Melatonin induces browning of inguinal white adipose tissue in Zucker diabetic fatty rats

Melatonin limits obesity in rodents without affecting food intake and activity, suggesting a thermogenic effect. Identification of brown fat (beige/brite) in white adipose tissue (WAT) prompted us to investigate whether melatonin is a brown‐fat inducer. We used Zücker diabetic fatty (ZDF) rats, a model of obesity‐related type 2 diabetes and a strain in which melatonin reduces obesity and improves their metabolic profiles. At 5 wk of age, ZDF rats and lean littermates (ZL) were subdivided into two groups, each composed of four rats: control and those treated with oral melatonin in the drinking water (10 mg/kg/day) for 6 wk. Melatonin induced browning of inguinal WAT in both ZDF and ZL rats. Hematoxylin–eosin staining showed patches of brown‐like adipocytes in inguinal WAT in ZDF rats and also increased the amounts in ZL animals. Inguinal skin temperature was similar in untreated lean and obese rats. Melatonin increased inguinal temperature by 1.36 ± 0.02°C in ZL and by 0.55 ± 0.04°C in ZDF rats and sensitized the thermogenic effect of acute cold exposure in both groups. Melatonin increased the amounts of thermogenic proteins, uncoupling protein 1 (UCP1) (by ~2‐fold, P < 0.01) and PGC‐1α (by 25%, P < 0.05) in extracts from beige inguinal areas in ZL rats. Melatonin also induced measurable amounts of UCP1 and stimulated by ~2‐fold the levels of PGC‐1α in ZDF animals. Locomotor activity and circulating irisin levels were not affected by melatonin. These results demonstrate that chronic oral melatonin drives WAT into a brown‐fat‐like function in ZDF rats. This may contribute to melatonin′s control of body weight and its metabolic benefits.     

Melatonin improves mitochondrial function in inguinal white adipose tissue of Zücker diabetic fatty rats

Mitochondrial dysfunction in adipose tissue may contribute to obesity‐related metabolic derangements such as type 2 diabetes mellitus (T2DM). Because mitochondria are a target for melatonin action, the goal of this study was to investigate the effects of melatonin on mitochondrial function in white (WAT) and beige inguinal adipose tissue of Zücker diabetic fatty (ZDF) rats, a model of obesity‐related T2DM. In this experimental model, melatonin reduces obesity and improves the metabolic profile. At 6 wk of age, ZDF rats and lean littermates (ZL) were subdivided into two groups, each composed of four rats: control (C‐ZDF and C‐ZL) and treated with oral melatonin in the drinking water (10 mg/kg/day) for 6 wk (M‐ZDF and M‐ZL). After the treatment period, animals were sacrificed, tissues dissected, and mitochondrial function assessed in isolated organelles. Melatonin increased the respiratory control ratio (RCR) in mitochondria from white fat of both lean (by 26.5%, P < 0.01) and obese (by 34.5%, P < 0.01) rats mainly through a reduction of proton leaking component of respiration (state 4) (28% decrease in ZL, P < 0.01 and 35% in ZDF, P < 0.01). However, melatonin treatment lowered the RCR in beige mitochondria of both lean (by 7%, P < 0.05) and obese (by 13%, P < 0.05) rats by maintaining high rates of uncoupled respiration. Melatonin also lowered mitochondrial oxidative status by reducing nitrite levels and by increasing superoxide dismutase activity. Moreover, melatonin treatment also caused a profound inhibition of Ca‐induced opening of mPTP in isolated mitochondria from both types of fat, white and beige, in both lean and obese rats. These results demonstrate that chronic oral melatonin improves mitochondrial respiration and reduces the oxidative status and susceptibility to apoptosis in white and beige adipocytes. These melatonin effects help to prevent mitochondrial dysfunction and thereby to improve obesity‐related metabolic disorders such as diabetes and dyslipidemia of ZDF rats.     

