Young Zucker Diabetic Fatty Rats Research Articles

Captopril treatment does not affect inflammatory and autophagy signaling in skeletal muscle of Zucker diabetic fatty rat

Skeletal muscle plays a key role in the development of Type 2 Diabetes Mellitus (T2DM) as its large capacity to sequester glucose and contribute to the regulation of energy metabolism is compromised. Previous studies indicate that in skeletal muscle, T2DM drives inflammatory signaling and impairs autophagy. Captopril (Cap), an angiotensin-converting enzyme (ACE) inhibitor, is a common treatment for hypertension in T2DM patients; however, the impact of Cap on inflammatory signaling and autophagy in insulin resistant skeletal muscle remains ill-defined. We hypothesized that inflammatory signaling would be elevated and degradation of autophagosomes would be impaired in obese Zucker diabetic fatty (ZDF) rats, which are hyperphagic as they lack the leptin receptor. We further hypothesized that daily Cap treatment would attenuate these disturbances. At 14 weeks of age, the absolute weights of gastrocnemius (p<0.0001), tibialis anterior (p<0.0001), soleus (p=0.002), and extensor digitorum longus (EDL, p<0.0001) muscles were decreased by 23-34% in ZDF compared to lean rats but were not altered by 8 weeks of Cap treatment (0.5g/kg/day). Immunoblotting revealed that in soleus and EDL, inflammatory signaling proteins TLR4, phosphorylated (p)IKK α/β (ser176/180), pNFkB p65 (Ser 536), IkBα, TNFα, and AP1 were similar amongst all groups, whereas MyD88 was elevated in ZDF soleus (p=0.03) but was not corrected by Cap. The autophagy protein, LC3A/B I was increased 32% (p=0.01) in EDL from ZDF rats compared to Con; however, PI3K III, pBeclin1 (ser 93), pULK1 (ser555), LAMP2, LC3A/B II, and the LC3A/B II/I ratio were similar amongst groups in the soleus and EDL and were not altered by Cap. Compared to control, P62, a key autophagy adapter protein, was similar in soleus and decreased 27% in EDL (p<0.01) from ZDF rats and was not altered by Cap. Counter to our expectations, although muscle mass was lower in ZDF groups, obesity and insulin resistance in young ZDF rats were not suffcient to drive inflammatory signaling or disrupt autophagy in glycolytic or oxidative skeletal muscle, and these processes appeared to be resistant to Cap treatment in ZDF rats. USDA-NIFA awarded to MJR (2021-67017-33937). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Cocoa-rich diet attenuates beta cell mass loss and function in young Zucker diabetic fatty rats by preventing oxidative stress and beta cell apoptosis.

We have recently shown that cocoa flavanols may have anti-diabetic potential by promoting survival and function of pancreatic beta-cells in vitro. In this work, we investigated if a cocoa-rich diet is able to preserve beta-cell mass and function in an animal model of type 2 diabetes and the mechanisms involved. Our results showed that cocoa feeding during the prediabetic state attenuates hyperglycaemia, reduces insulin resistant, and increases beta cell mass and function in obese Zucker diabetic rats. At the molecular level, cocoa-rich diet prevented beta-cell apoptosis by increasing the levels of Bcl-xL and decreasing Bax levels and caspase-3 activity. Cocoa diet enhanced the activity of endogenous antioxidant defenses, mainly glutathione peroxidase, preventing thus oxidative injury induced by the pre-diabetic condition and leading to apoptosis prevention. These findings provide the first in vivo evidence that a cocoa-rich diet may delay the loss of functional beta-cell mass and delay the progression of diabetes by preventing oxidative stress and beta-cell apoptosis.

Cocoa intake ameliorates hepatic oxidative stress in young Zucker diabetic fatty rats

Chronic hyperglycemia in diabetes is associated with oxidative stress-mediated tissue damage. The present study is aimed to explore the role of a cocoa-enriched diet in ameliorating the oxidative stress-induced damage in the liver of young type 2 diabetic Zucker diabetic fatty (ZDF) rats. Male ZDF rats were fed a control or cocoa-rich diet (10%), and Zucker Lean (ZL) animals received the control diet. ZDF rats fed with cocoa (ZDF-Ca) decreased body weight gain, glucose and insulin levels, and improved glucose tolerance and insulin resistance. Cocoa diet further reduced reactive oxygen species (ROS) levels and carbonyl content in the liver of ZDF animals. The diminished activity of superoxide dismutase (SOD) and the enhanced activity of heme oxygenase (HO-1) in ZDF-C were returned to ZL values upon cocoa administration. Cocoa did not restore the decreased glutathione-S-transferase (GST) activity in both ZDF groups in comparison to ZL rats. Glutathione (GSH) content and activities of glutathione peroxidase (GPx), glutathione reductase (GR) and catalase (CAT) remained unaltered among all animal groups. Moreover, cocoa-rich diet suppressed total and phosphorylated nuclear factor erythroid-derived 2-like 2 (Nrf2), as well as p65-nuclear factor-kappaB (NF-ĸB) enhanced levels observed in ZDF rats. The results indicate that cocoa protects the hepatocytes by improving the antioxidant competence in the liver of young type 2 diabetic ZDF rats.

