta-cell function, proliferation and survival in the Psammomys obesus model of diet-induced type 2 diabetes. Animals with marked hyperglycaemia after 6 days of high-energy diet were given twice-daily s.c. injection of 100 microg/kg S 23521 for 15 days. Food intake was significantly decreased in S 23251-treated P. obesus; however, there was no significant difference in body weight from controls. Progressive worsening of hyperglycaemia was noted in controls, as opposed to maintenance of pre-treatment glucose levels in the S 23521 group. Prevention of diabetes progression was associated with reduced mortality. In addition, the treated group had higher serum insulin, insulinogenic index and leptin, whereas plasma triglyceride and non-esterified fatty acid levels were decreased. S 23521 had pronounced effect on pancreatic insulin, which was 5-fold higher than the markedly depleted insulin reserve of control animals. Immunohistochemical analysis showed islet degranulation with disrupted morphology in untreated animals, whereas islets from S 23521-treated animals appeared intact and filled with insulin; beta-cell apoptosis was approximately 70% reduced, without a change in beta-cell proliferation. S 23521 treatment resulted in a 2-fold increase in relative beta-cell volume. Overall, S 23521 prevented the progression of diabetes in P. obesus with marked improvement of the metabolic profile, including increased pancreatic insulin reserve, beta-cell viability and mass. These effects are probably due to actions of S 23521 both directly on islets and via reduced food intake, and emphasize the feasibility of preventing blood glucose deterioration over time in type 2 diabetes.

Sialylated form of the neural cell adhesion molecule (NCAM): a new tool for the identification and sorting of beta-cell subpopulations with different functional activity.

To clarify the relationship between variations in beta-cell mass and pancreatic function, we investigated the possibility to analyze, quantify, and sort beta-cell subpopulations with different functional maturity. To this aim, we tested the reliability of the sialylated form of neural cell adhesion molecule (NCAM) (PSA-NCAM) as a marker of beta-cell functional activity. Islet cells isolated from adult rats were analyzed for their PSA-NCAM abundance using an anti-PSA-NCAM antibody. We found that PSA-NCAM is expressed only in beta-cells. The PSA-NCAM labeling was also studied with a fluorescence-activated cell sorter. We showed that the beta-cell population is heterogeneous for PSA-NCAM labeling. To directly determine the relationship between PSA-NCAM labeling and beta-cell activity, in vitro insulin secretion studies were performed on sorted beta-cell subpopulations using a perifusion technique. Two beta-cell subpopulations were analyzed: one that was highly labeled for PSA-NCAM and another that was poorly labeled. Insulin secretion from high PSA-NCAM-labeled beta-cells was significantly higher than that in low PSA-NCAM-labeled beta-cells. This differential expression in the beta-cell population was well correlated with differences in glucose responsiveness. PSA-NCAM seems thus suitable for use as a tool to identify beta-cell subpopulations according to their glucose responsiveness.

Neogenesis vs. apoptosis As main components of pancreatic beta cell ass changes in glucose-infused normal and mildly diabetic adult rats.

We have investigated in adult rats made mildly diabetic by a low dose of streptozotocin (35 mg/kg; STZ rats) and in nondiabetic rats (ND rats) the mechanisms leading to adaptive changes in the beta cell mass, during glucose infusion and several days after stopping infusion. As early as 24 h of glucose infusion, the beta cell mass was maximally increased in ND and STZ rats. In both groups, this increase was due mainly to a rapid activation of neogenesis of new endocrine cells rather than to an increase in beta cell proliferation. Seven days after stopping glucose infusion, the beta cell mass returned to basal values in both groups as a result of stimulation of beta cell apoptosis and a decrease in beta cell replication rate. In glucose-infused ND rats, changes in the beta cell mass were correlated to insulin secretion, whereas in STZ rats, insulin secretion in response to glucose was still impaired whatever the beta cell mass. In conclusion, the data stress the impressive plasticity of the endocrine pancreas of adult rats. They also show that changes in beta cell mass in ND and STZ rats resulted from a disruption in the balance between neogenesis and apoptosis.

Lipid infusion lowers sympathetic nervous activity and leads to increased beta-cell responsiveness to glucose.