Melatonin reduces hepatic mitochondrial dysfunction in diabetic obese rats

Hepatic mitochondrial dysfunction is thought to play a role in the development of liver steatosis and insulin resistance, which are both common characteristics of obesity and type 2 diabetes mellitus (T2DM). It was hypothesized that the antioxidant properties of melatonin could potentially improve the impaired functions of hepatic mitochondria in diabetic obese animals. Male Zucker diabetic fatty (ZDF) rats and lean littermates (ZL) were given either melatonin (10 mg/kg BW/day) orally for 6 wk (M‐ZDF and M‐ZL) or vehicle as control groups (C‐ZDF and C‐ZL). Hepatic function was evaluated by measurement of serum alanine transaminase and aspartate transaminase levels, liver histopathology and electron microscopy, and hepatic mitochondrial functions. Several impaired functions of hepatic mitochondria were observed in C‐ZDF in comparison with C‐ZL rats. Melatonin treatment to ZDF rats decreases serum levels of ALT (P < 0.001), alleviates liver steatosis and vacuolation, and also mitigates diabetic‐induced mitochondrial abnormalities, glycogen, and lipid accumulation. Melatonin improves mitochondrial dysfunction in M‐ZDF rats by increasing activities of mitochondrial citrate synthase (P < 0.001) and complex IV of electron transfer chain (P < 0.05) and enhances state 3 respiration (P < 0.001), respiratory control index (RCR) (P < 0.01), and phosphorylation coefficient (ADP/O ratio) (P < 0.05). Also melatonin augments ATP production (P < 0.05) and diminishes uncoupling protein 2 levels (P < 0.001). These results demonstrate that chronic oral melatonin reduces liver steatosis and mitochondria dysfunction in ZDF rats. Therefore, it may be beneficial in the treatment of diabesity.     

Melatonin increases brown adipose tissue mass and function in Zücker diabetic fatty rats: implications for obesity control

Melatonin limits obesity in rodents without affecting food intake and activity, suggesting a thermogenic effect. Previously we demonstrated that melatonin browns subcutaneous fat in Zücker diabetic fatty (ZDF) rats. Other works pointed to melatonin as a signal that increases brown adipose tissue (BAT) mass and function in rodents. However, direct proof of thermogenic properties (uncoupled mitochondria) of the newly recruited BAT in response to melatonin is still lacking. Therefore, in this work, we investigated if melatonin recruits thermogenic BAT in ZDF rats. Zücker lean (ZL) and ZDF animals were subdivided into two groups, control (C) and treated with oral melatonin (M) for 6 weeks. Mitochondrial mass, activity of citrate synthase (CS), and respiratory chain complexes I and IV were lower in C‐ZDF than in C‐ZL animals (P < .001). Melatonin treatment increased BAT weight in ZDF rats (P < .001). Also, it rose mitochondrial mass (P < .01) and activities of CS and complexes I and IV (P < .001) in both, ZDF and ZL rats. Uncoupling protein 1 (UCP1) mRNA and protein were 50% lower in BAT from obese rats. Also, guanosine diphosphate (GDP) binding was lower in ZDF than in lean rats (P < .01). Melatonin treatment of obese rats restored the expression of UCP1 and GDP binding to levels of lean rats and sensitized the thermogenic response to cold exposure. These data demonstrated that melatonin recruits thermogenic BAT in ZDF rats. This may contribute to melatonin's control of body weight and its metabolic benefits.     

Melatonin ameliorates low‐grade inflammation and oxidative stress in young Zucker diabetic fatty rats

