ZL Rats Research Articles

Synthesis and Anti-Diabetic Activity of an 8-Purine Derivative as a Novel DPP-4 Inhibitor in Obese Diabetic Zücker Rats.

Type 2 diabetes mellitus (T2DM) is one of the world's principal metabolic diseases characterized by chronic hyperglycemia. The gut incretin hormones, glucagon-like peptide 1 (GLP-1) and gastric inhibitory polypeptide (GIP), which has been proposed as a new treatment for T2DM, are extensively metabolized by Dipeptidyl peptidase 4 (DPP-4). Inhibitors of DPP-4 block the degradation of GLP-1 and GIP and may increase their natural circulating levels, favoring glycemic control in T2DM. A novel and potent selective inhibitor of DPP-4 with an 8-purine derived structure (1) has been developed and tested in vitro and in vivo in Zücker obese diabetic fatty (ZDF) rats, an experimental model of the metabolic syndrome and T2DM to assess the inhibitory activity using vildagliptin as reference standard. ZDF rats were subdivided into three groups (n = 7/group), control (C-ZDF), and those treated with compound 1 (Compound1-ZDF) and with vildagliptin (V-ZDF), both at 10 mg/kg/d rat body weight, in their drinking water for 12 weeks, and a group of lean littermates (ZL) was used. ZDF rats developed DM (fasting hyperglycemia, 425 ± 14.8 mg/dL; chronic hyperglycemia, HbA1c 8.5 ± 0.4%), compared to ZL rats. Compound 1 and vildagliptin reduced sustained HbAl1c (14% and 10.6%, P < 0.05, respectively) and fasting hyperglycemia values (24% and 19%, P < 0.05, respectively) compared to C-ZDF group (P < 0.001). Compound 1 and vildagliptin have shown a potent activity with an IC50 value of 4.92 and 3.21 µM, respectively. These data demonstrate that oral compound 1 administration improves diabetes in ZDF rats by the inhibitory effect on DPP-4, and the potential to be a novel, efficient and tolerable approach for treating diabetes of obesity-related T2DM, in ZDF rats.

Contribution of calcium dysregulation to impaired coronary artery contraction in Zucker diabetic fatty rats.

Diabetic coronary artery injury is closely associated with Ca2+ dysregulation, although the underlying mechanism remains unclear. This study explored the role and mechanism of Ca2+ handling in coronary artery dysfunction in type 2 diabetic rats. Zucker diabetic fatty (ZDF) rats were used as the type 2 diabetes mellitus model. The contractility of coronary artery rings induced by KCl, CaCl2 , 5-HT and U46619 was significantly lower in ZDF rats than in Zucker lean rats. Vasoconstriction induced by 5-HT and U46619 was greatly inhibited by nifedipine. However, in the presence of 1μM nifedipine or in the Ca2+ -free KH solution containing 1μM nifedipine, there was no difference in the vasoconstriction between Zucker lean and ZDF rats. Store-operated calcium channels (SOCs) were not involved in coronary vasoconstriction. The downregulation of contractile proteins and the upregulation of synthesized proteins were in coronary artery smooth muscle cells (CASMCs) from ZDF rats. Metformin reversed the reduction of vasoconstriction in ZDF rats. Taken together, L-type calcium channel is important for regulating the excitation-contraction coupling of VSMCs in coronary arteries, and dysregulation of this channel contributes to the decreased contractility of coronary arteries in T2DM.

Transcutaneous auricular vagal nerve stimulation inhibits limbic-regional P2X7R expression and reverses depressive-like behaviors in Zucker diabetic fatty rats

Zucker diabetic fatty (ZDF) rats develop type 2 diabetes (T2D) along with depressive-like behaviors. Transcutaneous auricular vagal nerve stimulation (taVNS) has antidiabetic and antidepressant-like effects in ZDF rats; however, the underlying antidepressant-like mechanisms are unclear. The purinergic receptor P2X7R, which is related to inflammation and depression, is upregulated in the limbic brain regions of depressed patients and rodents and is considered as a potential therapeutic target. Thus, this study aimed to provide preliminary evidence at the molecular level of taVNS antidepressant-like effect in ZDF rats through testing their limbic-regional P2X7R expression. ZDF rats were subjected to taVNS and transcutaneous non-vagal nerve stimulation (tnVNS). Body weight and blood glucose levels were monitored weekly. Depressive-like behaviors were evaluated with the open-field test (OFT) and forced swimming test (FST). Limbic-regional P2X7R expression was examined by western blotting (WB). P2X7R expressing cells were detected by immunohistochemistry (IHC). Compared to their lean littermates (ZL rats), ZDF rats developed obesity, hyperglycemia, and depressive-like behaviors with elevated limbic-regional P2X7R expression. taVNS but not tnVNS lowered body weight, reduced and stabilized blood glucose levels, suppressed limbic-regional P2X7R expression, and reversed the depressive-like behaviors. P2X7R was found primarily expressed in ZDF rats’ limbic-regional astrocytes. In conclusion, taVNS inhibits ZDF rats’ limbic-regional P2X7R expression, which may be one of the taVNS antidepressant-like mechanisms.