We investigated the possible involvement of the autonomic nervous system in the effect of a long-term elevation of plasma free fatty acid (FFA) concentration on glucose-induced insulin secretion (GIIS) in rats. Rats were infused with an emulsion of triglycerides (Intralipid) for 48 hours (IL rats). This resulted in a twofold increase in plasma FFA concentration. At the end of infusion, GIIS as reflected in the insulinogenic index (DeltaI/DeltaG) was 2.5-fold greater in IL rats compared with control saline-infused rats. The ratio of sympathetic to parasympathetic nervous activities was sharply decreased in IL rats relative to controls. GIIS was studied in the presence of increasing amounts of alpha- and beta-adrenoreceptor agonists and antagonists. The lowest concentrations of the alpha2A-adrenoreceptor agonist oxymetazoline, which were ineffective in control rats, reduced GIIS in IL rats. At the dose of 0.3 pmol/kg, GIIS became similar in IL and control rats. The use of beta-adrenoreceptor agonist (isoproterenol) or antagonist (propranolol) did not result in a significant alteration in GIIS in both groups. GIIS remained as high in IL vagotomized rats as in intact IL rats, indicating that changes in parasympathetic tone were of minor importance. Altogether, the data show that lipid infusion provokes beta-cell hyperresponsiveness in vivo, at least in part through changes in alpha2-adrenergic innervation.

Rilmenidine normalizes fructose-induced insulin resistance and hypertension in rats.

OBJECTIVE: The aim of this study was to determine the effects of rilmenidine (an antihypertensive drug that lowers blood pressure by decreasing sympathetic outflow) in an animal model of hypertension associated with insulin resistance, i.e. rats fed on a high-fructose diet. DESIGN: Wistar rats were fed for 4 weeks either on a standard diet (S group) or on a high-fructose diet (F group; 34.5% fructose). In half of the rats in the F group, rilmenidine (1 mg/kg per day) was added to the drinking water for the last 2 weeks of the diet (FR group). RESULTS: Body weight gain was higher in the F than in the S rats (66+/-8g versus 45+/-8g, P< 0.05), but was prevented by rilmenidine treatment (32+/-2g). Arterial systolic blood pressure was increased in F rats (162+/-2 versus 155+/-2 mmHg, P< 0.05), rilmenidine reduced this value to normal (149+/-3 mmHg). Glucose tolerance, glucose turnover rate, and insulin secretion were not modified by the diet or by the drug. However, during a euglycemic hyperinsulinemic clamp, glucose utilization was lower (10+/-1 versus 14+/-1.5 mg/min per kg; P< 0.05) and hepatic glucose production higher (1+/-0.01 versus 0 mg/min per kg, P< 0.01) in F than in S rats. These changes in insulin action were totally abolished by rilmenidine. CONCLUSIONS: These data demonstrate that rilmenidine can ameliorate the deleterious effects of a high-fructose diet, i.e. weight gain, hypertension, and resistance to the effects of insulin.

Effects of red blood cell hyperaggregation on the rat microcirculation blood flow.

This study presents the effects of red blood cell (RBC) hyperaggregation on the blood flow and pressure in the rat mesentery and cremaster network. We exclusively studied in situ non-vasodilated organs, in order to maintain the physiological regulation mechanisms. Dextran 500 was injected at different concentrations to increase RBC aggregation. The aggregation rate was measured on very small blood samples with an erythroaggregameter (SEFAM) which evaluated the disaggregating shear stress (tau D) needed to break the RBC aggregates. Microscopic observations and laser Doppler velocimetry were used to quantify the flow rate. The plasmatic dextran concentration (C) increase had different correlated effects: for example, tau D increased from 3 dynes cm-2 (for the control sample) to 14 dynes cm-2 (for C = 75 microM L-1); the flow rate was reduced threefold and very large aggregates were observed in the venules; the arteriolar pressure increased while venular pressure decreased. In order to differentiate the effects of RBC hyperaggregation from those of plasma hyperviscosity (due to dextran 500) on microcirculatory blood flow, we injected an RBC antiaggregating drug (troxerutine) (50 or 100 mg kg-1 i.v.). The consequences were a high reduction for (tau D) (from 14 dynes cm(-2)-9 dynes cm-2), smaller aggregates and higher blood flow in the venules. No effect of troxerutine was observed on plasma viscosity (plasma control: 1.9 cP with or without troxerutine; plasma with dextran at C = 75 microM L-1: 2.45 cP with or without troxerutine). The results strongly suggest that RBC aggregation has a significant influence on blood flow rate in the microcirculatory network.

Pancreatic beta-cell regeneration after 48-h glucose infusion in mildly diabetic rats is not correlated with functional improvement.