The aim of this study was to investigate the effects of melatonin on low‐grade inflammation and oxidative stress in young male Zucker diabetic fatty (ZDF) rats, an experimental model of metabolic syndrome and type 2 diabetes mellitus (T2DM). ZDF rats (n = 30) and lean littermates (ZL) (n = 30) were used. At 6 wk of age, both lean and fatty animals were subdivided into three groups, each composed of 10 rats: naive (N), vehicle treated (V), and melatonin treated (M) (10 mg/kg/day) for 6 wk. Vehicle and melatonin were added to the drinking water. Pro‐inflammatory state was evaluated by plasma levels of interleukin‐6 (IL‐6), tumor necrosis factor‐α (TNF‐α), and C‐reactive protein (CRP). Also, oxidative stress was assessed by plasma lipid peroxidation (LPO), both basal and after Fe²⁺/H₂O₂ inducement. ZDF rats exhibited higher levels of IL‐6 (112.4 ± 1.5 pg/mL), TNF‐α (11.0 ± 0.1 pg/mL) and CRP (828 ± 16.0 µg/mL) compared with lean rats (IL‐6, 89.9 ± 1.0, P < 0.01; TNF‐α, 9.7 ± 0.4, P < 0.01; CRP, 508 ± 21.5, P < 0.001). Melatonin lowered IL‐6 (10%, P < 0.05), TNF‐α (10%, P < 0.05), and CRP (21%, P < 0.01). Basal and Fe²⁺/H₂O₂‐induced LPO, expressed as malondialdehyde equivalents (µmol/L), were higher in ZDF rats (basal, 3.2 ± 0.1 versus 2.5 ± 0.1 in ZL, P < 0.01; Fe²⁺/H₂O₂‐induced, 8.7 ± 0.2 versus 5.5 ± 0.3 in ZL; P < 0.001). Melatonin improved basal LPO (15%, P < 0.05) in ZDF rats, and Fe²⁺/H₂O₂‐ induced LPO in both ZL (15.2%, P < 0.01) and ZDF rats (39%, P < 0.001). These results demonstrated that oral melatonin administration ameliorates the pro‐inflammatory state and oxidative stress, which underlie the development of insulin resistance and their consequences, metabolic syndrome, diabetes, and cardiovascular disease.     

Melatonin improves glucose homeostasis in young Zucker diabetic fatty rats

The aim of this study was to investigate the effects of melatonin on glucose homeostasis in young male Zucker diabetic fatty (ZDF) rats, an experimental model of metabolic syndrome and type 2 diabetes mellitus (T2DM). ZDF rats (n=30) and lean littermates (ZL) (n=30) were used. At 6wk of age, both lean and fatty animals were subdivided into three groups, each composed of ten rats: naive (N), vehicle treated (V), and melatonin treated (M) (10mg/kg/day) for 6wk. Vehicle and melatonin were added to the drinking water. ZDF rats developed DM (fasting hyperglycemia, 460±39.8mg/dL; HbA(1) c 8.3±0.5%) with both insulin resistance (HOMA-IR 9.28±0.9 versus 1.2±0.1 in ZL) and decreased β-cell function (HOMA1-%B) by 75%, compared with ZL rats. Melatonin reduced fasting hyperglycemia by 18.6% (P<0.05) and HbA(1) c by 11% (P<0.05) in ZDF rats. Also, melatonin lowered insulinemia by 15.9% (P<0.05) and HOMA-IR by 31% (P<0.01) and increased HOMA1-%B by 14.4% (P<0.05). In addition, melatonin decreased hyperleptinemia by 34% (P<0.001) and raised hypoadiponectinemia by 40% (P<0.001) in ZDF rats. Moreover, melatonin reduced serum free fatty acid levels by 13.5% (P<0.05). These data demonstrate that oral melatonin administration ameliorates glucose homeostasis in young ZDF rats by improving both insulin action and β-cell function. These observations have implications on melatonin's possible use as a new pharmacologic therapy for improving glucose homeostasis and of obesity-related T2DM, in young subjects.     

