Omapatrilat,an Angiotensin-Converting Enzyme and Neutral Endopeptidase Inhibitor,Attenuates Early Atherosclerosis in Diabetic and in Nondiabetic Low-Density Lipoprotein Receptor–Deficient Mice

Omapatrilat inhibits both angiotensin-converting enzyme (ACE) and neutral endopeptidase (NEP). ACE inhibitors have been shown to inhibit atherosclerosis in apoE-deficient mice and in several other animal models but failed in low-density lipoprotein (LDL) receptor– deficient mice despite effective inhibition of the reninangiotensin- aldosterone system. The aim of the present study was to examine the effect of omapatrilat on atherogenesis in diabetic and nondiabetic LDL receptor–deficient mice. LDL receptor–deficient male mice were randomly divided into 4 groups (n = 11 each). Diabetes was induced in 2 groups by low-dose STZ, the other 2 groups served as nondiabetic controls. Omapatrilat (70 mg/kg/day) was administered to one of the diabetic and to one of the nondiabetic groups. The diabetic and the nondiabetic mice were sacrificed after 3 and 5 weeks, respectively. The aortae were examined and the atherosclerotic plaque area was measured. The atherosclerotic plaque area was significantly smaller in the omapatrilat-treated mice, both diabetic and nondiabetic, as compared to nontreated controls. The mean plaque area of omapatrilattreated nondiabetic mice was 9357 ± 7293 μm², versus 71977 ± 34610 μm² in the nontreated mice (P = .002). In the diabetic animals, the plaque area was 8887 ± 5386 μm² and 23220 ± 10400 μm², respectively for treated and nontreated mice (P = .001). Plasma lipids were increased by omapatrilat: Meanplasma cholesterol in treated mice, diabetic and nondiabetic combined, was 39.31 ± 6.00 mmol/L, versus 33.12 ± 7.64 mmol/L in the nontreated animals (P = .008). The corresponding combined mean values of triglycerides were 4.83 ± 1.93 versus 3.00 ± 1.26 mmol/L (P = .02). Omapatrilat treatment did not affect weight or plasma glucose levels. Treatment with omapatrilat inhibits atherogenesis in diabetic as well as nondiabetic LDL receptor–deficient mice despite an increase in plasma lipids, suggesting a direct effect on the arterial wall.     

Changes in the Growth Hormone-IGF-I Axis in Non-obese Diabetic Mice

We investigated the changes in GH-IGF-I axis in non-obese diabetic (NOD)-mice, a model of insulin dependent diabetes mellitus. Diabetic female NOD mice and their age- and sex-matched controls were sacrificed at 4, 14, 21 and 30 days (30d DM) after the onset of glycosuria. Serum GH levels increased and serum IGF-I levels decreased in the 30d DM group (182 ± 32% and 45 ± 24% of age-matched controls respectively, p < 0.05). Another group (30d DM + I) was given SC insulin, and its serum IGF-I levels remained decreased. Liver GH receptor (GHR) and GH binding protein (GHBP) mRNA levels, as well as liver membrane GH binding assays were deeply decreased in the 30d DM group in comparison to controls. GHR message and binding capacity remained decreased in the 30d DM + I group. Renal GHR mRNA was decreased at 21d DM but not at 14d DM, whereas GHBP mRNA remained unchanged throughout the experiment. In conclusion, increased serum GH levels are documented in NOD diabetic mice, similarly to the changes described in humans. The decrease in GHR levels and decreased serum IGF-I in spite of increased circulating GH suggest a state of GH resistance.     

Octreotide,a Somatostatin Analogue,Fails to Inhibit Hypoxia-induced Retinal Neovascularization in the Neonatal Rat

