Lean Female Rats Research Articles

Fecal Microbiota Transplantation During Lactation Programs the Metabolism of Adult Wistar Rats in a Sex-specific Way

The intestinal microbiota is involved in many physiological processes. However, the effects of microbiota in metabolic programming still unknow. We evaluated whether the transplantation of fecal microbiota during early life can program health or disease during adulthood in a model of lean and obese male and female Wistar rats. Parental obesity were induced using a small litter (SL, 3 pups/dam) model. At 90 d old, normal litter (NL, 9 pups/dam) and SL males and females (parents) from different litters were mated: NL male vs. NL female; SL male vs. SL female. After birth, male and female offspring rats were also standardized in normal litters or small litters . From the 10th until 25th d of life, the NL and SL male and female offspring received via gavage of a solution containing the diluted feces of the opposite dam (fecal microbiota, M) or saline solution (S). At 90 d of age, biometric and biochemical parameters were assessed. NLM male rats transplanted with obese microbiota showed increased body weight, and fat pad deposition, hyperinsulinemia, glucose intolerance and dyslipidemia. SLM male rats transplanted with lean microbiota had decreased retroperitoneal and mesenteric fat, triglycerides and VLDL levels and improvement of glucose tolerance. Despite SLM female rats showed higher visceral fat, microbiota transplantation in female rats caused no changes in these parameters compared with control groups. Fecal microbiota transplantation during lactation induces long-term effects on the metabolism of male Wistar rats. However, female rats were resistant to metabolic alterations caused by the treatment.

Abstract 740: Obesity promotes liver pro-oxidative and DNA damage in mammary tumor of Zucker rats

Abstract Obesity is a major health problem in the US and world. Obesity is associated with the risk of cardiovascular disease, type 2 diabetes, and certain types of cancers such as breast cancer. Our laboratory reported that obesity increased mammary tumor model using obese Zucker rat model. However, the effects of obesity in the liver in regard to oxidative stress and DNA reaction is less known. In present study, we used a DMBA induced-mammary tumor model to further investigate the effects of obesity on creation of pro-oxidative environment in liver and oxidative DNA reaction. Lean and obese female rats were at 50 days of age received 7,12-dimethylbenz(α)anthracene (DMBA) and were sacrificed 20 weeks later. HPLC-ECD, HPLC-UV and LC-MS were used. Obesity capable significantly modify methylation status in liver caused higher (p < 0.001) SAM/SAH ratio (know as “methylation ratio”) and increased (p < 0.05) global DNA methylation. Also, obesity decrease GSH/GSSG ratio (known as “oxidative stress ratio”) with p < 0.0001 inducing higher oxidative stress and as results, increase presence both oxidized guanosine (8-OH-Guanosine) with p < 0.001 and oxidized 5metylCytosine (5hmCytosine) p<0.04 in liver DNA. 8-OH-Guanosine/5hmCytosine ratio was not significantly affected. Obesity caused significant oxidative stress in liver, causing oxidative DNA damage and change of DNA methylation pattern. These changes may contribute to the development of fatty liver in breast cancer models. Citation Format: Reza Hakkak, Stepan Melnyk, Soheila Korourian. Obesity promotes liver pro-oxidative and DNA damage in mammary tumor of Zucker rats [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 740.

Effects of a Low Dose of Caffeine Alone or as Part of a Green Coffee Extract, in a Rat Dietary Model of Lean Non-Alcoholic Fatty Liver Disease without Inflammation.

Non-alcoholic fatty liver disease is a highly prevalent condition without specific pharmacological treatment, characterized in the initial stages by hepatic steatosis. It was suggested that lipid infiltration in the liver might be reduced by caffeine through anti-inflammatory, antioxidative, and fatty acid metabolism-related mechanisms. We investigated the effects of caffeine (CAF) and green coffee extract (GCE) on hepatic lipids in lean female rats with steatosis. For three months, female Sprague-Dawley rats were fed a standard diet or a cocoa butter-based high-fat diet plus 10% liquid fructose. In the last month, the high-fat diet was supplemented or not with CAF or a GCE, providing 5 mg/kg of CAF. Plasma lipid levels and the hepatic expression of molecules involved in lipid metabolism were determined. Lipidomic analysis was performed in liver samples. The diet caused hepatic steatosis without obesity, inflammation, endoplasmic reticulum stress, or hepatic insulin resistance. Neither CAF nor GCE alleviated hepatic steatosis, but GCE-treated rats showed lower hepatic triglyceride levels compared to the CAF group. The GCE effects could be related to reductions of hepatic (i) mTOR phosphorylation, leading to higher nuclear lipin-1 levels and limiting lipogenic gene expression; (ii) diacylglycerol levels; (iii) hexosylceramide/ceramide ratios; and (iv) very-low-density lipoprotein receptor expression. In conclusion, a low dose of CAF did not reduce hepatic steatosis in lean female rats, but the same dose provided as a green coffee extract led to lower liver triglyceride levels.