Melatonin nephroprotective action in Zucker diabetic fatty rats involves its inhibitory effect on NADPH oxidase

Excessive activity of NADPH oxidase (Nox) is considered to be of importance for the progress of diabetic nephropathy. The aim of the study was to elucidate the effect of melatonin, known for its nephroprotective properties, on Nox activity under diabetic conditions. The experiments were performed on three groups of animals: (i) untreated lean (?/+) Zucker diabetic fatty (ZDF) rats; (ii) untreated obese diabetic (fa/fa) ZDF rats; and (iii) ZDF fa/fa rats treated with melatonin (20 mg/L) in drinking water. Urinary albumin excretion was measured weekly. After 4 wk of the treatment, the following parameters were determined in kidney cortex: Nox activity, expression of subunits of the enzyme, their phosphorylation and subcellular distribution. Histological studies were also performed. Compared to ?/+ controls, ZDF fa/fa rats exhibited increased renal Nox activity, augmented expression of Nox4 and p47^phox subunits, elevated level of p47^phox phosphorylation, and enlarged phospho‐p47^phox and p67^phox content in membrane. Melatonin administration to ZDF fa/fa rats resulted in the improvement of renal functions, as manifested by considerable attenuation of albuminuria and some amelioration of structural abnormalities. The treatment turned out to nearly normalize Nox activity, which was accompanied by considerably lowered expression and diminished membrane distribution of regulatory subunits, that is, phospho‐p47^phox and p67^phox. Thus, it is concluded that: (i) melatonin beneficial action against diabetic nephropathy involves attenuation of the excessive activity of Nox; and (ii) the mechanism of melatonin inhibitory effect on Nox is based on the mitigation of expression and membrane translocation of its regulatory subunits.     

Thermal, but not mechanical, nociceptive behavior is altered in the Zucker Diabetic Fatty rat and is independent of glycemic status.

This study investigated the possible link between developing hyperglycemia and mechanical and/or thermal hyperalgesia in the Zucker Diabetic Fatty (ZDF) rat. When normoglycemic (nonfasting blood glucose levels of 6 mM), 6-week-old ZDF rats were glucose intolerant compared to the nondiabetic Zucker lean control (ZL) rats, but there was no difference in their response to a noxious mechanical (paw pressure test) or thermal (hot plate) stimulus (mechanical nociceptive thresholds: ZDF 176.7+/-14.4 g, ZL 161.7+/-13.3 g; latencies to response to the thermal stimulus: ZDF 13.1+/-1.6 sec, ZL 16.7+/-1.5 sec). Blood glucose levels in untreated ZDF rats increased to 28.4+/-2.9 mM by 20 weeks of age, while ZDF rats treated with the insulin sensitizer, rosiglitazone, and ZL rats remained normoglycemic (< or =8 mM) throughout the study. Hyperglycaemia in ZDF rats was not associated with mechanical hyperalgesia, as the nociceptive threshold remained constant in both the rosiglitazone-treated and untreated ZDF rats and in the ZL rats throughout the study. In contrast, the latency to response to the thermal stimulus increased with time in ZL rats, but remained constant in hyperglycaemic ZDF rats such that the difference reached significance by 9 weeks of age (ZDF 11.6+/-1.7 sec, ZL 21.8+/-2.7 sec, p<0.01) and is consistent with hyperalgesia in the ZDF phenotype. However, this difference was not moderated by maintaining normoglycaemia in rosiglitazone-treated ZDF rats (12.8+/-1.3 sec). Together, the data suggest that hyperglycemia does not play a central role in the development of hyperalgesia in the ZDF rat.     

Effect of nutritional counseling on low-density lipoprotein cholesterol among Thai HIV-infected adults receiving antiretroviral therapy