Objective: Octreotide, a somatostatin analogue, has been shown to prevent angiogenesis in diverse in vitro models. We evaluated its effect on retinal neovascularization in vivo, using a neonatal rat retinopathy model. Methods: We used, on alternating days, hypoxia (10% O₂) and hyperoxia (50% O₂) during the first 14 days of neonatal rats, to induce retinal neovascularization. Half of the rats were injected subcutaneously with octreotide 0.7 μg/g BW twice daily. At day 18 the eyes were evaluated for the presence of epiretinal and vitreal hemorrhage, neovascularization and epiretinal proliferation. Octreotide pharmacokinetics and its effect on serum growth hormone (GH) and insulin-like growth factor I (IGF-I) were examined in 28 rats. Results: Serum octreotide levels were 667 μg/1 two hours after injection, 26.4 μg/1 after nine hours and 3.2 μg/1 after 14 hours. GH levels were decreased by 40% (p = 0.002) two hours after injection but thereafter returned to baseline. IGF-I levels were unchanged two hours after injection and were elevated by 26% 14 hours after injection (p = 0.02). Epiretinal membranes were highly associated with epiretinal hemorrhages (p < 0.001), while retinal neovascularization was notably associated with vitreal hemorrhages (p < 0.001). Conclusions: Twice-daily injections of octreotide failed to produce sustained decrease in serum GH, but produced rebound elevation of serum IGF-I. Accordingly, no statistically significant effect of injections on retinal pathology was noted. This finding, however, does not contradict our assumption that GH suppression may decrease the severity of retinopathy.     

GH administration and renal IGF-I system in arthritic rats

Experimental arthritis in rats results in a growth failure and a decrease in circulating and hepatic concentrations of insulin-like growth factor I (IGF-I). Renal damage has also been reported in arthritic rats. The aim of this study was 1) to analyse if alterations in the IGF-I system in the kidney occurs in adjuvant-induced arthritis and 2) to analyse if recombinant human GH (rhGH) administration is able to reverse these effects. Male Wistar rats were injected with complete Freund's adjuvant or vehicle and 22 days later they were killed. Arthritis increased serum creatinine levels, relative kidney weight and IGF-I concentrations in this organ. In a second experiment, arthritic and control rats received rhGH (3 UI/Kg sc) or 250 microl saline from day 14, after adjuvant or vehicle injection, until day 22. IGF-I concentrations were higher in both the renal cortex and medulla of arthritic rats. In contrast, kidney IGF-I mRNA was lower in both areas of arthritic animals. GH treatment significantly decreased serum creatinine levels and IGF-I concentrations in the kidney cortex and medulla of arthritic rats. However, the administration of rhGH to arthritic animals significantly increased the IGF-I gene expression in both the renal cortex and medulla. Serum and kidney concentrations of IGF-I binding proteins (IGFBPs) were increased in arthritic animals and they were reduced by GH administration. CONCLUSION: These data suggest that experimental arthritis causes renal dysfunction and GH treatment can ameliorate this effect.     

Growth hormone (GH)-independent stimulation of adiposity by GH secretagogues

Growth hormone secretagogues (GHSs) stimulate growth hormone (GH) secretion, which is lipolytic. Here we compared the effects of twice daily s.c. treatment of GH and the GHS, ipamorelin, on body fat in GH-deficient (lit/lit) and in GH-intact (+/lit and +/+) mice. In +/lit and lit/lit mice ipamorelin induced a small (15%) increase in body weight by 2 weeks, that was not further augmented by 9 weeks. GH treatment markedly enhanced body weight in both groups. Ipamorelin also increased fat pad weights relative to body weight in both lit/lit and +/lit mice. Two weeks GHS treatment (ipamorelin or GHRP-6) also increased relative body fat, quantified by in vivo dual energy X-ray absorpiometry (DEXA) in GH-intact mice. GH decreased relative fat mass in lit/lit mice and had no effect in GH-intact mice. Treatment with GHS, but not GH, increased serum leptin and food intake in GH-intact mice. Thus, GHSs increase body fat by GH-independent mechanisms that may include increased feeding.     

Preventive Effect of Dipeptidyl Peptidase-4 Inhibitor on Atherosclerosis Is Mainly Attributable to Incretin's Actions in Nondiabetic and Diabetic Apolipoprotein E-Null Mice

Aim

Several recent reports have revealed that dipeptidyl peptidase (DPP)-4 inhibitors have suppressive effects on atherosclerosis in apolipoprotein E-null (Apoe ^−/−) mice. It remains to be seen, however, whether this effect stems from increased levels of the two active incretins, glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP).

