ZF Rats Research Articles

Replenishment of vitamin A for 7 days partially restored hepatic gene expressions altered by its deficiency in rats.

We investigated the effects of vitamin A (VA) status on metabolism of Zucker rats with different genders and genotypes, and of short-term refeeding of a VA sufficient (VAS) diet on VA deficient (VAD) animals. First, male and female Zucker lean (ZL) and fatty (ZF) rats at weaning were fed a VAD or VAS diet for 8 weeks. Second, male VAD ZL rats were fed a VAS diet for 3 (VAD-VAS3d) or 7 (VAD-VAS7d) days. The body weight (BW), blood parameters, and hepatic expressions of genes for metabolism were determined. VA deficiency reduced BW gain in ZL and ZF rats of either gender. VAD ZL rats had lower plasma glucose, insulin, and leptin levels than VAS ZL rats. VAD-VAS3d and VAD-VAS7d rats had higher plasma glucose, insulin, and leptin levels than that in the VAD rats. The hepatic mRNA levels of Gck, Cyp26a1, Srebp-1c, Igf1, Rarb, Rxra, Rxrg, Pparg, and Ppard were lowered by VA deficiency. Refeeding of the VAS diet for 3 days restored the Gck and Cyp26a1 expressions, and for 7 days restored the Gck, Cyp26a1, Igf1, and Rxrb expressions significantly. The 7-day VA replenishment partially restored the hepatic gene expressions and metabolic changes in VAD ZL rats.

Silibinin improves L-cell mass and function through an estrogen receptor-mediated antioxidative mechanism

BackgroundSilibinin, a major component of milk thistle extract silymarin, promotes hypoglycemia by activating estrogen receptor (ER) α and β-mediated pathways in pancreatic β-cells. Glucagon-like peptide-1 (GLP-1) is the enteroendocrine peptide produced in L-cells, and it controls glucose homeostasis through multiple pathways. The effect of silibinin on L-cell mass and function is still unknown. PurposeThe protective effect of silibinin on palmitate (PA)-treated intestinal L-cell line GLUTag cells and the SHRSP•Z-Leprfa/Izm-Dmcr (SP•ZF) diabetic rat model was investigated in current study. MethodsAfter pre-incubation with 50 μM silibinin for 4 h, GLUTag cells were treated with 0.125 mM PA. MTT, Annexin V/PI apoptosis, Hoechst 33342 staining, western blot, DCFH-DA, GLP-1 ELISA, qRT-PCR and immunofluorescence analyses were undertaken to determine ER-dependent protection of silibinin against PA-induced cellular damage. The differential protein expression of GLUTag cells under different treatments was examined by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). The SP•ZF diabetic rat model was chosen for in vivo study. After 4 weeks of gastric gavage with 100 or 300 mg kg−1 of silibinin, the physiological indexes of the rats were measured. Cells expressing GLP-1, 8‑hydroxy-2′-deoxyguanosine (8-OHdG), ERα, and/or ERβ in duodenum tissues were detected by immunofluorescence. ResultsThe current study showed that the GLUTag cells preincubated with silibinin activated the transcription factor nuclear erythroid-2 like factor-2 (Nrf2)-antioxidant pathway, reduced reactive oxygen species (ROS) generation, and improved cell survival and GLP-1 content, while the antioxidative effect of silibinin was blocked by the selective ERα antagonist MPP or ERβ antagonist PHTPP in GLUTag cells. Our proteomics data further revealed that ERα or β inactivation reduced glutathione peroxide and proteins associated with endocytosis and reproduction, thus at least partially reversing the protective effect of silibinin. SP•ZF rats received silibinin treatment showed increased serum GLP-1 content and improved glucose homeostasis. Furthermore, silibinin upregulated ERα and β levels and reduced the level of 8-OHdG in GLP-1-positive cells. ConclusionsOur study showed that silibinin improved L-cell mass and function through an ER-mediated antioxidant pathway, and the proteomics analysis revealed for the first time the differential regulation of proteins by PA and silibinin in GLUTag cells.