Evaluation of the In Vitro Damage Caused by Lipid Factors on Stem Cells from a Female Rat Model of Type 2 Diabetes/Obesity and Stress Urinary Incontinence.

Human stem cell therapy for type 2 diabetes/obesity (T2D/O) complications is performed with stem cell autografts, exposed to the noxious T2D/O milieu, often with suboptimal results. We showed in the Obese Zucker (OZ) rat model of T2D/O that when their muscle-derived stem cells (MDSC) were from long-term T2D/O male rats, their repair efficacy for erectile dysfunction was impaired and were imprinted with abnormal gene- and miR-global transcriptional signatures (GTS). The damage was reproduced in vitro by short-term exposure of normal MDSC to dyslipidemic serum, causing altered miR-GTS, fat infiltration, apoptosis, impaired scratch healing, and myostatin overexpression. Similar in vitro alterations occurred with their normal counterparts (ZF4-SC) from the T2D/O rat model for female stress urinary incontinence, and with ZL4-SC from non-T2D/O lean female rats. In the current work we studied the in vitro effects of cholesterol and Na palmitate as lipid factors on ZF4-SC and ZL4-SC. A damage partially resembling the one caused by the female dyslipidemic serum was found, but differing between both lipid factors, so that each one appears to contribute specifically to the stem cell damaging effects of dyslipidemic serum in vitro and T2D/O in vivo, irrespective of gender. These results also confirm the miR-GTS biomarker value for MDSC damage.

Epigenomic Regulation of SATB2 Links Maternal Obesity to Impaired Osteoblast Differentiation (P11-034-19)

ObjectivesNutritional status during intrauterine and/or early postnatal life has substantial influences on adult offspring health. However, evidence on the impact of high fat diet (HFD)-induced maternal obesity on regulation of fetal bone development is sparse. Thus, we investigated the effects of maternal obesity in rodent and human cells on epigenetic regulation of osteoblast differentiation. MethodsFirst, female Sprague-Dawley rats were fed either a low-fat AIN-93G control diet or a high fat diet (HFD) (45% fat calories) for 10 wk starting at 6 wk of age. Lean (from control diet) and obese (from HFD) female rats were then time-impregnated (n = 6 per group) by control diet fed male rats. At gestational day 18.5 (E18.5), all fetuses were taken and embryonic osteogenic calvarial cells (EOCCs) were isolated. Second, human osteo-progenitors of mesenchymal stem cells were isolated from umbilical cord following delivery from pregnant mothers. ResultsWe found epigenetic regulation of polycomb-regulated gene Ezh2 (Enhancer of zeste homolog 2) in embryonic rats from HFD obese rat dams. Increased enrichment of repressive histone mark H3K27me3 on the gene body of SATB2 (ChIP Seq analysis) was associated with aberrant differentiation of EOCCs to mature osteoblasts. Knocking down Ezh2 in EOCCs and ST2 cells increased SATB2 expression; on the other hand, Ezh2 overexpression in EOCCs and ST2 cells decreased SATB2 expression. These data were consistent with ChIP experimental results showing strong association between H3K27me3, Ezh2 and SATB2. Second, human mesenchymal stem cells (MSCs) from umbilical cord (UC) were isolated following delivery from obese/overweight (pre-pregnancy BMI ≥ 25 kg/m2) and control (pre-pregnancy BMI between 19–25 kg/m2) pregnant mothers. We found: 1) UC-MSCs from pregnant obese/overweight mothers showed increased Ezh2 expression and decreased SATB2 mRNA expression, which was concurrent lower osteoblastogenesis potential in EOCCs; 2) ChIP experiments using H3K27me3 IP (immune-precipitation) showed significant association between H3K27me3, Ezh2 and SATB2. ConclusionsThese findings indicate maternal HFD-induced obesity-associated decrease of fetal pre-osteoblastic cell differentiation is under epigenetic control through SATB2 expression. Funding SourcesSupported by USDA-ARS Project.