Methods

Nontreated Apoe ^−/− mice, streptozotocin-induced diabetic Apoe ^−/− mice, and db/db diabetic mice were administered the DPP-4 inhibitor vildagliptin in drinking water and co-infused with either saline, the GLP-1 receptor blocker, exendin(9–39), the GIP receptor blocker, (Pro³)GIP, or both via osmotic minipumps for 4 weeks. Aortic atherosclerosis and oxidized low-density lipoprotein-induced foam cell formation in exudate peritoneal macrophages were determined.

Results

Vildagliptin increased plasma GLP-1 and GIP levels without affecting food intake, body weight, blood pressure, or plasma lipid profile in any of the animals tested, though it reduced HbA1c in the diabetic mice. Diabetic Apoe ^−/− mice exhibited further-progressed atherosclerotic lesions and foam cell formation compared with nondiabetic counterparts. Nondiabetic and diabetic Apoe ^−/− mice showed a comparable response to vildagliptin, namely, remarkable suppression of atherosclerotic lesions with macrophage accumulation and foam cell formation in peritoneal macrophages. Exendin(9–39) or (Pro³)GIP partially attenuated the vildagliptin-induced suppression of atherosclerosis. The two blockers in combination abolished the anti-atherosclerotic effect of vildagliptin in nondiabetic mice but only partly attenuated it in diabetic mice. Vildagliptin suppressed macrophage foam cell formation in nondiabetic and diabetic mice, and this suppressive effect was abolished by infusions with exendin(9–39)+(Pro³)GIP. Incubation of DPP-4 or vildagliptin in vitro had no effect on macrophage foam cell formation.

Conclusions

Vildagliptin confers a substantial anti-atherosclerotic effect in both nondiabetic and diabetic mice, mainly via the action of the two incretins. However, the partial attenuation of atherosclerotic lesions by the dual incretin receptor antagonists in diabetic mice implies that vildagliptin confers a partial anti-atherogenic effect beyond that from the incretins.     

Effect of Mild Hypoinsulinemia on Renal Hypertrophy: Growth Hormone/Insulin-Like Growth Factor I System in Mild Streptozotocin Diabetes

The metabolic aberrations associated with diabetes mellitus profoundly alter the growth hormone/insulin-like growth factor I (GH/IGF-I) system. In severe experimental diabetes, serum IGF-I level is reduced, reflecting altered hepatic expression. On the other hand, increased levels of kidney IGF-I have been implicated in the development of diabetic kidney disease. This study aimed to examine the effect of mild experimental diabetes with hypoinsulinemia on both the systemic and renal GH/IGF-I systems in a low-dose streptozotocin (STZ)-induced diabetic rat. Diabetic animals with mild hypoinsulinemia developed renal hyperfiltration within 3 days of diabetes, whereas the renal size increased significantly only between 30 and 48 days of diabetes. Plasma GHlevels were unchanged during the entire course of the study, but a decrease in serum IGF-I, IGF-binding protein 3 (IGFBP-3), and IGF-binding protein 4 (IGFBP-4) occurred after 10, 30, and 48 days. Kidney IGF-I and IGF-binding protein 1 (IGFBP-1) mRNA expression increased after 10 and 30 days of diabetes. A significant increase in kidney IGFBP-1/2, IGFBP-3, and IGFBP-4 proteins was seen after 48 days of diabetes.Apositive correlations was found between renal growth and insulin/glucose ratio (r = .57), kidney IGF-I (r = .57), IGFBP-1 mRNA(r = .43), IGFBP-1/2 (r = .41), and IGFBP-4 levels (r = .40). These results demonstrate hyperfiltration within 3 days of diabetes and a similar response in the IGF-I system in mildly and severely hypoinsulinemic rats; however, renomegaly develops slower in mildly diabetic rats at least partly due to delayed changes in the renal IGF and IGF BPs.     