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

Effect of resveratrol, carnitin, quercetin and aromatic amino acids on the xenobiotic metabolising and antioxidant enzymes in the liver during obesity in rats with different genotypes

The use of minor biologically active substances (BAS) in specialized products is one of the promising areas in the diet therapy for obesity and other alimentary-dependent diseases (metabolic syndrome, type 2 diabetes mellitus, etc.). The effects of the BAS in patients are often ambiguous, depending on a number of factors, one of which is the state of the organism enzyme systems (enzymes of xenobiotic metabolism and antioxidant defense), the patient's genotype, and many others. The aim was to study the effect of BAS [quercetin (Q), L-carnitine (L-Car), resveratrol (Res), aromatic amino acids tyrosine (Tyr) and tryptophan (Trp)] on the activities of phase I and II of xenobiotic-metabolising enzymes and antioxidant enzymes in rats using various in vivo models of obesity and with impaired dopamine transport. Material and methods. The activities of cytochrome P450 enzymes (CYP1A1 and CYP3A), glutathione transferase (GT), UDPglucuronosyltransferase (UDP-GT), hemoxygenase-1 (HO-1), and quinone reductase (QR) were determined by fluorimetric, spectrophotometric methods and HPLC in microsomes and cytosolic fraction of rat's liver. We used rats of outbred Wistar line, Zucker ZF line with hereditarily determined obesity and DAT-KO line with a knockout of the DAT dopamine transporter gene, which for 62 days were fed a standard balanced control or high-carbohydrate high-fat diet (30% fat by weight and 20% fructose solution instead of water) supplemented with BAS, such as Q, Res, L-Car, Tyr and Trp in doses 50, 25, 300, 1250 and 250 mg/kg of body weight respectively. Results and discussion. The presence of a DAT knockout led to a small but statistically significant decrease in the activity of GT in the liver in both homozygous and heterozygous animals. The CYP1A1 activity was significantly decreased in all carriers of the DAT knockout gene, while HO-1 activity, on the contrary, was increased, independently of the composition of the diet used. In Zucker ZF rats of all groups, in comparison with Wistar rats fed the corresponding diets, the activities of GT, UDP-GT, CYP1A1, CYP3A and QR were significantly reduced in terms of the total protein content. HO-1 activity was reduced in Zucker ZF rats in comparison with Wistar rats to a lesser extent, however, the addition of Q significantly influenced the difference between the two lines. Trp consumption led to a significant increase in GT activity in Wistar rats. In DAT-KO homozygotes this effect was insignificant, while in heterozygotes it was absent. Similarly, consumption of Trp resulted in a significant increase in CYP1A1 activity only in Wistar rats, but not in DAT-KO rats. The activity of UDP-GT under Trp intake increased only in DAT heterozygotes. The genotype significantly influenced the response of QR activity to Trp consumption, but in an ambiguous way - there was an increase in activity in heterozygotes and a decrease in homozygotes for DAT knockout. CYP1A1 activity was significantly increased in rats treated with Tyr. Conclusion. The data obtained indicate that the effect of various dietary supplements used in the treatment of obesity and metabolic syndrome on the xenobiotic-metabolising enzymes and antioxidant enzymes can have a different nature and direction depending on the genotype and the level of spontaneous physical activity and energy expenditure determined by it, which should be taken into account when approaches to personalized diet therapy of alimentary-dependent diseases are developing.

P0988THE PROTECTIVE EFFECT OF RED ORANGE AND LEMON EXTRACT (RLE) IN DIABETIC OBESE ZUCKER RATS IS RELATED TO ITS ACTION ON THE NADPH OXIDASE SUBUNIT