Galectin‐3 Levels in Hepatic Tissue from Lean and Obese Zucker Rats

Obesity and metabolic syndrome are of major concern for physicians worldwide. Galectin‐3 is a β‐galactoside‐binding protein that has been shown to have regulatory functions in inflammatory responses as well as fibrosis of the liver, lung, heart and blood vessels. Galectin‐3 is widely expressed in multiple types of immune cells as well as epithelial and endothelial cells, and its properties have been explored as a therapeutic target for nonalcoholic fatty liver disease liver fibrosis, heart disease and other obesity‐associated diseases. We previously observed elevation of galectin‐3 in mesenteric fat from male and female 12‐week‐old obese Zucker rats compared to their lean counterparts. We hypothesized that galectin‐3 expression levels would be elevated in the liver in obese animals. Livers were collected from 20 Zucker rats (5 male, 5 female lean rats and 5 male, 5 female obese Zucker rats) for histology and protein analysis. After protein lysates were collected and total protein levels were normalized, galectin‐3 levels were quantified with the Wes Simple Western capillary electrophoresis assay. The results showed that livers of obese rats of both sexes display typical fatty liver pathology and have a significantly higher level of Galectin‐3 expression. The results show that increased galectin‐3 expression accompanies fatty liver in obese rats. Galectin‐3 may represent a future target for preventing tissue dysfunction in obesity and metabolic syndromeSupport or Funding InformationAPS Frontiers in Physiology FellowshipThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The RELATION OF SERUM IRISIN LEVEL WITH METABOLIC AND HORMONAL CHANGES IN RAT MODEL OF POLYCYSTIC OVARY

ABSTRACT Background : Polycystic ovary syndrome (PCOS) is one of the leading causes of infertility in female. It is usually associated with insulin resistance (IR), obesity, hyperlipidemia, and type II diabetes. Irisin is amyokine which increases energy expenditure and protects against insulin resistance and obesity. There were controversial studies about the levels of irisin and its relation to hormonal and metabolic changes in PCOS. This study was designed to estimate serum level of irisin in letrozole-induced PCOS in both lean and obese female rats, and to study the link between serum irisin level and some metabolic and hormonal parameters in PCOS. Material and Methods: This study was conducted on 48 young virgin female of local strain albino rats. They were randomly divided into 2 equal groups: Group I: lean group were fed on normal laboratory chow diet and Group II: A high fat diet induced obesity group and each group was subdivided into group A (control) and group B (letrozole induced PCOS). Fasting serum irisin, Lutenizig hormone (LH), Folliclular stimulating hormone (FSH), testosterone, estradiol, progesterone, insulin, glucose, triglycerides, cholesterol, LDL and HDL levels were estimated. In addition, body mass index (BMI) and Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) were calculated in all groups at the end of the experiment. Results : Serum irisin level was elevated in both lean and obese PCOS groups when compared to both lean and obese control groups. In addition, obese PCOS group had significant higher level of irisin when compared to lean PCOS group. Conclusion: Serum irisin level was positively correlated with BMI, HDL and testosterone levels and negatively correlated with HOMA- IR, insulin, cholesterol, triglycerides and LDL levels. Irisin may have a role in the development of polycystic ovary syndrome. Key words: Irisin – PCOS- lean – obese.

Intergenerational Influence of Paternal Obesity on Metabolic and Reproductive Health Parameters of the Offspring: Male-Preferential Impact and Involvement of Kiss1-Mediated Pathways.

Obesity and its comorbidities are reaching epidemic proportions worldwide. Maternal obesity is known to predispose the offspring to metabolic disorders, independently of genetic inheritance. This intergenerational transmission has also been suggested for paternal obesity, with a potential negative impact on the metabolic and, eventually, reproductive health of the offspring, likely via epigenetic changes in spermatozoa. However, the neuroendocrine component of such phenomenon and whether paternal obesity sensitizes the offspring to the disturbances induced by high-fat diet (HFD) remain poorly defined. We report in this work the metabolic and reproductive impact of HFD in the offspring from obese fathers, with attention to potential sex differences and alterations of hypothalamic Kiss1 system. Lean and obese male rats were mated with lean virgin female rats; male and female offspring were fed HFD from weaning onward and analyzed at adulthood. The increases in body weight and leptin levels, but not glucose intolerance, induced by HFD were significantly augmented in the male, but not female, offspring from obese fathers. Paternal obesity caused a decrease in luteinizing hormone (LH) levels and exacerbated the drop in circulating testosterone and gene expression of its key biosynthetic enzymes caused by HFD in the male offspring. LH responses to central kisspeptin-10 administration were also suppressed in HFD males from obese fathers. In contrast, paternal obesity did not significantly alter gonadotropin levels in the female offspring fed HFD, although these females displayed reduced LH responses to kisspeptin-10. Our findings suggest that HFD-induced metabolic and reproductive disturbances are exacerbated by paternal obesity preferentially in males, whereas kisspeptin effects are affected in both sexes.

Effects of Obesity on Pro-Oxidative Conditions and DNA Damage in Liver of DMBA-Induced Mammary Carcinogenesis Models.