Validation of serum IGF-I as a biomarker to monitor the bioactivity of exogenous growth hormone agonists and antagonists in rabbits

The development of new growth hormone (GH) agonists and growth hormone antagonists (GHAs) requires animal models for pre-clinical testing. Ideally, the effects of treatment are monitored using the same pharmacodynamic marker that is later used in clinical practice. However, intact rodents are of limited value for this purpose because serum IGF-I, the most sensitive pharmacodynamic marker for the action of GH in humans, shows no response to treatment with recombinant human GH and there is little evidence for the effects of GHAs, except when administered at very high doses or when overexpressed. As an alternative, more suitable model, we explored pharmacodynamic markers of GH action in intact rabbits. We performed the first validation of an IGF-I assay for the analysis of rabbit serum and tested precision, sensitivity, linearity and recovery using an automated human IGF-I assay (IDS-iSYS). Furthermore, IGF-I was measured in rabbits of different strains, age groups and sexes, and we monitored IGF-I response to treatment with recombinant human GH or the GHA Pegvisomant. For a subset of samples, we used LC-MS/MS to measure IGF-I, and quantitative western ligand blot to analyze IGF-binding proteins (IGFBPs). Although recovery of recombinant rabbit IGF-I was only 50% in the human IGF-I assay, our results show that the sensitivity, precision (1.7–3.3% coefficient of variation) and linearity (90.4–105.6%) were excellent in rabbit samples. As expected, sex, age and genetic background were major determinants of IGF-I concentration in rabbits. IGF-I and IGFBP-2 levels increased after single and multiple injections of recombinant human GH (IGF-I: 286±22 versus 434±26 ng/ml; P<0.01) and were highly correlated (P<0.0001). Treatment with the GHA lowered IGF-I levels from the fourth injection onwards (P<0.01). In summary, we demonstrated that the IDS-iSYS IGF-I immunoassay can be used in rabbits. Similar to rodents, rabbits display variations in IGF-I depending on sex, age and genetic background. Unlike in rodents, the IGF-I response to treatment with recombinant human GH or a GHA closely mimics the pharmacodynamics seen in humans, suggesting that rabbits are a suitable new model to test human GH agonists and antagonists.KEY WORDS: Pharmacodynamic marker, Acromegaly, Growth hormone deficiency, Animal model     

Circulating microRNA profile in humans and mice with congenital GH deficiency

Reduced inflammation, increased insulin sensitivity, and protection against cancer are shared between humans and mice with GH/IGF1 deficiency. Beyond hormone levels, miRNAs are important regulators of metabolic changes associated with healthy aging. We hypothesized that GH deficiency in humans alters the abundance of circulating miRNAs and that a subset of those miRNAs may overlap with those found in GH‐deficient mice. In this study, subjects with untreated congenital isolated GH deficiency (IGHD; n = 23) and control subjects matched by age and sex (n = 23) were recruited and serum was collected for miRNA sequencing. Serum miRNAs from young (6 month) and old (22 month) Ames dwarf (df/df) mice with GH deficiency and their WT littermates (n = 5/age/genotype group) were used for comparison. We observed 14 miRNAs regulated with a genotype by age effect and 19 miRNAs regulated with a genotype effect independent of age in serum of IGHD subjects. These regulated miRNAs are known for targeting pathways associated with longevity such as mTOR, insulin signaling, and FoxO. The aging function was overrepresented in IGHD individuals, mediated by hsa‐miR‐31, hsa‐miR‐146b, hsa‐miR‐30e, hsa‐miR‐100, hsa‐miR‐181b‐2, hsa‐miR‐195, and hsa‐miR‐181b‐1, which target the FoxO and mTOR pathways. Intriguingly, miR‐181b‐5p, miR‐361‐3p, miR‐144‐3p, and miR‐155‐5p were commonly regulated in the serum of humans and GH‐deficient mice. In vitro assays confirmed target genes for the main up‐regulated miRNAs, suggesting miRNAs regulated in IGHD individuals can regulate the expression of age‐related genes. These findings indicate that systemic miRNAs regulated in IGHD individuals target pathways involved in aging in both humans and mice.     

Prostaglandin E2 (PGE2)-induced growth hormone (GH) release: Effect of intrahypothalamic and intrapituitary implants