COX-2-Derived Prostanoids and Oxidative Stress Additionally Reduce Endothelium-Mediated Relaxation in Old Type 2 Diabetic Rats

Endothelial dysfunction in resistance arteries alters end organ perfusion in type 2 diabetes. Superoxides and cyclooxygenase-2 (COX-2) derivatives have been shown separately to alter endothelium-mediated relaxation in aging and diabetes but their role in the alteration of vascular tone in old diabetic subjects is not clear, especially in resistance arteries. Consequently, we investigated the role of superoxide and COX-2-derivatives on endothelium-dependent relaxation in 3 and 12 month-old Zucker diabetic fatty (ZDF) and lean (LZ) rats. Mesenteric resistance arteries were isolated and vascular tone was investigated using wire-myography. Endothelium (acetylcholine)-dependent relaxation was lower in ZDF than in LZ rats (60 versus 84% maximal relaxation in young rats and 41 versus 69% in old rats). Blocking NO production with L-NAME was less efficient in old than in young rats. L-NAME had no effect in old ZDF rats although eNOS expression level in old ZDF rats was similar to that in old LZ rats. Superoxide level and NADPH-oxidase subunits (p67phox and gp91phox) expression level were greater in ZDF than in LZ rats and were further increased by aging in ZDF rats. In young ZDF rats reducing superoxide level with tempol restored acetylcholine-dependent relaxation to the level of LZ rats. In old ZDF rats tempol improved acetylcholine-dependent relaxation without increasing it to the level of LZ rats. COX-2 (immunolabelling and Western-blot) was present in arteries of ZDF rats and absent in LZ rats. In old ZDF rats arterial COX-2 level was higher than in young ZDF rats. COX-2 blockade with NS398 restored in part acetylcholine-dependent relaxation in arteries of old ZDF rats and the combination of tempol and NS398 fully restored relaxation in control (LZ rats) level. Accordingly, superoxide production and COX-2 derivatives together reduced endothelium-dependent relaxation in old ZDF rats whereas superoxides alone attenuated relaxation in young ZDF or old LZ rats.

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 Fe(2+)/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 Fe(2+)/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; Fe(2+)/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 Fe(2+)/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.

Postprandial Insulin Resistance in Zucker Diabetic Fatty Rats is Associated with Parasympathetic‐Nitric Oxide Axis Deficiencies

The Zucker diabetic fatty (ZDF) rat is an obesity and type 2 diabetes model. Progression to diabetes is well characterised in ZDF rats, but only in the fasted state. We evaluated the mechanisms underlying postprandial insulin resistance in young ZDF rats. We tested the hypothesis that the overall postprandial action of insulin is affected in ZDF rats as a result of impairment of the hepatic parasympathetic-nitric oxide (PSN-NO) axis and/or glutathione (GSH), resulting in decreased indirect (PSN-NO axis) and direct actions of insulin. Nine-week-old male ZDF rats and lean Zucker rats (LZR, controls) were used. The action of insulin was assessed in the fed state before and after parasympathetic antagonism atropine. Basal hepatic NO and GSH were measured, as well as NO synthase (NOS) and γ-glutamyl-cysteine synthethase (GCS) activity and expression. ZDF rats presented postprandial hyperglycaemia (ZDF, 201.4 ± 12.9 mg/dl; LZR, 107.7 ± 4.3 mg/dl), but not insulinopaenia (ZDF, 5.9 ± 0.8 ng/ml; LZR, 1.5 ± 0.3 ng/ml). Total postprandial insulin resistance was observed (ZDF, 78.6 ± 7.5 mg glucose/kg; LZR, 289.2 ± 24.7 mg glucose/kg), with a decrease in both the direct action of insulin (ZDF, 54.8 ± 7.0 mg glucose/kg; LZR, 173.3 ± 20.5 mg glucose/kg) and the PSN-NO axis (ZDF, 24.5 ± 3.9 mg glucose/kg; LZR, 115.9 ± 19.4 mg glucose/kg). Hepatic NO (ZDF, 117.2 ± 11.4 μmol/g tissue; LZR, 164.6 ± 4.9 μmol/g tissue) and GSH (ZDF, 4.9 ± 0.3 μmol/g; LZR, 5.9 ± 0.2 μmol/g) were also compromised as a result of decreased NOS and GCS activity, respectively. These results suggest a compromise of the mechanism responsible for potentiating insulin action after a meal in ZDF rats. We show that defective PSN-NO axis and GSH synthesis, together with an impaired direct action of insulin, appears to contribute to postprandial insulin resistance in this model.