Expression of nucleic acid oxidation metabolites 8-Oxo-GSn and β-amyloid protein in the urine and cerebral tissues of diabetic rats

The aim of the study is to analyze the changes of 8-Oxo GSn expression and to explore the possible mechanism of type 2 diabetes mellitus combined with cognitive disorder using Zucker Diabetic Fatty rats (ZDF). Five-month-old ZDF rats and the wild-type control rats were employed in this study. Immunohistochemical (IHC) staining and isotope dilution high performance liquid chromatography - tandem mass spectrometry (ID - LC - MS/MS) method were used for analyzing 8-Oxo GSn in brain tissues and urine. The biochemical markers (Aβ40 and Aβ42) were also analyzed. In the cerebral cortex and hippocampus of ZDF rats, 8-Oxo GSn level was increased significantly than control rats. Aβ40 and Aβ42 levels were significantly increased in the brain of ZDF rats than control animals (P < 0.05). APP expression level was maximum in ZDF rats than that of control rats (P < 0.05). After eight months of 8-Oxo GSn injection into the brain of ZL rats, Aβ expression was considerably increased in ZL rats than control (P < 0.05). Overexpression of 8-Oxo GSn is one of the main reasons for ZL rats combined cognitive disorder.

Melatonin induces fat browning by transdifferentiation of white adipocytes and de novo differentiation of mesenchymal stem cells.

The role of melatonin in obesity control is extensively accepted, but its mechanism of action is still unclear. Previously we demonstrated that chronic oral melatonin acts as a brown-fat inducer, driving subcutaneous white adipose tissue (sWAT) into a brown-fat-like function (beige) in obese diabetic rats. However, immunofluorescence characterization of beige depots in sWAT and whether melatonin is a beige-fat inducer by de novo differentiation and/or transdifferentiation of white adipocytes are still undefined. Lean (ZL) and diabetic fatty (ZDF) Zücker rats were subdivided into two groups, control (C) and oral melatonin-supplemented (M, 10 mg kg-1 day-1) for 6 weeks. Mesenchymal stem cells (MSCs) were isolated from both rat inguinal fat and human lipoaspirates followed by adipogenesis assays with or without melatonin (50 nM for 12 h in a 24 h period, 12 h+/12 h-) mimicking the light/dark cycle. Immunofluorescence and western-blot assays showed the partial transdifferentiation of white adipocytes in both ZL and ZDF rats, with increasing thermogenic and beige markers, UCP1 and CITED1 and decreasing white adipocyte marker ASC-1 expression. In addition, melatonin increased UCP1, CITED1, and PGC1-α expression in differentiated adipocytes in both rats and humans. These results demonstrate that melatonin increases brown fat in obese diabetic rats by both adipocyte transdifferentiation and de novo differentiation. Furthermore, it promotes beige MSC adipogenesis in humans. This may contribute to the control of body weight attributed to melatonin and its metabolic benefits in human diabesity.

Electroacupuncture improves glucose and lipid metabolism by regulating APN/AMPK/PPARα signaling of skeletal muscle in Zucker diabetic obese rats

To observe the effect of electroacupuncture (EA) on skeletal muscle adiponectin receptor (Adipor1) / adenylate activated protein kinase (AMPK) / peroxisome proliferator-activated receptor α (PPARα) signaling pathway and skeletal muscle morphology by the secretion of serum adiponectin in Zucker diabetic obese (ZDF) rats, so as to explore its mechanism underlying regulating glucose and lipid metabolism of type 2 diabetes mellitus (T2DM) and improving skeletal muscle insulin resistance (IR). Twelve male ZDF rats and six Zucker thin (ZL) rats were selected. The rats were fed with Purina#5008 high-fat diet for four weeks to induce T2DM model after adaptive feeding with normal diet for one week. The ZDF rats were randomly divided into model group and EA group according to blood glucose level after modeling and 6 ZL rats were used as the blank control group. Rats in the EA group were treated with "Pishu" (BL20), EA stimulation of "Yishu" (EX-B3), "Zusanli" (ST36) and "Sanyinjiao" (SP6), once a day and 6 times a week for 4 weeks. Rats of the model and blank control groups were grabbed and fixed in the same way as EA group. Fasting blood glucose (FBG) was measured before and after EA intervention. Serum levels of insulin (INS), C-peptide (C-P), adiponectin (APN) were measured by radioimmunoassay, and those of free fatty acid (FFA), low-density lipoprotein (LDL), cholesterol (TC), triglyceride (TG) content determined by enzyme colorimetry and insulin resistance index (HOMA-IR) was calculated. The protein expression levels of AdipoR1, AMPK and PPARα proteins in the quadriceps femoris tissues were detected by Western blot and histopathological changes of quadriceps femoris muscle were observed by H.E. staining. Compared with the blank control group, the levels of FBG, serum INS, C-P, FFA, LDL, TC, TG and HOMA-IR in the model group were significantly increased (P<0.01, P<0.05), while the levels of serum APN and the expressions of AdipoR1, AMPK and PPARα proteins in the skeletal muscle were significantly decreased (P<0.01, P<0.05). Compared with the model group, the levels of FBG, serum INS, C-P, FFA, LDL, TC and HOMA-IR in the EA group were significantly decreased (P<0.01), and those of serum APN and expression levels of AdipoR1, AMPK and PPARα proteins in the skeletal muscle significantly increased (P<0.01), but the serum TG level had no remarkable change in the EA group (P>0.05). In addition, H.E. staining showed disordered arrangement of skeletal muscle cells, rupture and fuzziness of muscle fibers, enlargement of the space between muscular fibers and infiltration of small number of adipose cells which were relatively milder in the EA group. EA can reduce blood glucose and lipid levels, and improve IR in ZDF rats, which may be related to its functions in up-regulating AdipoR1/AMPK/PPARα signaling and in promoting APN secretion.