Overiectomized rats were unilaterally implanted with a 23-gauge stainless steel cannula in different hypothalamic areas or in the pituitary gland and subsequently were treated with estrogen (sc, 10 μg estradiol benzoate, Eb). Two days after the estrogen injection, an inner cannula containing PGE₂ or PHF_2α at its tip was inserted into the cannula. Other animals were implanted with empty inner cannula. Plasma GH concentrations were measured by RIA in blood samples drawn from the jugular vein while the animals were lightly etherized before (−2) and at 20, 40, 60 and 120 min following the implantation. Plasma GH levels in control animals bearing an empty cannula in the body of the arcuate nucleus-median eminence region (BARH-ME) were signifantly depressed by the ether stress. The implantation of PGF_2α in this area was completely ineffective in preventing ether stress-induced decline in plasma GH. By contrast, PGE₂ implanted in BARH-ME or the post-chiasmatic region of the hypothalamus (HARH-ME) elevated plasma GH 20 min following its implantation and partially prevented the subsequent decrease in GH levels induced by ether stress. PGE₂ implants located in several other hypothalamic areas failed to induce GH release or to prevent the decline in GH levels induced by ether stress. However, PGE₂ implanted in the pituitary gland elicited a marked increase in plasma GH at 20 min and completely prevented the subsequent ether stress-induced decline in GH levels.The results suggest that PGE₂ can act at both hypothalamic (ARH-ME) and pituitary levels to stimulate GH release. At the hypothalamus, PGE₂ may inhibit GH-inhibiting factor (GIF) release or induce release of GH releasing factor (GHRF).     

Effect of Resveratrol on Behavioral Performance of Streptozotocin-induced Diabetic Mice inAnxiety Tests

The aim of this study was to evaluate with anxiety tests the effect of resveratrol (RSV) on streptozotocin (STZ)-induced diabetic mouse behavioral performance at the second and fourth week of treatment. Confirmed diabetic mice (>250 mg/dl of glucose in blood after STZ injection) were treated with RSV (RDM, n=12) or control treated (DM, n=12) for 4 weeks. DM and RDM were tested in the Open Field Test (OFT) and Elevated Plus Maze (EPM). In the second week of RSV treatment, a higher grooming frequency (P<0.05) and a lower defecation and rearing frequency (P<0.05) were detected in the OFT in the RDM group compared with the DM. There was a higher grooming frequency (P<0.05) and higher percentage of entries in open arms (P<0.05) in the RDM group than in the DM group in the EPM. However, in the fourth week of RSV treatment, the only effect observed was a higher grooming frequency in the RDM group than in the DM group (P<0.05) in the EPM. In conclusion, RSV treatment in diabetic mice provoked anxiolytic-like effects in both tests (OFT and EPM), and these effects were observed in a short time window (2 weeks). It is suggested that RSV may help diabetic animals to adapt to new stressing and anxiety situations and thus to improve their welfare.     

Diabetes reduces the cholesterol exporter ABCA1 in mouse macrophages and kidneys

Accumulation of cholesterol in arterial macrophages may contribute to diabetes-accelerated atherosclerotic cardiovascular disease. The ATP-binding cassette transporter ABCA1 is a cardioprotective membrane protein that mediates cholesterol export from macrophages. Factors elevated in diabetes, such as reactive carbonyls and free fatty acids, destabilize ABCA1 protein in cultured macrophages, raising the possibility that impaired ABCA1 plays an atherogenic role in diabetes. We therefore examined the modulation of ABCA1 in two mouse models of diabetes. We isolated peritoneal macrophages, livers, kidneys, and brains from type 1 non-obese diabetic (NOD) mice and mice made diabetic by viral-induced autoimmune destruction of pancreatic β-cells, and we measured ABCA1 protein and mRNA levels and cholesterol contents. ABCA1 protein levels and cholesterol export activity were reduced by 40–44% (P < 0.01) in peritoneal macrophages and protein levels by 48% (P < 0.001) in kidneys in diabetic NOD mice compared with nondiabetic animals, even though ABCA1 mRNA levels were not significantly different. A similar selective reduction in ABCA1 protein was found in peritoneal macrophages (33%, P < 0.05) and kidneys (35%, P < 0.05) from the viral-induced diabetic mice. In liver and brain, however, diabetes had no effect or slightly increased ABCA1 protein and mRNA levels. The reduced ABCA1 in macrophages and kidneys was associated with increased cholesterol content. Impaired ABCA1-mediated cholesterol export could therefore contribute to the increased atherosclerosis and nephropathy associated with diabetes.     