HIV-infected patients receiving antiretroviral therapy have increased risk of metabolic syndrome, including dyslipidemia. In this study, we determined whether individual nutritional counseling reduced dyslipidemia, particularly low-density lipoprotein (LDL) cholesterol, among HIV-infected patients with dyslipidemia not currently taking lipid-lowering medication. We conducted a randomized 24-week trial among HIV-infected patients with dyslipidemia who were on antiretroviral therapy and were eligible to initiate therapeutic lifestyle changes according to the Thai National Cholesterol Education Program. Participants were randomly assigned to an intervention group that received individual counseling with a nutritionist for seven sessions (baseline, weeks 2, 4, 8, 12, 18, and 24) and a control group that received standard verbal diet information at baseline and nutritional counseling only at week 24. A 24-h recall technique was used to assess dietary intake for both groups at baseline and week 24. Lipid profile (total cholesterol, LDL, high-density lipoprotein (HDL), and triglyceride) was measured at baseline and after 12 and 24 weeks of therapy. An intention-to-treat and linear mixed model were used. Seventy-two patients were randomly assigned, and 62 (86%) participants completed their lipid profile test. After 12 weeks of follow-up, there were significant reductions in the intervention group for total cholesterol (?14.4?±?4.6?mg/dL, P?=?.002), LDL cholesterol (?13.7?±?4.1?mg/dL, P?=?.001), and triglyceride (?30.4?±?13.8?mg/dL, P?=?.03). A significant reduction in LDL cholesterol was also observed in the control group (?7.7?±?3.8?mg/dL, P?=?.04), but there were no significant differences in change of mean lipid levels between the groups at 12 weeks of follow-up. After 24 weeks, participants assigned to the intervention group demonstrated significantly greater decreases in serum total cholesterol (?19.0?±?4.6?vs. 0.2?±?4.3?mg/dL, P?=?.003) and LDL cholesterol (?21.5?±?4.1?vs. ?6.8?±?3.8?mg/dL, P?=?.009). There were no significant changes in HDL cholesterol or triglyceride levels in either group.     

The novel sodium glucose transporter 2 inhibitor dapagliflozin sustains pancreatic function and preserves islet morphology in obese, diabetic rats

Aims: To investigate whether glucose lowering with the selective sodium glucose transporter 2 (SGLT2) inhibitor dapagliflozin would prevent or reduce the decline of pancreatic function and disruption of normal islet morphology. Methods: Female Zucker diabetic fatty (ZDF) rats, 7–8 weeks old, were placed on high‐fat diet. Dapagliflozin (1 mg/kg/day, p.o.) was administered for ～33 days either from initiation of high‐fat diet or when rats were moderately hyperglycaemic. Insulin sensitivity and pancreatic function were evaluated using a hyperglycaemic clamp in anaesthetized animals (n = 5–6); β‐cell function was quantified using the disposition index (DI) to account for insulin resistance compensation. Pancreata from a matched subgroup (n = 7–8) were fixed and β‐cell mass and islet morphology investigated using immunohistochemical methods. Results: Dapagliflozin, administered from initiation of high‐fat feeding, reduced the development of hyperglycaemia; after 24 days, blood glucose was 8.6 ± 0.5 vs. 13.3 ± 1.3 mmol/l (p < 0.005 vs. vehicle) and glycated haemoglobin 3.6 ± 0.1 vs. 4.8 ± 0.26% (p < 0.003 vs. vehicle). Dapagliflozin improved insulin sensitivity index: 0.08 ± 0.01 vs. 0.02 ± 0.01 in obese controls (p < 0.03). DI was improved to the level of lean control rats (dapagliflozin 0.29 ± 0.04; obese control 0.15 ± 0.01; lean 0.28 ± 0.01). In dapagliflozin‐treated rats, β‐cell mass was less variable and significant improvement in islet morphology was observed compared to vehicle‐treated rats, although there was no change in mean β‐cell mass with dapagliflozin. Results were similar when dapagliflozin treatment was initiated when animals were already moderately hyperglycaemic. Conclusion: Sustained glucose lowering with dapagliflozin in this model of type 2 diabetes prevented the continued decline in functional adaptation of pancreatic β‐cells.     