Vitamin A Status Regulates the Development of Obesity and Type 2 Diabetes in Zucker Diabetic Fatty Rats

Abstract Objectives Here, we studied the effects of VA status on the development of obesity and type 2 diabetes in Zucker diabetic fat (ZDF) rats. Methods Zucker Lean (ZL) and ZDF rats at weaning were divided into 6 groups, VA deficient with basal fat (VAD-BF, 0 mg retinyl palmitate (RP)/kg and 22.1% fat energy), VA marginal with BF (VAM-BF, 0.35 mg RP/kg), VA sufficient with BF (VAS-BF, 4.0 mg RP/kg), VAD with high-fat (VAD-HF, 60% fat energy), VAM-HF and VAS-HF diets, and fed for 8 weeks (w). The body mass (BM), and peripheral blood glucose (PBG) were measured weekly. An oral glucose tolerance test (OGTT) were done at 6.5 and 7.5w, respectively. Plasma levels of glucose, insulin, triacylglycerol and cholesterol l were determined using commercially available kits. The expression levels of genes and proteins in the liver of rats were analyzed using PCR and Western blot. Results VAS-BF ZL and ZDF rats from 6w had respectively higher BM than VAD/VAM-BF ZL and ZDF rats. VAS-HF ZL and ZDF rats from 4w had respectively higher BM than VAD/VAM-HF ZL and ZDF rats. VAS-BF/HF ZDF rats from 6w had respectively higher PBG levels than VAD/VAM-BF/HF ZDF rats. The OGTT AUC values of VAS-BF/HF ZL/ZDF rats were respectively higher than that of VAD/VAM-BF/HF ZL/ZDF rats. The levels of glucose, insulin, triacylglycerol and cholesterol in VAD/VAM/VAS-BF ZDF and VAD/VAM/VAS-HF ZDF rats were higher than that in BF ZL rats (except for the glucose level) and HF ZL rats, respectively. The hepatic Gck mRNA and its protein levels in VAD-BF ZL rats were lower than that in VAS-BF ZL rats. The hepatic levels of Fas, and Acl mRNA and FAS, and ACL proteins in VAM/VAS-HF ZF rats were higher than that in VAM/VAS-HF ZL rats. The hepatic retinol content of VAD-BF/HF ZL/ZDF rats were lower than that of VAM groups, which are lower than that of VAS-BF/HF ZL/ZDF rats. Conclusions VA statuses affect BM gain in ZL and ZDF rats fed a BF or a HF diet. The expression levels of mRNAs and proteins in the fatty acid biosynthesis pathways were reduced in VAD-HF ZDF rats. The effects of VA on fatty acid biosynthesis in ZDF rats were masked in a HF diet setting. Reduced VA intake prevents obesity, and type 2 diabetes in ZDF rats. Funding Sources Diabetes Action Research and Education Foundation

Transcutaneous auricular vagal nerve stimulation inhibits hypothalamic P2Y1R expression and attenuates weight gain without decreasing food intake in Zucker diabetic fatty rats.