Effect of maternal diabetes and ethanol interactions on embryo development in the mouse

The aim of this study was to determine the possible fetal effects of interaction between maternal diabetes and acute doses of alcohol. Pregnant TO mice were made diabetic by a single injection of streptozotocin (STZ) on gestation day (GD) 2. Single dose of 0.003 or 0.03 ml/g body weight of fresh ethanol (25% v/v of absolute alcohol in normal saline) was injected into groups of diabetic and nondiabetic animals on GD 7 or 8. One group of diabetic animals had a daily dose of 6–8 IU of insulin subcutaneously. Fetuses were collected on GD 18. There was a significant increase in the incidence of implantation failure in the diabetes plus ethanol groups and insulin control group. Ethanol injection on GD 7 accentuated diabetes-related embryonic resorption and intrauterine growth retardation (IUGR). This effect was less marked in the diabetic group treated with ethanol on GD 8. Diabetes alone produced a greater incidence of IUGR than ethanol alone. Midfacial hypoplasia and minor anomalies were found more frequently in the combination treatment groups. Holoprosencephaly and thymus hypoplasia observed in diabetic groups were found to be reduced in frequency in the diabetes plus ethanol groups, suggesting an antagonistic type of ethanol--diabetes interaction, stage-dependently. Since severely malformed embryos are known to be resorbed/killed in utero in mice, this reduction might reflect the magnitude of early death of severely malformed embryos. These data suggest that the interaction effects are possibly related to alterations in fundamental developmental processes of early embryos. (Mol Cell Biochem 261: 43–56, 2004)     

Differential expression of insulin-like growth factor-I and insulin-like growth factor binding protein-1 in the diabetic rat

Summary Multiple factors contribute to the growth retardation which is a characteristic feature of uncontrolled diabetes. In this report we have examined the effects of streptozotocin-induced (STZ) diabetes on expression of insulin-like growth factor-I (IGF-I) and insulin-like growth factor binding protein-1 (IGFBP-1) in various tissues. As early as 7 days after STZ administration there was a modest reduction in IGF-I mRNA abundance. The reduction (10–30%) was of similar magnitude in each of the 7 tissues examined; liver, kidney, lung, diaphragm, quadraceps, heart and adipose tissue. However, the reduction achieved statistical significance only in the lung (p < 0.05) and diaphragm (p < 0.01). A further reduction in IGF-I mRNA abundance was seen in many tissues, 32 and 91 days after STZ administration. In contrast to the decrease in IGF-I mRNA, IGFBP-1 mRNA was significantly increased in the liver and kidney of diabetic rats. IGFBP-1 mRNA was detectable at only very low levels in other tissues but was increased in diabetic rats compared non-diabetic rats. In diabetic rats, a highly significant correlation (R = 0.75, p < 0.001) between hepatic IGFBP-1 mRNA and glucose was observed whereas there was no significant correlation between serum glucose and hepatic IGF-I mRNA abundance (R = 0.24, p = NS). Treatment of diabetic rats with insulin resulted in a small, non significant increase in hepatic and renal IGF-I mRNA and a significant decrease in renal IGFBP-1 mRNA abundance. The observations reported here are consistent with the hypothesis that diminished IGF-I expression and inhibition of available IGF-1 by increased levels of IGFBP-1 may explain the impaired growth seen in diabetic animals.     

Streptozotocin Stimulates the Ion Channel TRPA1 Directly: INVOLVEMENT OF PEROXYNITRITE*

Streptozotocin (STZ)-induced diabetes is the most commonly used animal model of diabetes. Here, we have demonstrated that intraplantar injections of low dose STZ evoked acute polymodal hypersensitivities in mice. These hypersensitivities were inhibited by a TRPA1 antagonist and were absent in TRPA1-null mice. In wild type mice, systemic STZ treatment (180 mg/kg) evoked a loss of cold and mechanical sensitivity within an hour of injection, which lasted for at least 10 days. In contrast, Trpa1^−/− mice developed mechanical, cold, and heat hypersensitivity 24 h after STZ. The TRPA1-dependent sensory loss produced by STZ occurs before the onset of diabetes and may thus not be readily distinguished from the similar sensory abnormalities produced by the ensuing diabetic neuropathy. In vitro, STZ activated TRPA1 in isolated sensory neurons, TRPA1 cell lines, and membrane patches. Mass spectrometry studies revealed that STZ oxidizes TRPA1 cysteines to disulfides and sulfenic acids. Furthermore, incubation of tyrosine with STZ resulted in formation of dityrosine, suggesting formation of peroxynitrite. Functional analysis of TRPA1 mutants showed that cysteine residues that were oxidized by STZ were important for TRPA1 responsiveness to STZ. Our results have identified oxidation of TRPA1 cysteine residues, most likely by peroxynitrite, as a novel mechanism of action of STZ. Direct stimulation of TRPA1 complicates the interpretation of results from STZ models of diabetic sensory neuropathy and strongly argues that more refined models of diabetic neuropathy should replace the use of STZ.     