Comparison of the antilipolytic effects of an A1 adenosine receptor partial agonist in normal and diabetic rats

Introduction and Aims: Elevated plasma free fatty acid (FFA) concentrations play a role in the pathogenesis of type 2 diabetes (2DM). Antilipolytic agents that reduce FFA concentrations may be potentially useful in the treatment of 2DM. Our previous observation that CVT‐3619 lowered plasma FFA and triglyceride concentrations in rats and enhanced insulin sensitivity in rodents with dietary‐induced forms of insulin resistance suggested that it might be of use in the treatment of patients with 2DM. The present study was undertaken to compare the antilipolytic effects of CVT‐3619 in normal (Sprague Dawley, SD) and Zucker diabetic fatty (ZDF) rats. Results: ZDF rats had significantly higher fat pad weight, glucose, insulin and FFA concentrations than those of SD rats. EC₅₀ values for forskolin‐stimulated FFA release from isolated adipocytes from SD and ZDF rats were 750 and 53 nM, respectively (p < 0.05). Maximal forskolin stimulation of FFA release was significantly (p < 0.01) less in ZDF rats (133 ± 60 μM) compared with SD rats (332 ± 38 μM). EC₅₀ values for isoproterenol to increase lipolysis in adipocytes from SD and ZDF rats were 2 and 7 nM respectively. Maximal isoproterenol‐stimulated lipolysis was significantly (p < 0.01) lower in adipocytes from ZDF rats (179 ± 23 μM) compared with SD rats (343 ± 27 μM). Insulin inhibited lipolysis in adipocytes from SD rats with an IC₅₀ value of 30 pM, whereas adipocytes from ZDF rats were resistant to the antilipolytic actions of insulin. In contrast, IC₅₀ values for CVT‐3619 to inhibit the release of FFA from SD and ZDF adipocytes were essentially the same (63 and 123 nM respectively). CVT‐3619 inhibited lipolysis more than insulin in both SD (86 vs. 46%, p < 0.001) and ZDF (80 vs. 13%, p < 0.001) adipocytes. In in vivo experiments, CVT‐3619 (5 mg/kg, PO) lowered FFA to a similar extent in both groups. Plasma concentrations of CVT‐3619 were not different in SD and ZDF rats. There was no significant difference in the messenger RNA expression of the A₁ receptors relative to β‐actin expression in adipocytes from SD (0.98 ± 0.2) and ZDF rats (0.99 ± 0.3). Conclusion: The antilipolytic effects of CVT‐3619 appear to be independent of insulin resistance and animal model.     

Altered gap junctional communication and renal haemodynamics in Zucker fatty rat model of type 2 diabetes

Aims/hypothesis  

We examined the link between altered gap junctional communication and renal haemodynamic abnormalities in diabetes in studies performed on Zucker lean (ZL) and the Zucker diabetic fatty (ZDF) rat model of type 2 diabetes.     

Sitagliptin Protects Cardiac Function by Reducing Nitroxidative Stress and Promoting Autophagy in Zucker Diabetic Fatty (ZDF) Rats

Purpose

The purpose of the study is to identify potential mechanisms involved in the cardiac protective effects of sitagliptin in Zucker diabetic fatty (ZDF) rats.

Methods and Results

Male non-diabetic lean Zucker rats (Lean) and ZDF rats treated with saline (ZDF) or sitagliptin (ZDF + sita) were used in this study. The blood pressure and lipid profiles were increased significantly in ZDF rats compared with Lean rats. ZDF + sitagliptin rats had decreased systolic blood pressure compared with ZDF rats. Sitagliptin treatment decreased total cholesterol (TC), triglycerides (TGs), low-density lipoprotein (LDL), and high-density lipoprotein (HDL) levels. Ejection fraction (EF) and fractional shortening (FS) were decreased in ZDF rats, which improved with sitagliptin from 59.8%?±?3.0 and 34.5%?±?3.1 to 66.9%?±?3.4 and 40.9%?±?4.2, respectively. Moreover, the nitroxidative stress level was increased while autophagy levels were decreased in ZDF rats, which was reversed by the administration of sitagliptin. Treatment with sitagliptin or FeTMPyP improved the autophagy level in high-glucose cultured H9c2 cells by increasing autolysosome numbers from 15?±?4 to 21?±?3 and 22?±?3, respectively. We detected a positive correlation between DPP-4 activity and 3-nitrotyrosine levels (r?=?0.3903; P?<?0.01), a negative correlation between Beclin-1 levels and DPP-4 activity (r?=???0.3335; P?<?0.01), and a negative correlation between 3-nitrotyrosine and Beclin-1 levels (r?=???0.3794; P?<?0.01) in coronary heart disease patients.