Zucker diabetic fatty (ZDF) rats that harbor a mutation in the leptin receptor innately develop type 2 diabetes (T2D) with obesity. Transcutaneous auricular vagal nerve stimulation (taVNS) has an antidiabetic effect in ZDF rats. However, the underlying mechanisms of the weight-gain attenuating effect in ZDF rats by taVNS is still unclear. This study aimed to assess whether the weight-gain attenuating effect of taVNS in ZDF rats is associated with changes in the central nervous system (CNS) expression of P2Y1 receptors (P2Y1R). Adult male ZDF rats were subjected to taVNS and transcutaneous non-vagal nerve stimulation (tnVNS). Their food intake and body weight were recorded daily and weekly, respectively. P2Y1R expression in the hypothalamus, amygdala, and hippocampus was evaluated by western blotting. Hypothalamic P2Y1R expressing cells were detected using immunohistochemistry. Naïve ZDF rats were much heavier (p < 0.05) than their lean littermates (ZL rats), with elevated hypothalamic P2Y1R expression (p < 0.05). Further, taVNS but not tnVNS attenuated weight gain (p < 0.05) without decreasing food intake (p > 0.05) and suppressed hypothalamic P2Y1R expression in ZDF rats (p < 0.05). Moreover, P2Y1R showed major expression in astrocytes of ZDF rats' hypothalamus. ZDF rats innately develop obesity associated with elevated hypothalamic P2Y1R expression. taVNS attenuates weight gain in ZDF rats without changes in food intake, suggesting increased energy expenditure. Whether the reduced hypothalamic P2Y1R expression in response to taVNS is mechanistically linked to the increased energy expenditure remains to be determined.

Melatonin Improves Endoplasmic Reticulum Stress-Mediated IRE1α Pathway in Zücker Diabetic Fatty Rat.

Obesity and diabetes are linked to an increased prevalence of kidney disease. Endoplasmic reticulum stress has recently gained growing importance in the pathogenesis of obesity and diabetes-related kidney disease. Melatonin, is an important anti-obesogenic natural bioactive compound. Previously, our research group showed that the renoprotective effect of melatonin administration was associated with restoring mitochondrial fission/fusion balance and function in a rat model of diabesity-induced kidney injury. This study was carried out to further investigate whether melatonin could suppress renal endoplasmic reticulum (ER) stress response and the downstream unfolded protein response activation under obese and diabetic conditions. Zücker diabetic fatty (ZDF) rats and lean littermates (ZL) were orally supplemented either with melatonin (10 mg/kg body weight (BW)/day) (M–ZDF and M–ZL) or vehicle (C–ZDF and C–ZL) for 17 weeks. Western blot analysis of ER stress-related markers and renal morphology were assessed. Compared to C–ZL rats, higher ER stress response associated with impaired renal morphology was observed in C–ZDF rats. Melatonin supplementation alleviated renal ER stress response in ZDF rats, by decreasing glucose-regulated protein 78 (GRP78), phosphoinositol-requiring enzyme1α (IRE1α), and ATF6 levels but had no effect on phospho–protein kinase RNA–like endoplasmic reticulum kinase (PERK) level. In addition, melatonin supplementation also restrained the ER stress-mediated apoptotic pathway, as indicated by decreased pro-apoptotic proteins phospho–c–jun amino terminal kinase (JNK), Bax, and cleaved caspase-3, as well as by upregulation of B cell lymphoma (Bcl)-2 protein. These improvements were associated with renal structural recovery. Taken together, our findings revealed that melatonin play a renoprotective role, at least in part, by suppressing ER stress and related pro-apoptotic IRE1α/JNK signaling pathway.

SO022EVALUATION OF GLOMERULAR HEMODYNAMIC CHANGES BY SGLT2 INHIBITION IN TYPE 2 DIABETIC RATS USING IN VIVO IMAGING TECHNIQUES

Abstract Background and Aims In recent clinical trials, the SGLT2 inhibitor (SGLT2i) slowed the progression of kidney disease compared with the placebo in patients with type 2 diabetes. Improvement of glomerular hyperfiltration via tubuloglomerular feedback (TGF) is considered to be one of the possible pathways for renal protection with SGLT2 inhibition (SGLT2i) in diabetic kidney disease (DKD). We have successfully developed the novel method to measure single-nephron GFR (SNGFR) in mice using multiphoton laser microscopy and demonstrated that the adenosine/adenosine A1 receptor (A1aR) pathway plays a pivotal role in the TGF mechanism in the type 1 diabetic model, Akita mice (Kengo Kidokoro, David Z. I. Cherney et al. Circulation. 2019). It has been suggested that the mechanism of improvement effects in glomerular hyperfiltration by SGLT2i is different in type 1 diabetes and type 2 diabetes. However, the detailed regulatory mechanism of GFR by SGLT2i is not fully understood in type 2 diabetes. This study aims to clarify the effects of SGLT2i on glomerular hemodynamics in type 2 diabetic rats. Method Zucker lean (ZL) rats and Zucker diabetic fatty (ZDF) rats were used. In the first experiment, SNGFR and diameters of glomerular afferent/efferent arterioles were measured in both groups. Next, we examined the change of SNGFR and diameters of glomerular afferent/efferent arterioles, as well as urinary excretions of glucose and sodium in ZDF after a single-dose administration of SGLT2i (luseogliflozin; 10mg/kg, gavage) for 120 minutes, which generated the following three groups: SGLT2i group, SGLT2i + adenosine A1 receptor (A1aR) antagonist (8-cyclopentyl-1,3-dipropylxanthine, 1mg/kg) group, and insulin group. Results SNGFR in the ZDF group was significantly higher than in the ZL group. The diameter of the afferent arteriole and efferent arteriole was also wider in ZDF rats than in ZL rats. The SNGFR and diameter of the afferent arteriole were significantly decreased after a single-dose administration of SGLT2i in ZDF. However, there was no significant diameter change in the efferent arteriole. Moreover, a decrease of SNGFR was not observed in the A1aR antagonist group after SGLT2i administration. Urinary excretions of glucose and sodium showed a similar pattern in the SGLT2i and SGLT2i+ A1aR antagonist groups. Conclusion The adenosine/A1aR pathway plays an important role in the regulation of the tonus of the afferent arteriole and is involved in the suppression of glomerular hyperfiltration by SGLT2 inhibition in type 2 diabetes.