Abstract Background and Aims Type 2 diabetes mellitus is a pathological and chronical condition that induce a rearrangement in carbohydrate metabolism (Kaneto H et al. Antioxid Redox Signal. 2007). Even if the pathological mechanism of DN is not yet clear, NADPH oxidase (NOX) has been demonstrated to be involved in kidney ROS (Reactive Oxygen Species) generation during diabetic state (Palicz A. et al. J Biol Chem. 2001). Therefore, NOX-4, whose p22-phox subunit have shown a pivotal role in diabetes development, has been considered a potential pharmacological target in DN (Gill PS and Wilcox CS. Antioxid Redox Signal. 2006). In previous experiments performed on diabetic rat animal model with DN, we have found a beneficial effect of an extract rich in anthocyanins (ANT), members of flavonoids group (Shih PH. J Agric Food Chem. 2007). The aim of the present work was to investigate the molecular mechanisms involved in the DN and the protective effects of the new extract particularly abundant in ANT, the Red orange and Lemon Extract (RLE). Method Zucker diabetic fatty rats were divided into three groups and treated by gavage daily until 30 weeks of age, as follows: control group (ZF rats) received 1 mL of normal saline; ZDF Group (ZDF rats) received 1mL of normal saline; ZDF + RLE group (ZDF rats) treated with 90 mg/ kg of RLE dissolved in 1mL of normal saline. We examined the gene and protein expression of NOX4 catalytic subunit and its regulatory subunit p22-phox through Real Time PCR and western blot analysis. Results The expression gene levels of NOX4 significantly increased in diabetic rats ZDF, compared to control ZF (***p<0.001). Exposure of ZDF rats with RLE induces a decreased expression of NOX4 (Figure 1) compared to ZDF. On the contrary, there were not significant expression levels variations for regulatory subunit p22-phox. Western blot results are in agreement with Real Time PCR data, in fact an increase in NOX4 protein levels (*p<0.05) is evident, but no significant increase in p22-phox subunit was observed. Conclusion In conclusion, we hypothesize that the protective effect of RLE on the development of DN is related to its ability to inhibit NADPH oxidase activity and opens new perspectives for the treatment of DN.

Activation of Dorsomedial Hypothalamic Neurons Promotes Physical Activity and Decreases Food Intake and Body Weight in Zucker Fatty Rats.

Previous reports have shown that running wheel activity or voluntary exercise prevents hyperphagia and obesity in various animal models of obesity, but such effects seem only minimal in obese animals lacking leptin or leptin receptors. The mechanisms underlying this ineffectiveness remain unclear. Here, we identified the action of neuronal activation in the dorsomedial hypothalamus (DMH) in modulating physical activity, food intake and body weight using leptin receptor mutant obese Zucker (Lepr(fa), ZF) and Koletsky (Lepr(fak), SHROB) rats. Ad lib-fed SHROB rats with locked running wheels became hyperphagic and gained body weight rapidly. These alterations were not ameliorated in ad lib-fed SHROB rats with voluntary access to running wheels, but the body weight of SHROB rats with running wheel access was significantly decreased when they were pair-fed to the amounts consumed by lean controls. Determinations of hypothalamic gene expression revealed that sedentary ad lib-fed SHROB rats had increased expression of neuropeptide Y (Npy) and decreased expression of pro-opiomelanocortin (Pomc) in the arcuate nucleus (ARC). Both ARC Npy and Pomc expression were further altered under running and pair-fed conditions, indicating that both genes are appropriately regulated in response to increased energy demands or alterations caused by running activity and food restriction. Strikingly, c-Fos immunohistochemistry revealed that while voluntary running activity elevated the number of c-Fos positive cells in the DMH (particularly in the ventral and caudal subregions) of intact rats, such activation was not observed in ZF rats. Using adeno-associated virus (AAV)-mediated expression of the designer receptors hM3D(Gq) in the ventral and caudal DMH of ZF rats, we found that chemogenetic stimulation of neurons in these DMH subregions via injection of the designer drug clozapine N-oxide (CNO) significantly increased their running activity and reduced their food intake and body weight. Together, these results demonstrate that activation of ventral and caudal DMH neurons promotes physical activity and decreases food intake and body weight and suggest that intact DMH neural signaling is likely crucial for exercise-induced reductions of food intake and body weight in obese rats lacking leptin receptors.

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.

Role of Alpha Lipoic Acid on Metabolic Hepatosteatosis : An Experimental , Biochemical , and Microscopic Study

Background and aim of work: metabolic hepatosteatosis is a common serious prevalent condition in KSA. The current study investigated alpha lipoic acid (ALA) effects on hepatic lipid accumulation in severely fatty rats and secondarily on blood lipid profile. Materials and methods: sixty male Zucker rats were selectively used half of which (Lean) weighed 200±25g & others weighed 375±30g. Fatty animals were allowed free access to food and water for one week before experiment. The animals were divided into lean untreated (group I), lean ALA – managed (group II), fatty untreated (group III) and fatty ALA – managed (group IV) (fifteen animals per each group). ALA was taken orally (20 mg/kg/day) for six months. Animals were sacrificed and weighed (BW). Their liver was weighed (LW) and its portion was sliced to study its lipid content. Right tibia length (TL) was measured and LW: TL ratio was calculated. Results: ZF ALA-untreated rats showed high LW: TL ratio. ALA therapy significantly reduced BW, TL, LW and LW: TL ratio in managed ZF rats compared to untreated ones while it didn't affect these parameters significantly in lean (ZL) rats. ZF rats exhibited significant hepatosteatosis evidenced by excessive liver triglyceride (TG) and total cholesterol (TC) contents and microscopically by large cytoplasmic vacuoles. Interestingly, 6-months' ALA therapy in ZF caused significantly diminished serum triglycerides and cholesterol levels as well as diminished hepatic triglycerides and TC component in addition to decreased its lipid vacuoles compared to untreated ZF rats. The results were insignificantly changed between managed and untreated lean groups. Conclusion: our findings support effectiveness of ALA therapy in excessive hepatosteatosis and in hyperlipidemia via improving abnormal lipid metabolism.