We investigated the effect of glucose infusion on beta-cell regeneration in rats made mildly diabetic by a single injection of low dosage (35 mg/kg) streptozotocin (STZ). Nondiabetic (ND) and STZ rats were submitted to a 48-h glucose infusion (hyperglycemia approximately 22 mmol/l in both groups: ND and STZ hyperglycemic-hyperinsulinemic [ND HG-HI and STZ HG-HI rats]). Before infusion, beta-cell mass was 65% lower in STZ rats than in ND rats (2.0 +/- 0.02 vs. 5.5 +/- 0.6 mg), 1.6-fold increased in ND HG-HI rats (8.7 +/- 1.7 mg), and 2.7-fold increased in STZ HG-HI rats (5.4 +/- 0.9 mg). In ND HG-HI rats, beta-cell enlargement was related to an increase in beta-cell responsiveness to nutrient secretagogues both in vivo and in vitro, whereas in STZ HG-HI rats, no significant improvement in insulin secretion could be noticed. To determine the respective role of hyperglycemia and hyperinsulinemia on beta-cell area changes, ND and STZ rats were submitted to a 48-h hyperinsulinemic-euglycemic clamp. No modification of beta-cell mass was detected in either group. In conclusion, 48-h superimposed hyperglycemia was enough to restore beta-cell mass previously reduced by STZ injection. This effect seemed to be due to hyperglycemia rather than hyperinsulinemia alone. The data stress the dissociation between beta-cell regeneration and improvement in islet function in diabetic rats. Our model seems suitable for studying factors that can improve the plasticity and function of the pancreas in NIDDM.

Pioglitazone induces in vivo adipocyte differentiation in the obese Zucker fa/fa rat.

Thiazolidinediones are potent antidiabetic compounds, in both animal and human models, which act by enhancing peripheral sensitivity to insulin. Thiazolidinediones are high-affinity ligands for peroxisome proliferator-activated receptor-gamma, a key factor for adipocyte differentiation, and they are efficient promoters of adipocyte differentiation in vitro. Thus, it could be questioned whether a thiazolidinedione therapy aimed at improving insulin sensitivity would promote the recruitment of new adipocytes in vivo. To address this problem, we have studied the in vivo effect of pioglitazone on glucose metabolism and gene expression in the adipose tissue of an animal model of obesity with insulin resistance, the obese Zucker (fa/fa) rat. Pioglitazone markedly improves insulin action in the obese Zucker (fa/fa) rat, but doubles its weight gain after 4 weeks of treatment. The drug induces a large increase of glucose utilization in adipose tissue, where it stimulates the expression of genes involved in lipid metabolism such as the insulin-responsive GLUT, fatty acid synthase, and phosphoenolpyruvate carboxykinase genes, but decreases the expression of the ob gene. These changes are related to both an enhanced adipocyte differentiation, as shown by the large increase in the number of small adipocytes in the retroperitoneal fat pad, and a direct effect of pioglitazone on specific gene expression (phosphoenolpyruvate carboxykinase and ob genes) in mature adipocytes.

[Effects of rilmenidine on rats made insulin resistant and hypertensive by a high fructose diet]. / Effets de la rilménidine chez le rat rendu insulinorésistant et hypertendu par un régime riche en fructose.

This study was aimed to determine the effects of rilmenidine, an hypertensive drug, in an animal model of hypertension associated with insulin resistance, i.e. rats fed on a high fructose diet. Wistar rats were fed during four weeks either on a standard diet (S) or on a high fructose diet (F, 34.5% de fructose). In half of the F groups, rilmenidine (1 mg/kg/day) was added to the drinking water during the two last weeks of the diet (FR). Arterial blood pressure as well as insulin efficiency were determined at the end of the four weeks. Body weight gain was higher in F than in S rats (66 +/- 8 g versus 45 +/- 8 g; p < 0.05), this was prevented by rilmenidine treatment (32 +/- 2 g). Arterial systolic blood pressure was increased in F rats (162 +/- 2 vs 155 +/- 2 mmHg; p < 0.05), rilmenidine brought this value back to normal (149 +/- 3 mmHg). During the euglycemic hyperinsulinemic clamp, glucose utilization was lower (10 +/- 1 vs 14 +/- 1.5 mg/min/kg; p < 0.05) and hepatic glucose production higher (1 +/- 0.01 vs 0 mg/min/kg; p < 0.01) in F than in S rats. These changes in insulin action were totally abolished by rilmenidine. These data demonstrate that rilmenidine can ameliorate the deleterious effects of a high fructose diet, i.e. weight gain, hypertension and resistance to the effects of insulin Rilmenidine could represent a potential therapeutic agent for the treatment of hypertension associated with metabolic disorders such as syndrom X and obesity.

Circulatory alterations induced by intra-arterial injection of iodized oil and emulsions of iodized oil and doxorubicin: experimental study.

PURPOSE: To evaluate circulatory alterations induced by intra-arterial injection of iodized oil and emulsions of iodized oil with an anticancer drug. MATERIALS AND METHODS: The viscosities and stability were evaluated in arterial flow of iodized oil and the emulsions. Doppler ultrasound was used to quantify the embolic effect in the arteries of rabbits. The behavior of doxorubicin and iodized oil in the rat cremaster muscle was studied with videomicroscopy. RESULTS: The emulsions did not break up after injection, and the size of discontinuous-phase droplets did not change. The embolic effect did not correlate with viscosity. The thinnest water-in-oil emulsion had the lowest embolic effect and induced homogeneous distribution of iodized oil droplets in the arterial tree. CONCLUSION: Changes in the formulation of emulsions obtained with the same proportions of drug and iodized oil change the distribution of iodized oil in the arterial tree, the location at which the drug is released, and the embolic effect.