Vitamin A Status Regulates the Development of Obesity and Type 2 Diabetes in Zucker Diabetic Fatty Rats

Abstract Objectives We have shown that vitamin A (VA) status regulates obesity and fuel metabolism in rats. Here, we studied the effects of VA status on the development of obesity and type 2 diabetes in Zucker diabetic fat (ZDF) rats. Methods Zucker Lean (ZL) and ZDF rats at weaning were divided into 6 groups, VA deficient with basal fat (VAD-BF, 0 mg retinyl palmitate (RP)/kg and 22.1% fat energy), VA marginal with BF (VAM-BF, 0.35 mg RP/kg), VA sufficient with BF (VAS-BF, 4.0 mg RP/kg), VAD with high-fat (VAD-HF, 60% fat energy), VAM-HF and VAS-HF diets, and fed for 8 weeks (w). The body mass (BM), and peripheral blood glucose (PBG) were measured weekly. Insulin tolerance test (ITT) and oral glucose tolerance test (OGTT) were done at 6.5 and 7.5 w, respectively. At the end of the feeding, blood, liver and white adipose tissue (WTA) samples were collected. Results VAS-BF ZL and ZDF rats from 6 w had respectively higher BM than VAD/VAM-BF ZL and ZDF rats. VAS-HF ZL and ZDF rats from 4 w had respectively higher BM than VAD/VAM-HF ZL and ZDF rats. The liver/BM and WAT/BM ratios in VAD/VAM-BF/HF ZL and ZDF V rats were respectively lower than that of VAS-BF/HF groups. VAS-BF/HF ZDF rats from 6 w had respectively higher PBG levels than VAD/VAM-BF/HF ZDF rats. In ITT, PBG levels of VAD/VAM/VAS-BF ZL rats dropped until 15 mins. PBG levels of VAD-HF and VAM/VAS-HF ZL rats declined until 30 mins and 15 mins, respectively. PBG levels of VAM-BF and VAS-BF ZDF rats dropped until 15 mins and 5 mins, respectively. PBG levels of VAD-BF ZDF rats start to dropped after 10 mins and stopped after 20 mins. PBG levels of VAD/VAM-HF and VAS-HF ZDF rats dropped until 15 mins and 20 mins, respectively. The OGTT results showed that PBG levels of VAS-BF ZL rats peaked at 10 mins, and VAS-BF/HF ZL rats had respectively higher PBG levels than VAD/VAM-BF/HF ZL rats. PBG levels of all ZDF rats peaked at 60 mins (except for VAS-BF ZDF rats at 30) before dropped. TheOGTT area under the curve values of VAS-BF/HF ZL or ZDF rats were respectively higher than that of VAD/VAM-BF/HF ZL or ZDF rats, and that of VAM-HF ZL rats were higher than that of VAD-HF ZL rats. Conclusions VA statuses affect BM gain in ZL and ZDF rats in BF and HF diets. Reduced VA intake prevents obesity, and type 2 diabetes in ZDF rats. Funding Sources Diabetes Action Research and Education Foundation.

Pioglitazone downregulates Twist-1 expression in the kidney and protects renal function of Zucker diabetic fatty rats

AimsRenal interstitial fibrosis and glomerulosclerosis are the characteristic presentation of diabetic nephropathy progression. Twist-1 overexpression contributes to renal fibrosis. Previous studies have demonstrated that pioglitazone (PIO), a PPAR-γ agonists, can ameliorate renal fibrosis and protect renal function. However, whether PIO attenuates renal fibrosis and delays diabetic nephropathy progression by regulating Twist-1 expression remains unclear. MethodsMale Zucker diabetic fatty (ZDF) rats were randomly divided into 3 groups: (1) ZDF group, (2) ZDF + PIO group treated with PIO for 10 weeks, (3) ZDF + PIO + GW9662 group treated with GW9662 (a PPAR-γ antagonist) and PIO for 10 weeks. Age-matched Zucker lean rats (ZL group) were used as a control group. Urinary albumin/creatinine ratio (UACR) and renal blood flow were measured. Renal histopathology and Twist-1 expression were determined by immunohistochemistry. The protein and mRNA levels of Twist-1 and PPAR-γ were analyzed by Western blot and qRT-PCR. ResultsPIO considerably reduced UACR and improved renal blood flow. This was associated with amelioration of glomerulosclerosis and tubulointerstitial fibrosis evidenced by the expression decrease of collagen I, aquaporin 1, α-SMA, transforming growth factor β1 and nephrin, although glycaemia remained high. Moreover, Twist-1 protein and mRNA expression in kidney of ZDF rats were significantly increased compared with ZL rats and PIO significantly decreased Twist-1 levels. ConclusionsThis study shows that PIO can downregulate Twist-1 expression in the kidney, inhibit renal fibrosis and protect renal function in ZDF rats. These PIO-mediated effects are independent of glycemic control.