Chronic alcohol consumption,type 2 diabetes mellitus,insulin-like growth factor-I (IGF-I), and growth hormone (GH) in ethanol-treated diabetic rats

Aims

Alcohol has deleterious influences on glucose metabolism which may contribute to the development of type 2 diabetes mellitus (T2DM). Insulin-like growth factor I (IGF-I) and growth hormone (GH), which interact with insulin to modulate metabolic control, have been shown to be related to impaired glucose tolerance. This study was conducted to assess the possibility that altered circulating IGF-I and GH levels contribute to the exacerbation of T2DM by alcohol use in type 2 diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats and non-diabetic Long-Evans Tokushima Otsuka (LETO) rats.

Main method

OLETF rats were pair-fed a Lieber-DeCarli Regular Ethanol diet and LETO rats were pair-fed a control diet for 6 weeks. At 6 weeks, an Intraperitoneal Glucose Tolerance Test (IP-GTT) was performed and IGF-I and GH levels were evaluated.

Key findings

Prior to an IP-GTT, OLETF-Ethanol (O-E) group had significantly a decrease in the mean glucose levels compared to OLETF-Control (O-C) group. At 120 min post IP-GTT, the O-E group had significantly an increase in the mean glucose levels compared to O-C group. The serum IGF-I levels were significantly lower and the serum GH levels were significantly higher in the O-E group than in L-C group.

Significance

These results suggest that IGF-I and GH are prominent in defining the risk and development of T2DM, and may be adversely affected by heavy alcohol use, possibly mediating its diabetogenic effects. Thus, the overall glucose intolerance in the setting of alcoholism may be attributable to inappropriate alteration of IGF-I and GH levels.     

Kidney growth in normal and diabetic mice is not affected by human insulin-like growth factor binding protein-1 administration

Insulin-like growth factor I (IGF-I) accumulates in the kidney following the onset of diabetes, initiating diabetic renal hypertrophy. Increased renal IGF-I protein content, which is not reflected in messenger RNA (mRNA) levels, suggests that renal IGF-I accumulation is due to sequestration of circulating IGF-I rather than to local synthesis. It has been suggested that IGF-I is trapped in the kidney by IGF binding protein 1 (IGFBP-1). We administered purified human IGFBP-1 (hIGFBP-1) to nondiabetic and diabetic mice as three daily sc injections for 14 days, starting 6 days after induction of streptozotocin diabetes when the animals were overtly diabetic. Markers of early diabetic renal changes (i.e., increased kidney weight, glomerular volume, and albuminuria) coincided with accumulation of renal cortical IGF-I despite decreased mRNA levels in 20-day diabetic mice. Human IGFBP-1 administration had no effect on increased kidney weight or albuminuria in early diabetes, although it abolished renal cortical IGF-I accumulation and glomerular hypertrophy in diabetic mice. Increased IGF-I levels in kidneys of normal mice receiving hIGFBP-1 were not reflected on kidney parameters. IGFBP-1 administration in diabetic mice had only minor effects on diabetic renal changes. Accordingly, these results did not support the hypothesis that IGFBP-1 plays a major role in early renal changes in diabetes.     