Preliminary evaluation of β3-adrenoceptor agonist-induced 18F-FDG metabolic activity of brown adipose tissue in obese Zucker rat

BackgroundWe have investigated β3-adrenoceptor agonist mediated brown adipose tissue (BAT) activation using 18F-FDG PET/CT in Zucker lean (ZL) and obese (ZF) rats. Methods18F-FDG was injected into ZL and ZF rats pretreated with saline or agonist CL316,243 for scans. 18F-FDG metabolic activity was computed as standard uptake values. ResultsCL316,243 in ZL activated BAT up to 4-fold compared to saline, while ZF BAT was only up by 2 fold. The decreased activation was consistent with lower β3-adrenoceptor levels in ZF rats. ConclusionsThe genetically modified ZL and ZF rats may provide a useful rat model to evaluate the significance of β3-adrenoceptor agonist-induced BAT activation in obesity.

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.

Vitamin A and feeding statuses modulate the insulin‐regulated Pck1 and Srebp‐1c expression in primary hepatocytes from lean and obese rats (645.1)

The hepatic insulin resistance occurs with profound changes of the expression of genes for glucose and lipid metabolism. Previously, we compared insulin‐regulated gene expression in hepatocytes of Zucker lean (ZL) and fatty (ZF) rats. We found that insulin‐regulated Srebp‐1c and Pck1 expressions are impaired in hepatocytes of ZF, but not ZL, rats fed ad libitum, which can be corrected by fasting. As vitamin A (VA) deficiency causes reduction of body mass and food intake in rats, we set up a pair‐feeding experiment to study the effects of energy intake and VA status on the insulin‐regulated gene expression. Rats were fed a VA deficient (VAD) diet ad libitum (VAD‐AD) for 8 weeks. An isocaloric VA sufficient (VAS) diet equal to the amount of the VAD diet taken by the VAD‐AD rat in 24‐hour was given to the pair‐fed rats in two ways. On the last day, VAS‐PF‐AD rat was allowed to ingest the VAS diet ad libitum to avoid the fasting state. VAS‐PF‐4M rat was given one fourth of the daily ration every 6 hours. Here, we report that the insulin‐ and retinoic acid (RA)‐regulated Srebp1c and Pck1 expressions were impaired in hepatocytes of ZF rats fed ad libitum, confirming our previous results. The impairment was partially corrected in hepatocytes of ZF rats fed the VAD diet ad libitum. The ZL and ZF VAS‐PF‐AD or ‐4M rats had higher body mass and feed efficiency than their VAD‐AD controls before the reduction of food intake occurred. The VAS‐PF rats overate when enough diet was present at the end of the pair‐feeding paradigm. Insulin‐inhibited Pck1 (VAS‐PF‐AD ZL rats) and induced Srebp1c (VAS‐PF‐AD ZL and ZF rats) expressions ware partially impaired. The expressions of genes for hepatic VA metabolism were altered in those rats. VA and feeding statuses modulate the insulin‐regulated gene expression.Grant Funding Source: Supported by AHA

Up‐Regulation of Stearoyl‐CoA Desaturase 1 Increases Liver MUFA Content in Obese Zucker but Not Goto‐Kakizaki Rats