Renal Protective Effects of N-Acetyl-Seryl-Aspartyl-Lysyl-Proline (Ac-SDKP) in Obese Rats on a High-Salt Diet

BACKGROUNDObesity is a public health problem, associated with salt sensitive hypertension, kidney inflammation, and fibrosis. N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) is a tetra peptide with anti-inflammatory and anti-fibrotic properties. However, its effect on preventing kidney damage in obesity is unknown. We hypothesized that Zucker obese (ZO) rats on a high-salt (HS) diet develop renal damage, inflammation, fibrosis, and this is prevented with Ac-SDKP treatment.METHODSZucker lean (ZL) and ZO rats (8 weeks old) were treated with Ac-SDKP (1.6 mg/kg/day) while maintained on either a normal-salt (NS; 0.4%) or HS (4%) diet for 8 weeks. Systolic blood pressure (SBP), albuminuria, renal inflammation, and fibrosis were evaluated.RESULTSHS diet increased macrophage infiltration in the kidneys of both ZL and ZO rats but was significantly higher in ZO rats receiving the HS diet (ZL + NS, 13.9 ± 1.3 vs. ZL + HS, 19.14 ± 1.5 and ZO + NS, 25.5 ± 1.4 vs. ZO + HS, 87.8 ± 10.8 cells/mm2; P < 0.05). Ac-SDKP prevented macrophage infiltration in ZO rats (ZO + HS + Ac-SDKP, 32.18 ± 2.4 cells/mm2; P < 0.05). Similarly, glomerulosclerosis, cortical, and medullary interstitial fibrosis were increased in ZO rats fed the HS diet, and Ac-SDKP attenuated these alterations (P < 0.05). SBP was increased in ZO rats fed the HS diet (ZO + NS, 121.3 ± 8.9 vs. ZO + HS, 164 ± 6.9 mm Hg; P < 0.05), and it was significantly decreased with Ac-SDKP treatment (ZO + HS + Ac-SDKP, 144.05 ± 14.1 mm Hg; P = 0.004). Albuminuria was higher in ZO rats than in ZL rats; however, neither HS nor Ac-SDKP treatment affected it.CONCLUSIONSAc-SDKP treatment in ZO rats fed a HS diet prevented renal damage by reducing inflammation, fibrosis, and SBP.

Obesity‐associated glomerular inflammation increases albuminuria without renal histological changes

Obesity is one of risk factors for chronic kidney disease (CKD), but the precise mechanism involved is unclear. This study characterizes the effect of obesity‐induced glomerular inflammation, oxidative stress, and albuminuria in obese rats. Glomerular samples were collected from fatty (ZF) and lean (ZL) Zucker rats. After 2 months of feeding, body weight and albuminuria were significantly increased in ZF rats when compared to ZL rats. Expression of the inflammatory markers TNF‐α and CCR2 was significantly increased in the glomeruli of ZF rats. However, expression of IL‐6 mRNA was not increased. Analysis of renal pathology showed no glomerular expansion. As inflammatory and oxidative stress markers are associated with NF‐κB, we evaluated whether NF‐κB activation was increased in the glomeruli of mice on a high‐fat diet. Immunohistochemistry showed increased NF‐κB activation in the glomeruli when transgenic mice overexpressing an NF‐κB‐dependent enhanced green fluorescent protein were fed with a high‐fat diet. These results suggest that obesity of only 2 months duration can cause albuminuria, due to increased inflammation or oxidative stress, but may not be long enough to develop renal pathological changes.

In Situ Activation and Preservation of Penile Progenitor Cells Using Icariside II in an Obesity-Associated Erectile Dysfunction Rat Model.