Conclusions

Sitagliptin alleviates diabetes-induced cardiac injury by reducing nitroxidative stress and promoting autophagy. This study indicates a novel target pathway for the treatment of cardiovascular complications in type 2 diabetes mellitus.

     

The vasopeptidase inhibitor AVE7688 ameliorates Type 2 diabetic nephropathy

Aim/hypothesis Pharmacological inhibition of the renin angiotensin system has proven clinical efficacy in nephropathies of various origins, including diabetic nephropathy. We tested the effects of the dual inhibition of both angiotensin converting enzyme and neutral endopeptidase by the vasopeptidase inhibitor AVE7688 in an animal model of Type 2 diabetic nephropathy.Methods We treated 56 obese Zucker diabetic fatty (ZDF, Gmi-fa/fa) rats aged 34-weeks with either placebo (n=9) or the vasopeptidase inhibitor AVE7688 in four different doses (each n=9; 3, 10, 30, or 60 mg/kg/d in chow). We used 11 heterozygous (+/fa) rats which received placebo and served as non-diabetic, lean controls. Urinary albumin/creatinine ratio was assessed as a marker of nephropathy at baseline (age 34-weeks) and after 10 weeks of chronic treatment.Results All obese animals had established diabetes mellitus that was not influenced by AVE7688 (HbA_1c >12%, stable in all dose groups). There was massive albuminuria in the homozygous ZDF rats (albumine/creatinine ratio >20 mg/mg vs minimal albuminuria in lean controls) that was decreased by AVE7688 in a dose dependent manner (Placebo 2.0±4.4 vs 11.9±1.8, 13.4±0.7, 13.6±2.8, and 19.8±2.8 mg/mg in the 3, 10, 30, and 60 mg/kg/d groups, respectively; all treatment groups p<0.05 vs Placebo).Conclusion/interpretation AVE7688 ameliorates proteinuria in Zucker diabetic fatty rats with established diabetes mellitus. Vasopeptidase inhibition represents an effective novel therapeutic principle for intervention in Type 2 diabetic nephropathy independent of metabolic control.Abbreviations ACE angiotensin converting enzyme - ZDF Zucker diabetic fatty     

Tissue-specific alterations of methyl group metabolism with DNA hypermethylation in the Zucker (type 2) diabetic fatty rat

Background Altered methyl group and homocysteine metabolism were tissue‐specific, persistent, and preceded hepatic DNA hypomethylation in type 1 diabetic rats. Similar metabolic perturbations have been shown in the Zucker (type 2) diabetic fatty (ZDF) rat in the pre‐diabetic and early diabetic stages, but tissue specificity and potential impact on epigenetic marks are unknown, particularly during pathogenesis. Methods ZDF (fa/fa) and lean (+/?) control rats were killed at 12 and 21 weeks of age, representing early and advanced diabetic conditions. Blood and tissues were analysed with respect to methyl group and homocysteine metabolism, including DNA methylation. Results At 12 weeks, hepatic glycine N‐methyltransferase (GNMT), methionine synthase, and cystathionine β‐synthase (CBS) activity and/or abundance were increased in ZDF rats. At 21 weeks, GNMT activity was increased in liver and kidney; however, only hepatic CBS protein abundance (12 weeks) and betaine‐homocysteine S‐methyltransferase mRNA expression (21 weeks) were significantly elevated (78 and 100%, respectively). Hepatic phosphatidylethanolamine N‐methyltransferase expression was also elevated in the ZDF rat. Homocysteine concentrations were decreased in plasma and kidney, but not in liver, at 12 and 21 weeks. In contrast to hepatic DNA hypomethylation in the type 1 diabetic rat, genomic DNA was hypermethylated at 12 and 21 weeks in the liver of ZDF rats, concomitant with an increase in DNA methyltransferase 1 expression at 21 weeks. Conclusions The pathogenesis of type 2 diabetes in the ZDF rat was associated with tissue and disease stage‐specific aberrations of methyl group and homocysteine metabolism, with persistent hepatic global DNA hypermethylation. Copyright © 2011 John Wiley & Sons, Ltd.     