Local sympathetic denervation of white adipose tissue in rats induces preadipocyte proliferation without noticeable changes in metabolism.

The direct influence of the sympathetic nervous system on white adipose tissue was studied by performing a unilateral surgical denervation of the retroperitoneal fat pad in rats, the contralateral pad being used as a control. One week after surgery, the weight of the denervated pad was significantly higher than that of the intact pad. In vivo, glucose utilization was not altered by denervation. The expression of GLUT4 as well as the expression and activity of fatty acid synthase, lipoprotein lipase, and hormone-sensitive triglyceride lipase were similar in the two pads. Lipolysis in response to norepinephrine, determined in vitro, was not modified by denervation although the ratio between alpha 2- and beta-adrenergic receptors was changed. Denervation induced an increase in DNA content without change in the number of mature adipocytes. The expression of A2COL6/pOb24, a marker of the early step of adipocyte differentiation, was significantly enhanced in the denervated pad, suggesting an increased number of preadipocytes. This was confirmed by an increased cell number observed in the denervated fat pad 1 month after surgery. In conclusion, surgical denervation of the white fat pad does not alter the glucose and lipid metabolisms. By contrast, it accelerated adipocyte differentiation and led to the recruitment of new precursors.

Regulation of glucose homeostasis in pre-obese Zucker rats before and after weaning.

This study was undertaken to determine the changes which could occur in glucose homeostatis during the suckling-weaning transition in the genetically obese Zucker (fa/fa) rat. Glucose kinetics and glucose utilization in individual tissues were determined in 15-day-old suckling and 30-day-old weaned obese Zucker rats either in the post-absorptive state or during a glucose infusion. During suckling, glucose turnover rates in the basal state as well as glucose production and utilization during the glucose infusion were identical in lean and obese rats. Furthermore, individual tissue glucose utilizations were similar in the two groups of rats, except in brown adipose tissue where utilization was lower in obese than in lean rats during the glucose challenge. After weaning, glucose turnover rates and glucose utilization in individual tissues were identical in the two genotypes in the basal state. During the glucose infusion, hepatic glucose production was less suppressed in the obese. Furthermore, glucose utilization was significantly lower in muscles (extensor digitorum longus, tibialis anterior, diaphragm) and higher in white adipose tissue of obese rats. These data show that, before weaning, pre-obese Zucker rats present a perturbation only in the uptake of glucose in brown adipose tissue. Major defects in the regulation of glucose homeostatis occur after weaning.

Effects of counterregulatory hormones on insulin-induced glucose utilization by individual tissues in rats.

The effect of counterregulatory hormones (epinephrine, norepinephrine and glucocorticoids) on insulin-induced glucose utilization in individual tissues of normal rats was investigated in vivo. This was done in normoglycaemic conditions, using the euglycaemic hyperinsulinaemic clamp combined with an injection of 2-[1-3H]-deoxyglucose. The main effect of these hormones was to reduce insulin-induced glucose utilization in skeletal muscles and particularly in the oxidative one. No changes were observed in heart diaphragm and adipose tissues. These results emphazise the role of counterregulatory hormones on glucose utilization and demonstrate that muscles are their major site of action. They support the notion that the increase in plasma concentrations of these hormones could play a role in states of insulin resistance like obesity and diabetes.

Inheritance of diabetes mellitus as consequence of gestational hyperglycemia in rats.

Our study investigated whether a deterioration of glucose homeostasis and insulin secretion in adult female rats from hyperglycemic dams could be transmitted to the next generation independent of genetic interferences. Dams (F0) were rendered hyperglycemic by continuous glucose infusion during the last week of pregnancy. Females born of these rats (F1) exhibited glucose intolerance and impaired insulin secretion in vivo at adulthood. When they were 3 mo old, they were matched with males born of control dams. During pregnancy, their glucose tolerance remained impaired compared with that of controls. Consequently, F2 newborns of F1 hyperglycemic dams showed the main features of newborns from diabetic mothers: they were hyperglycemic, hyperinsulinemic, and macrosomic. As adults, they displayed basal hyperglycemia and defective glucose tolerance and insulin secretion. This indicates that the long-range deteriorating effects on glucose homeostasis of gestational hyperglycemia in the F1 generation are transmitted to the F2 generation and suggests that a perturbed fetal metabolic environment contributes to the inheritance of diabetes mellitus.