Obesity-associated erectile dysfunction (ED) involves pathologic change that may be related to deficit of the penile endogenous stem/progenitor cells. Therefore, an in-depth study of the penile stem/progenitor cells in the pathogenesis of ED is warranted. For this study, eight Zucker Lean (ZUC-Leprfa 186; ZL group) and 16 Zucker Fatty (ZUC-Leprfa 185; ZF) male rats received an intraperitoneal injection of 5-ethynyl-2-deoxyuridine (EdU) to track endogenous stem cells. Twelve weeks later, the ZF rats were randomized to gavage feeding with 1.5 mg/kg/day of icariside II (ZF + ICA II group) or the solvent (ZF group). Treatment lasted 4 weeks and was followed by a 1-week washout period. ZF rats had impaired erectile function with related pathologic changes compared with ZL rats. ICA II treatment restored erectile function and prevented smooth muscle atrophy, endothelial dysfunction, and lipid accumulation compared with no treatment. EdU label-retaining cell levels were higher in the ZF + ICA II group compared with the ZF group. Histone 3 phosphorylation at Ser 10, a specific mitotic cell marker, was additionally used to identify dividing cells. ICA II activated more penile stem cells to proliferate in ZF rats compared with ZL rats. These results suggest that ZF rats can be used as a model for obesity-associated ED and that ICA II improves erectile function and pathologic changes through endogenous progenitor cell preservation and proliferation.

Deciphering bacterial community changes in zucker diabetic fatty rats based on 16S rRNA gene sequences analysis.

The aim of the present pilot study was deciphering bacterial community changes in Zucker diabetic fatty rats (ZDF rats), a model of type 2 diabetes. Recent studies unmasked that the status of gastrointestinal tract microbiota has a marked impact on nutrition-related syndromes such as obesity and type-2 diabetes (T2D). In this study, samples taken from the gastrointestinal tracts (GI tracts) of ZDF and their lean littermates (ZL rats) were subjected to 16S rRNA gene sequence-based analysis to examine the characteristic bacterial communities, including those located in the stomach, duodenum, jejunum, ileum, cecum and feces. Results revealed that the Firmicutes/Bacteroidetes ratio was increased and greater numbers of Lactobacillus were detected along GI tracts in ZDF rats compared to ZL rats. In conclusion, this work is the first study to systematically characterize bacterial communities along ZDF rat GI tract and provides substantial evidence supporting a prospective strategy to alter the GI microbial communities improving obesity and T2D.

Oral treatment with a zinc complex of acetylsalicylic acid prevents diabetic cardiomyopathy in a rat model of type-2 diabetes: activation of the Akt pathway.

BackgroundType-2 diabetics have an increased risk of cardiomyopathy, and heart failure is a major cause of death among these patients. Growing evidence indicates that proinflammatory cytokines may induce the development of insulin resistance, and that anti-inflammatory medications may reverse this process. We investigated the effects of the oral administration of zinc and acetylsalicylic acid, in the form of bis(aspirinato)zinc(II)-complex Zn(ASA)2, on different aspects of cardiac damage in Zucker diabetic fatty (ZDF) rats, an experimental model of type-2 diabetic cardiomyopathy.MethodsNondiabetic control (ZL) and ZDF rats were treated orally with vehicle or Zn(ASA)2 for 24 days. At the age of 29–30 weeks, the electrical activities, left-ventricular functional parameters and left-ventricular wall thicknesses were assessed. Nitrotyrosine immunohistochemistry, TUNEL-assay, and hematoxylin-eosin staining were performed. The protein expression of the insulin-receptor and PI3K/AKT pathway were quantified by Western blot.ResultsZn(ASA)2-treatment significantly decreased plasma glucose concentration in ZDF rats (39.0 ± 3.6 vs 49.4 ± 2.8 mM, P < 0.05) while serum insulin-levels were similar among the groups. Data from cardiac catheterization showed that Zn(ASA)2 normalized the increased left-ventricular diastolic stiffness (end-diastolic pressure–volume relationship: 0.064 ± 0.008 vs 0.084 ± 0.014 mmHg/µl; end-diastolic pressure: 6.5 ± 0.6 vs 7.9 ± 0.7 mmHg, P < 0.05). Furthermore, ECG-recordings revealed a restoration of prolonged QT-intervals (63 ± 3 vs 83 ± 4 ms, P < 0.05) with Zn(ASA)2. Left-ventricular wall thickness, assessed by echocardiography, did not differ among the groups. However histological examination revealed an increase in the cardiomyocytes’ transverse cross-section area in ZDF compared to the ZL rats, which was significantly decreased after Zn(ASA)2-treatment. Additionally, a significant fibrotic remodeling was observed in the diabetic rats compared to ZL rats, and Zn(ASA)2-administered ZDF rats showed a similar collagen content as ZL animals. In diabetic hearts Zn(ASA)2 significantly decreased DNA-fragmentation, and nitro-oxidative stress, and up-regulated myocardial phosphorylated-AKT/AKT protein expression. Zn(ASA)2 reduced cardiomyocyte death in a cellular model of oxidative stress. Zn(ASA)2 had no effects on altered myocardial CD36, GLUT-4, and PI3K protein expression.ConclusionsWe demonstrated that treatment of type-2 diabetic rats with Zn(ASA)2 reduced plasma glucose-levels and prevented diabetic cardiomyopathy. The increased myocardial AKT activation could, in part, help to explain the cardioprotective effects of Zn(ASA)2. The oral administration of Zn(ASA)2 may have therapeutic potential, aiming to prevent/treat cardiac complications in type-2 diabetic patients.