The equivocal metabolic response to endotoxaemia in type 2 diabetic and obese ZDF rats

Aims/hypothesis The metabolic and endocrine disturbances associated with obesity and type 2 diabetes may impair the normal metabolic response to injury. Our objective was to investigate amino acid metabolism in endotoxaemic type 2 diabetic obese rats.Materials and methods A metabolic study was performed over 4 days using male Zucker diabetic fatty (ZDF) rats (fa/fa) and lean littermates (fa/+) divided into three groups: ad libitum-fed groups which underwent no treatment, lipopolysaccharide (LPS)-treated groups receiving E. coli LPS by i.p. injection, and pair-fed groups to the respective LPS groups. We evaluated the effect of endotoxaemia on body weight, food intake and tissue weights. Nitrogen loss and muscular proteolysis were measured daily by determination of urinary 3-methylhistidine (3-MH) excretion. Plasma, intestine and muscle amino acid levels were measured.Results The data showed that ad libitum-fed ZDF rats had lower plasma arginine and glutamine levels than ad libitum-fed control rats. Compared with control rats, the LPS-treated ZDF rats presented lower thymic involution, a lower 3-MH:creatinine ratio and higher cumulative nitrogen balance.Conclusions/interpretation Against our working hypothesis, ZDF rats did not show an impaired metabolic response, and even appeared to be less sensitive to the stress.L. Belabed and G. Senon made an equal contribution to this study.     

Modulation of adipoinsular axis in prediabetic zucker diabetic fatty rats by diazoxide

Dysregulation of the adipoinsular axis in male obese Zucker diabetic fatty (ZDF; fa/fa) rats, a model of type 2 diabetes, results in chronic hyperinsulinemia and increased de novo lipogenesis in islets, leading to beta-cell failure and diabetes. Diazoxide (DZ; 150 mg/kg.d), an inhibitor of insulin secretion, was administered to prediabetic ZDF animals for 8 wk as a strategy for prevention of diabetes. DZ reduced food intake (P < 0.02) and rate of weight gain only in ZDF rats (P < 0.01). Plasma insulin response to glucose load was attenuated in DZ-Zucker lean rats (ZL; P < 0.01), whereas DZ-ZDF had higher insulin response to glucose than controls (P < 0.001). DZ improved hemoglobin A1c (P < 0.001) and glucose tolerance in ZDF (P < 0.001), but deteriorated hemoglobin A1c in ZL rats (P < 0.02) despite normal tolerance in the fasted state. DZ lowered plasma leptin (P < 0.001), free fatty acid, and triglyceride (P < 0.001) levels, but increased adiponectin levels (P < 0.02) only in ZDF rats. DZ enhanced beta3-adrenoreceptor mRNA (P < 0.005) and adenylate cyclase activity (P < 0.01) in adipose tissue from ZDF rats only, whereas it enhanced islet beta3- adrenergic receptor mRNA (P < 0.005) but paradoxically decreased islet adenylate cyclase activity (P < 0.005) in these animals. Islet fatty acid synthase mRNA (P < 0.03), acyl coenzyme A carboxylase mRNA (P < 0.01), uncoupling protein-2 mRNA (P < 0.01), and triglyceride content (P < 0.005) were only decreased in DZ-ZDF rats, whereas islet insulin mRNA and insulin content were increased in DZ-ZDF (P < 0.01) and DZ-ZL rats (P < 0.03). DZ-induced beta-cell rest improved the lipid profile, enhanced the metabolic efficiency of insulin, and prevented beta-cell dysfunction and diabetes in diabetes-prone animals. This therapeutic strategy may be beneficial in preventing beta-cell failure and progression to diabetes in humans.