Ventricular myocyte contraction, intracellular calcium and expression of genes encoding cardiac muscle proteins in young and ageing Zucker diabetic fatty rat heart

Diabetes mellitus and its complications is a serious global health problem and the total number of people with this disease is projected to rise from 171 million in 2000 to 366 million in 2030. A recent study among Emirati citizens reported age-standardized rates for diabetes mellitus (diagnosed and undiagnosed) and pre-diabetes in those 30–64 years old as 29.0% and 24.2%, respectively. The association between type 2 diabetes mellitus and obesity is very strong and cardiovascular disease is the leading cause of morbidity and mortality among diabetic patients. The changes in ventricular myocyte contraction, intracellular calcium and the expression of genes encoding cardiac muscle proteins that take place in young (9–13 weeks) and ageing (30–34 weeks) Zucker diabetic fatty (ZDF) rat heart have been reviewed. Diabetes mellitus was associated with a fourfold elevation in non-fasting blood glucose in young and ageing ZDF rat compared with age-matched Zucker lean controls. Amplitude of shortening was unaltered in myocytes from young and ageing ZDF rats. Time to peak and time to half relaxation of shortening was prolonged in myocytes from young ZDF rats and was unaltered in myocytes from ageing ZDF rats compared with controls. Amplitude of the Ca2+ transient was unaltered in myocytes from young and ageing ZDF rats. Time to peak Ca2+ transient was prolonged in myocytes from young and ageing ZDF rats. L-type Ca2+ current was significantly reduced in myocytes from young and ageing ZDF rats. Sarcoplasmic reticulum Ca2+ transport did not appear to be altered in myocytes from young or ageing ZDF rats. Expression of genes encoding L-type Ca2+ channel proteins, plasma membrane transporters, sarcoplasmic reticulum Ca2+ and regulatory proteins and cardiac muscle proteins were variously up-regulated, down-regulated or unaltered in ventricles from young and ageing ZDF rats. Up-regulated genes in young ZDF rat heart included CACNA1C, CACNA1G, CACNA1H, ATP1A1 and MYH7, whereas down-regulated genes in young ZDF rat heart included ATP1B1, SLC9A1, ATP2A2, CALM1, MYH6, MYL2, ACTC1, TNNI3, TNNT2 and TNNC1. Up-regulated genes in ageing ZDF rat heart included CACNA1G, CACNA1H, ATP2A1 and MYL2, whereas down-regulated genes in ageing ZDF rat heart included CACNA2D3, SLC9A1, ATP2A2, MYH6 and TNNT2. Subtle changes in expression of genes encoding various cardiac muscle proteins may underlie functional changes in hearts of young and ageing ZDF rats compared with age-matched controls.

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.

Beneficial effects of melatonin on obesity and lipid profile in young Zucker diabetic fatty rats

The study objective was to investigate the effects of melatonin on obesity and obesity-associated systolic hypertension and dyslipidemia in young male Zucker diabetic fatty (ZDF) rats, an experimental model of the metabolic syndrome. 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 (n=10): naive (N), vehicle-treated (V), and melatonin-treated (M) (10mg/kg/day) for 6wk. Vehicle and melatonin were added to the drinking water. Melatonin reduced mean weight gain (51±2/100g BW) versus N-ZDF group (58±3, P<0.05) without food intake differences. M-ZDF rats showed an apparent reduction in systolic hypertension that proved not to be statistically significant, and a significant improvement in dyslipidemia, with a reduction in hypertriglyceridemia from 580±40 to 420.6±40.9mg/dL (P<0.01). Melatonin raised high-density-lipoprotein (HDL) cholesterol in ZDF (from 81.6±4.9 to 103.1±4.5mg/dL, P<0.01) and ZL rats (from 62.8±4.8 to 73.5±4.8mg/dL, P<0.05) and significantly reduced low-density-lipoprotein (LDL) cholesterol in ZDF rats from 5.20±0.4 to 4.14±0.3 mg/dL (P<0.05) but had no effect on total cholesterol levels. To our knowledge, this is the first evidence of a positive effect of melatonin on overweight and lipid pattern of obese Zucker diabetic rats, supporting the proposition that melatonin administration may ameliorate overweight and lipid metabolism in humans. Because these benefits occurred in youth, before advanced metabolic and vascular complications, melatonin might help to prevent cardiovascular disease associated with obesity and dyslipidemia.