Acute exacerbation of sleep apnea by hyperoxia impairs cognitive flexibility in Brown-Norway rats.

To determine whether learning deficits occur during acute exacerbation of spontaneous sleep related breathing disorder (SRBD) in rats with high (Brown Norway; BN) and low (Zucker Lean; ZL) apnea propensity. Spatial acquisition (3 days) and reversal learning (3 days) in the Morris water maze (MWM) with polysomnography (12:00-08:00): (1) with acute SRBD exacerbation (by 20-h hyperoxia immediately preceding reversal learning) or (2) without SRBD exacerbation (room air throughout). Randomized, placebo-controlled, repeated-measures design. 14 BN rats; 16 ZL rats. 20-h hyperoxia. Apneas were detected as cessation of respiration ≥ 2 sec. Swim latency in MWM, apnea indices (AI; apneas/hour of sleep) and percentages of recording time for nonrapid eye movement (NREM), rapid eye movement (REM), and total sleep were assessed. Baseline AI in BN rats was more than double that of ZL rats (22.46 ± 2.27 versus 10.7 ± 0.9, P = 0.005). Hyperoxia increased AI in both BN (34.3 ± 7.4 versus 22.46 ± 2.27) and ZL rats (15.4 ± 2.7 versus 10.7 ± 0.9) without changes in sleep stage percentages. Control (room air) BN and ZL rats exhibited equivalent acquisition and reversal learning. Acute exacerbation of AI by hyperoxia produced a reversal learning performance deficit in BN but not ZL rats. In addition, the percentage of REM sleep and REM apnea index in BN rats during hyperoxia negatively correlated with reversal learning performance. Acute exacerbation of sleep related breathing disorder by hyperoxia impairs reversal learning in a rat strain with high apnea propensity, but not a strain with a low apnea propensity. This suggests a non-linear threshold effect may contribute to the relationships between sleep apnea and cognitive dysfunctions, but strain-specific differences also may be important.

Transcutaneous auricular vagus nerve stimulation triggers melatonin secretion and is antidepressive in Zucker diabetic fatty rats.

Decreased circulating melatonin is implicated in depression. We recently found that Zucker diabetic fatty rats (ZDF, fa/fa) develop depression-like behaviors and that transcutaneous auricular vagus nerve stimulation (taVNS) is antidepressive in ZDF rats. Here we studied whether the ZDF rats could be used as a depression rodent model and whether the antidepressive effect of taVNS is mediated through modulation of melatonin secretion. Adult male ZDF and Zucker lean (ZL, fa/+) littermates were used. 30 min-taVNS procedures (2/15 Hz, 2 mA) were administered once daily under anesthesia for 34 consecutive days in pineal intact ZDF (n = 8) and ZL (n = 6) rats, as well as in pinealectomized ZDF rats (n = 8). Forced swimming test (FST) was used to determine depression-like behavior and ELISA to detect plasma melatonin concentration on day 35. We found that naïve ZDF rats had a longer immobility time in FST and that long-term (34 days) taVNS treatment ameliorated the depression-like behavior. In both pineal intact and pinealectomized ZDF rats, taVNS induced acute melatonin secretion, both during and after the taVNS session. A low melatonin level is related to the poor FST performance in ZDF rats (R = −0.544) in contrast to ZL rats (R = 0.247). In conclusion, our results show that ZDF rats are ideal candidates of innate depression and that taVNS is antidepressive through triggering melatonin secretion and increasing its production.

Obesity-associated Gingival Vascular Inflammation and Insulin Resistance

Obesity is a risk factor for periodontitis, but the pathogenic mechanism involved is unclear. We studied the effects of insulin in periodontal tissues during the state of obesity-induced insulin resistance. Gingival samples were collected from fatty (ZF) and lean (ZL, control) Zucker rats. Endothelial nitric oxide synthase (eNOS) expression was decreased, and activities of protein kinase C (PKC) α, ß2, δ, and ϵ isoforms were significantly increased in the gingiva from ZF rats compared with those from ZL rats. Expression of oxidative stress markers (mRNA) and the p65 subunit of NF-κB was significantly increased in ZF rats. Immunohistochemistry revealed that NF-κB activation was also increased in the gingival endothelial cells from transgenic mice overexpressing NF-κB-dependent enhanced green fluorescent protein (GFP) and on a high-fat vs. normal chow diet. Analysis of the gingiva showed that insulin-induced phosphorylation of IRS-1, Akt, and eNOS was significantly decreased in ZF rats, but Erk1/2 activation was not affected. General PKC inhibitor and an anti-oxidant normalized the action of insulin on Akt and eNOS activation in the gingiva from ZF rats. This provided the first documentation of obesity-induced insulin resistance in the gingiva. Analysis of our data suggested that PKC activation and oxidative stress may selectively inhibit insulin-induced Akt and eNOS activation, causing endothelial dysfunction and inflammation.