The Goto-Kakizaki (GK) rat is an animal model for spontaneous-onset, non-obese type 2 diabetes. Despite abundant evidence about disorders in metabolism, little information is available about fatty acid metabolism in the liver of GK rats. This study aimed to investigate the characteristics of the fatty acid profile, particularly MUFA, and the mechanism underlying the alterations in fatty acid profiles in the liver of GK rats. The activities of enzymes that participate in the biosynthesis of MUFA, expressions of genes encoding these enzymes, and the fatty acid profile in the liver were compared with those of obese Zucker (fa/fa) (ZF) rats, which are obese and non-diabetic. Stearoyl-CoA desaturase (SCD) activity and SCD1 gene expression were considerably up-regulated in GK rats, and these levels were largely comparable to those in ZF rats. However, the proportions and contents of oleic acid and palmitoleic acid were very low considering the highly elevated activity of SCD in the liver of GK rats, when compared with ZF rats. Palmitoyl-CoA chain elongation (PCE) activity and fatty acid elongase (Elovl6) gene expression were markedly up-regulated in ZF rats, whereas PCE activity was up-regulated much less and Elovl6 gene expression was unchanged in GK rats. These results suggest the possibility that up-regulation of gene expression of Elovl6 along with SCD1 is indispensable to elevate the proportions and contents of oleic acid in the liver.

Vitamin A status affects obesity development and hepatic expression of key genes for fuel metabolism in Zucker fatty rats

We hypothesized that vitamin A (VA) status may affect obesity development. Male Zucker lean (ZL) and fatty (ZF) rats after weaning were fed a synthetic VA deficient (VAD) or VA sufficient (VAS) diet for 8 weeks before their plasma parameters and hepatic genes' expression were analyzed. The body mass (BM) of ZL or ZF rats fed the VAD diet was lower than that of their corresponding controls fed the VAS diet at 5 or 2 weeks, respectively. The VAD ZL and ZF rats had less food intake than the VAS rats after 5 weeks. The VAD ZL and ZF rats had lower plasma glucose, triglyceride, insulin, and leptin levels, as well as lower liver glycogen content, net mass of epididymal fat, and liver/BM and epididymal fat/BM ratios (ZL only) than their respective VAS controls. VAD rats had lower hepatic Cyp26a1, Srebp-1c, Fas, Scd1, Me1, Gck, and Pklr (ZL and ZF); and higher Igfbp1 (ZL and ZF), Pck1(ZF only), and G6pc (ZF only) mRNA levels than their respective VAS controls. We conclude that ZL and ZF rats responded differently to dietary VA deficiency. VA status affected obesity development and altered the expression of hepatic genes for fuel metabolism in ZF rats. The mechanisms will help us to combat metabolic diseases.

Effects of vitamin A status on the responses of the hepatic genes’ expression to insulin and retinoic acid in primary hepatocytes from Zucker lean and fatty rats

Insulin‐mediated regulation of hepatic gene expression plays a key role in glucose and lipid homeostasis. This study was aimed to examine the hepatic gene expression in response to insulin and retinoic acid (RA) treatments in primary hepatocytes from Zucker rats of different vitamin A (VA) status. Primary hepatocytes were isolated from Zucker lean (ZL) or fatty (ZF) rats fed either vitamin A sufficient (VAS) or deficient diet (VAD) for 8 weeks after weaning. The mRNA levels of Gck, Pck1, Srebp‐1c and Pklr after insulin and RA stimulation for 6 hours were determined using real‐time PCR. In hepatocytes of ZL rats on either diet, insulin synergized with RA to induce the expression of Gck and Srebp‐1c. This was impaired in hepatocytes of ZF rats on VAS, but partially recovered in those on VAD diet. In lean hepatocytes, the Pck1 expression was suppressed by insulin and induced by RA respectively. The insulin‐suppressed Pck1 expression was impaired in hepatocytes from ZF rats fed VAS diet and partially restored in that fed VAD diet. The RA‐induced Pck1 expression was normal in hepatocytes from ZF rats fed VAS, and was enhanced in that fed VAD. The Pklr expression was similar between ZL and ZF and not affected by VA status. We conclude that VA status modulate the insulin‐ and RA‐regulated hepatic gene expression in hepatocytes of ZF rats. It suggests that micronutrients contribute to the hepatic insulin response.Grant Funding Source: AHA

Effects of retinoid catabolism on the gene expression in primary rat hepatocytes and hepatoma cells