Sympathetic modulation of baseline hindlimb blood flow and vascular conductance in a model of prediabetes using young Zucker Diabetic Fatty rats

Although associated with vascular disease, few studies have addressed the impact of prediabetes on vascular function. Thus, we sought to identify differences in baseline sympathetic vascular control between male Zucker (pre)Diabetic Fatty rats (PD) and lean controls (CTL) (n = 11/group). Hindlimb blood flow (Qh) and vascular conductance (VC) were assessed under local infusion of BIBP3226 (NPY Y1‐receptor (Y1R) antagonist), prazosin (α1‐receptor antagonist) and dual blockade. With no differences in baseline, BIBP3226 increased Qh 0.3 ± 0.1 ml/min in both groups and VC increased 4.2 ± 0.9 and 4.5 ± 1.1 μl/min/mmHg in PD and CTL respectively (P &lt; 0.05). Prazosin increased Qh 0.2 ± 0.04 and 0.2 ± 0.1 ml/min and VC increased 3.3 ± 0.5 and 4.2 ± 1.0 μl/min/mmHg in PD and CTL respectively (P &lt; 0.05). Dual blockade increased Qh 0.2 ± 0.05 ml/min in both groups and VC increased 4.6 ± 1.2 and 4.4 ± 0.72 μl/min/mmHg in PD and CTL respectively (P &lt; 0.05). Recovery time from maximum after drug infusion to 37% of baseline (1/e) for Qh and VC was shorter in PD vs. CTL (% of CTL, Qh and VC respectively: BIBP3226 68% and 53%, Prazosin 75% and 81%, dual blockade 78% and 70%) (P &lt; 0.05). In support, red and white vastus muscle Y1R, α1R, and tyrosine hydroxylase expression was higher in PD vs. CTL (P &lt; 0.05). These data suggest that the SNS is upregulated in prediabetes leading to persisting effects on skeletal muscle blood flow. NSERC and NIH.

Chronic treatment with the thiazolidinedione, MCC-555, is associated with reductions in nitric oxide synthase activity and beta-cell apoptosis in the pancreas of the Zucker Diabetic Fatty rat.

The Zucker Diabetic Fatty (ZDF) rat is a model of impaired insulin sensitivity arising from hyperphagia owing to a mutation in the leptin receptor. In time, young ZDF rats, which are not initially diabetic, develop impaired pancreatic beta-cell function leading to apoptotic cell death. This results in an inability to fully compensate for the reduction in insulin sensitivity with hypersecretion of insulin. Young, pre-diabetic ZDF rats were treated, over a 4-week period, with the thiazolidinedione compound MCC-555, and the islet morphology studied in comparison to ZDF rats not given MCC-555. In particular, changes in the apoptotic incidence, as measured using TUNEL staining to localize apoptotic cells, were studied over the 4-week period. Changes in the induction of nitric oxide synthase and in the accumulation of nitrate/nitrite within the pancreas were also studied during the time course of administration of MCC-555. The study has demonstrated that the administration of MCC-555 significantly decreases the apoptotic incidence in the islets of Langerhans of pre-diabetic ZDF rats given the compound, as compared to those not given MCC-555, as well as decreasing the accumulation of nitrate/nitrite within the pancreas.

The development of overt diabetes in young Zucker Diabetic Fatty (ZDF) rats and the effects of chronic MCC-555 treatment.

1. Young (6-week-old) pre-diabetic Zucker Diabetic Fatty (ZDF) rats displaying impaired glucose tolerance (IGT), moderate hyperglycaemia and hyperinsulinaemia were treated with the novel thiazolidinedione, MCC-555, for 28 days, during which time beta-cell failure and progression to overt diabetes occurs. 2. Treated ZDF rats exhibited consistently lower blood glucose levels than vehicle-treated diabetic controls, with a delayed rise and lower plateau levels. MCC-555 maintained plasma insulin levels throughout the treatment period, whereas these fell by 40% in untreated ZDF rats. 3. The rise in body weight was maintained in MCC-555-treated rats, whereas vehicle-treated rats exhibited blunted body weight gain after 8 weeks of age. Daily food intake was higher in diabetic, as compared to non-diabetic rats, but treatment did not modify food intake in diabetic rats. Water intake was lower in treated ZDF rats, concomitant with lowering of blood glucose. 4. The hyperinsulinaemic-euglycaemic clamp technique was applied to all rats after treatment to examine the effects of MCC-555 on insulin sensitivity. The glucose infusion rate to maintain normoglycaemia was lower in diabetic than in non-diabetic rats, demonstrating reduced glucose entry into insulin-sensitive tissues in diabetic rats. Increased glucose infusion rates were required to maintain euglycaemia in treated diabetic rats, demonstrating increased insulin sensitivity in these animals. 5. In conclusion, chronic MCC-555 treatment of young ZDF rats displaying IGT attenuates the development of overt diabetes through improved insulin sensitivity and maintenance of beta-cell function. MCC-555 may thus be beneficial in humans with IGT, to prevent or delay the progression of diabetes.