Glucose regulates ghrelin, neuropeptide Y, and the GH/IGF-I axis in the tilapia, Oreochromis mossambicus

In general, a fish's ability to clear glucose is sluggish in relation to mammals, which has lead to the idea that fish are glucose intolerant. It has been reported that circulating glucose levels do fluctuate in response to environmental challenges. Recent reports suggest that glucose may function as a metabolic signal regulating ‘glucosensors’ in the brain in fish, as has been reported in mammals. The current study was designed to investigate the effect of glucose on ghrelin and neuropeptide Y (NPY) signaling in the brain, and on the growth hormone/insulin-like growth factor-I (GH/IGF-I) in the tilapia, Oreochromis mossambicus. Glucose treatment significantly increased plasma and stomach mRNA levels of ghrelin. In the brain, mRNA levels of the ghrelin receptor (GRLN-R) were significantly reduced, whereas NPY mRNA levels were significantly elevated; suggesting that NPY containing neurons may be a “glucosensor” as reported in mammals. Glucose treatment resulted in changes in the GH/IGF-I axis. Liver mRNA levels of both GH receptors (GHR1 and GHR2) were significantly elevated, whereas liver IGF-I mRNA were unaltered by glucose treatment. No change in plasma or pituitary mRNA levels of GH was observed. Glucose significantly reduced plasma IGF-I levels. These data show that glucose regulates endocrine factors involved in appetite, growth, and possibly energy homeostasis, and suggests that glucose may be acting as a signal of metabolic status in fish.     

Increased plasma corticosterone levels in bovine growth hormone (bGH) transgenic mice: Effects of ACTH,GH and IGF-I onin vitro adrenal corticosterone production

Previous work from our laboratory provided evidence for increased plasma corticosterone levels in mice transgenic for human and bovine growth hormone (GH). Corticosterone was elevated in both sexes, under both basal and ether-induced stress conditions. The objectives of the present study were to investigate thein vitro adrenal sensitivity to ACTH, GH and/or IGF-I in normal and bGH transgenic mice, to examine plasma corticosterone levels at different times of the day, and to determine plasma levels of ACTH in these animals. For the measurement of plasma corticosterone and ACTH levels, transgenic and normal siblings were housed 2 per cage and decapitated simultaneously within 20 seconds of the first disturbance of the cage. The corticosterone production byin vitro adrenal incubations did not differ between adrenals from normal and transgenic mice at the basal level or in the presence of different doses of ACTH. Growth hormone or IGF-I did not have any effect on corticosterone productionin vitro when given alone, and did not modify the effects of ACTH on the accumulation of corticosterone in the media. Plasma corticosterone concentrations were higher in transgenic than in normal animals in both morning and evening. Plasma concentrations of ACTH in animals killed in the morning were sharply increased in transgenic males as compared with their normal siblings. The results indicate that increased circulating levels of corticosterone in transgenic mice are not due to a potentiation of ACTH actions by GH or IGF-I, but rather to a chronic increase in plasma ACTH levels. The increase in ACTH is presumably a reflection of GH actions in the hypothalamic-pituitary system.     

Pentoxifylline Diminishes the Oxidative Damage to Renal Tissue Induced by Streptozotocin in the Rat

Oxidative damage has been suggested to be a contributing factor in the development to diabetic nephropathy (DN). Recently, there has been evidence that pentoxifylline (PTX) has free radical-scavenging properties; thus, its antiinflammatory and renoprotective effects may be related to a reduction in reactive oxygen species production. It is likely that the pharmacological effects of PTX include an antioxidant mechanism as shown in in vitro assays. The aim of this study was to evaluate whether the reported renoprotective effects of PTX could be the result of its antioxidant actions in streptozotocin (STZ)-induced DN in rats. The administration of PTX over a period of 8 weeks, in addition to displaying renoprotective effects, caused a significant reduction in lipoperoxide levels (LPOS) in the diabetic kidney (P < 0.05), compared to untreated rats. These levels were comparable to those in the healthy kidney of experimental animals (P > 0.05). All untreated STZ rats exhibited an increase in LPOS as opposed to healthy controls (H) (P < 0.001). The total antioxidant activity (TAA) in plasma was increased significantly already after 2 days of STZ (P < 0.05). When we examined the progression of TAA in STZ rats, there was a significant decrease over 8 weeks (P < 0.05). PTX treatment caused an increase in TAA when compared to untreated STZ rats (P < 0.05). Renal hypertrophy was less evident in PTX-treated STZ than in untreated STZ rats, evaluated by kidney weight/body weight ratio. These results indicate that PTX decreases the oxidative damage induced by these experimental procedures and may increase antioxidant defense mechanisms in STZ-induced diabetes in rats.