Vitamin A (retinol, ROL) is reversibly oxidized into retinal (RAL) by retinol dehydrogenases; retinal is irreversibly oxidized by retinal dehydrogenases into retinoic acid (RA) in cells. RA regulates gene expression via activation of retinoic acid receptor (RAR) and retinoid X receptor (RXR). We have shown that retinoids synergize with insulin to induce Srebp‐1c expression in primary rat hepatocytes. Therefore, we hypothesize that RA production controls the regulation of hepatic gene expression.The expression of Cyp26a1 (an RA responsive gene) was examined using real‐time PCR and used as a marker for RA production from ROL and RAL in primary hepatocytes and hepatoma cells. Here, we show that retinol and retinal dose‐dependently induced Cyp26a1 expression via activation of RAR and RXR in those cells. The expression levels of Rdh2, Rdh10, Raldh1, and Raldh3 (major retinoid catabolic enzymes) were examined in hepatocytes from Zucker lean (ZL) and fatty (ZF) rats. Only Rdh2 and Raldh1 were expressed at comparable level as that of 36B4 gene (the invariable control gene) in hepatocytes and hepatoma cells. In addition, the hepatocytes from ZF rats had higher expression level of Raldh1 than that from ZL rats.We conclude that hepatocytes actively metabolize retinol into RA, which directly regulate gene expression. The elevated Raldh1 expression in ZF hepatocytes may contribute to the altered lipid metabolism in ZF liver.Sponsor: Guoxun Chen, 1215 W. Cumberland Ave. 229 Jessie Harris Building, Knoxville, TN 37996Grant Funding Source : AHA

Progression of Obesity‐Related Heart Dysfunction in Two Rat Models

The prevalence of obesity in the US has increased substantially over the past decades, with upwards of 33.8% of the population obese and 68% overweight. In childhood, 11.7% – 14.6% of preschoolers in developed countries and 16.4% of adolescents in the US are considered obese. Several animal models have been utilized to study the impact of obesity on the heart. In rats, the most prevalent models are the Zucker Fatty rats (ZF), a genetically modified rat with a leptin receptor deficiency, and normal Sprague Dawley rats (SD) fed a high fat diet. In the present study, we examined echocardiographic indices for each model in order to evaluate heart function. Values for IVRT/HR were significantly higher in ZF rats compared to Zucker Lean rats (0.058±0.010 ms/bpm vs, 0.045±0.009 ms/bpm, p<0.05) at 19 weeks of age, however the values for SD rats were not significantly different after eating a high fat diet for 10 weeks. E/A ratio was not significantly different in ZF rats compared to lean controls, however was significantly decreased in SD rats fed a high fat diet compared to rats fed a normal diet (0.98 vs. 1.11; p<0.05). Diastolic dysfunction was evident in both models, although the values for the ZF rats indicated less relative diastolic impairment than the SD rats fed a high fat diet. Continued work will examine the progression and mechanistic differences in these models, and their relevance to human obesity.

Differential Adipose Tissue Inflammatory State in Obese Nondiabetic Zucker Fatty Rats Compared to Obese Diabetic Zucker Diabetic Fatty Rats

Adipose tissue (AT) inflammation is linked to the pathogenesis of diabetes in obesity. Here, we compare the AT inflammatory state of 2 animal models of obesity and obesity plus diabetes, respectively. Obese nondiabetic ZF rats exhibited a trend towards increased proportions of CD11b positive cells in the adipose tissue stroma vascular fraction suggesting a state of increased AT inflammation compared to their lean littermates, but no alterations in systemic inflammatory parameters. In contrast, obese diabetic ZDF rats exhibited systemic as well as local AT inflammation with elevated levels of circulating Regulated upon Activation, Normal T-cell Expressed and Secreted Protein (Rantes), interleukin 1β (IL-1β) and monocyte chemotactic protein 1 (MCP-1), and an increased infiltration of adipose tissue CD11b positive cells. Our data provide a novel phenotypic characterisation of 2 common metabolic animal models and suggest an association of obesity with local inflammation in adipose tissue, and an association of diabetes with local inflammation in adipose tissue plus systemic inflammation. AT inflammation in obesity might therefore initiate a process that above a certain limits finally results in systemic inflammation and diabetes.

Protection by Red Wine Polyphenols against Metabolic and Cardiovascular Alterations Associated with Obesity: A Possible Link with Estrogen Alpha Receptor

Epidemiological studies report an inverse association between dietary flavonoid consumption and mortality from cardiovascular and metabolic diseases. Red wine contains a wide variety of polyphenols (RWPC), which mainly derive from grape solids (skin and seeds) and can be divided into two classes, flavonoids and non-flavonoids. RWPC exert numerous effects including antioxidant and free radical properties, anti-platelet aggregating and anti-thrombotic activities. Moreover, RWPC are powerful vasodilators and contribute to the preservation of the integrity of the endothelium and to the inhibition of smooth muscle cell proliferation and migration. All these effects of red wine may interfere with atherosclerotic plaque development and stability, vascular thrombosis and occlusion and explain the prevention of ischemic heart disease, stroke and metabolic diseases observed in different experimental models. With regard to obesity, we studied the effects of dietary supplementation of RWPC in Zucker fatty rats (ZF). We show that RWPC lead to an improvement of glucose and lipid metabolism in ZF rats, enhances cardiac function and decreases peripheral resistance. In addition, RWPC improve endothelium-dependent relaxation in both aorta and small mesenteric arteries from ZF rats, an effect accompanied by an increase of vascular nitric oxide (NO) production and a decrease of superoxide anions release, resulting in an increase of NO bioavailability. Thus, RWPC lead to an overall improvement in obesity-associated alterations, including glucose and lipid metabolism, as well as endothelial and cardiac functions. A molecular basis of the effect of RWPC has been provided on the endothelium, indentifying the alpha isoform of the estrogen receptor (ERα) as the key receptor, or at least one of those transducing vascular effects exerted by these compounds, particularly delphinidin with respect to NO production. These results are of importance inasmuch as RWPC, by interacting with ERα, can activate other targets such the sirtuin-1-adenosine monophosphate protein kinase network, both of which are amongst the most potent regulators of cellular metabolism, including vascular cells, adipocytes and hepatocytes. Altogether, the available evidence indicates that RWPC might be of therapeutic benefit for cardiovascular and metabolic protections, although prospective controlled clinical trials are still needed.

Protective Role of Pomegranate on Fatty Liver in Obesity: An Experimental Chemical & Histopathological Study

Aim of the work: Fatty liver is the most common cause of abnormal liver function tests. Since obesity and fatty liver can result in serious drawbacks and represent a considerable incidence rate in Saudi Arabia. This country is a famous one of pomegranate. This study aimed at investigating the ameliorating effects of pomegranate on hepatic lipid accumulation in rats with severe fatty liverin order to provide a cheap natural protective medicine since this health problem can affect the entire world economy. Material and methods: Twenty-four male Zucker rats aged 13–15 weeks were used. Animals were allowed free access to food and water for 1 week before starting the experiment. According to the body weight (BW), plasma triglycerides (TG) and total cholesterol (TC) levels were measured before treatment. The animals were divided into lean control, lean pomegranate (PGF)-treated, fatty control and fatty PGF- treated groups (six animals per group). Finally, the animals were weighed after a prompt dislocation of their neck vertebra. Then, the liver was rapidly excised, washed with saline on ice and weighed. In addition, one part of it was cut into slices for studies of lipid determination analysis. The right tibia length (TL) was measured for calculation of the ratio of liver weight to TL. A portion of the liver was homogenized and the lipids were extracted with isopropanol. Triglycerides and TC contents in the supernatant were determined. Results: Fatty rats (ZF) had shorten TL but their BWs were increased compared to lean rats. Also, ZF rats showed hepatomegaly reflected by increased liver weight and increased ratio of liver weight to TL. Treatment with PGF extract did not change BW and TL, but it reduced liver weight and the ratio in ZF rats. PGF did not affect these parameters in lean ( ZL) rats. Fatty rats exhibited severe hepatic steatosis, reflected by a marked increase in hepatic TG as well as hepatic TC contents and fatty droplets visualized by either lightmicroscopy as large vacuoles occupying the cytoplasm of hepatocytes pushing their nuclei to the periphery against the cell membrane giving signet-ring appearance or by electron microscopy as membrane-bound inclusions closely near the endoplasmic reticulum with indentation of the nuclei of the affected cells. Interestingly, 6-week treatment of fatty rats with PGF extract reduced hepatic TG contents and fatty droplets whereas it did not alter hepatic TC contents. This treatment showed minimal effects in ZL rats. Conclusion: These findings are potentially important for supporting the effectiveness of pomegranate in the treatment of obesity - induced fatty liver disease through the improvement of abnormal lipid metabolism.