The prevalence of the overweight and obesity is on the rise worldwide. Obesity can increase the risk of certain cancers and liver steatosis development. Previously, we reported that obesity increased liver steatosis in a mammary tumor model, but little is known about the effects of obesity in the liver in regard to global DNA methylation, DNA damage, and oxidative/nitrosative stress. Using a mammary tumor model, we investigated the effects of obesity on oxidative stress and DNA reaction. Five-week-old lean and obese female rats were used. At 50 days of age, all rats received 7,12-dimethylbenz(α)anthracene (DMBA) and were sacrificed 155 days later. HPLC with electrochemical and ultraviolet detection and LC-MS were used. Obesity caused higher (p < 0.0004) methionine levels, had no effect (p < 0.055) on SAM levels, caused lower (p < 0.0005) SAH levels, caused higher (p < 0.0005) SAM/SAH ratios, and increased (p < 0.02) global DNA methylation. Levels of free reduced GSH were not significantly lower (p < 0.08), but free oxidized GSSG was higher (p < 0.002) in obese rats. The GSH/GSSG ratio was lower (p < 0.0001), and oxidized guanosine was higher (p < 0.002) in DNA of obese rats compared to lean rats. Obesity caused significant oxidative/nitrosative stress, oxidative DNA damage, and change of DNA methylation pattern in the liver, and these changes may contribute to the development of liver steatosis in breast cancer models.

Maternal obesity is associated with ovarian inflammation and upregulation of early growth response factor 1.

Obesity impairs reproductive functions through multiple mechanisms, possibly through disruption of ovarian function. We hypothesized that increased adiposity will lead to a proinflammatory gene signature and upregulation of Egr-1 protein in ovaries from obese (OB; n = 7) compared with lean (LN; n = 10) female Sprague-Dawley rats during the peri-implantation period at 4.5 days postcoitus (dpc). Obesity was induced by overfeeding (40% excess calories for 28 days) via total enteral nutrition prior to mating. OB dams had higher body weight (P < 0.001), greater fat mass (P < 0.001), and reduced lean mass (P < 0.05) and developed metabolic dysfunction with elevated serum lipids, insulin, leptin, and CCL2 (P < 0.05) compared with LN dams. Microarray analyses identified 284 differentially expressed genes between ovaries from LN vs. OB dams (±1.3 fold, P < 0.05). RT-qPCR confirmed a decrease in expression of glucose transporters GLUT4 and GLUT9 and elevation of proinflammatory genes, including CCL2, CXCL10, CXCL11, CCR2, CXCR1, and TNFα in ovaries from OB compared with LN (P < 0.05). Protein levels of PI3K and phosphorylated Akt were significantly decreased (P < 0.05), whereas nuclear levels of Egr-1 (P < 0.05) were increased in OB compared with LN ovaries. Moreover, Egr-1 was localized to granulosa cells, with the highest expression in cumulus cells of preovulatory follicles. mRNA expression of VCAN, AURKB, and PLAT (P < 0.05) correlated with %visceral fat weight (r = 0.51, -0.77, and -0.57, respectively, P ≤ 0.05), suggesting alterations in ovarian function with obesity. In summary, maternal obesity led to an upregulation of inflammatory genes and Egr-1 expression in peri-implantation ovarian tissue and a concurrent downregulation of GLUTs and Akt and PI3K protein levels.

Feeding butter with elevated content of trans-10, cis-12 conjugated linoleic acid to obese-prone rats impairs glucose and insulin tolerance.

BackgroundWe recently demonstrated that feeding a natural CLAt10,c12-enriched butter to lean female rats resulted in small, but significant increases in fasting glucose and insulin concentrations, and impaired insulin tolerance. Our goal was to extend these findings by utilizing the diabetes-prone female fatty Zucker rat. Rats were fed custom diets containing 45 % kcal of fat derived from control and CLAt10,c12-enriched butter for 8 weeks.MethodsCLA t10,c12-enriched butter was prepared from milk collected from cows fed a high fermentable carbohydrate diet to create subacute rumen acidosis (SARA); control (non-SARA) butter was collected from cows fed a low grain diet. Female fatty Zucker rats (10 weeks old) were randomly assigned to one of four diet treatments: i) low fat (10 % kcal), ii) 45 % kcal lard, iii) 45 % kcal SARA butter, or iv) 45 % kcal non-SARA butter. A low fat fed lean Zucker group was used as a control group. After 8 weeks, i) glucose and insulin tolerance tests, ii) insulin signaling in muscle, adipose and liver, and iii) metabolic caging measurements were performed.ResultsGlucose and insulin tolerance were significantly impaired in all fatty Zucker groups, but to the greatest extent in the LARD and SARA conditions. Insulin signaling (AKT phosphorylation) was impaired in muscle, visceral (perigonadal) adipose tissue and liver in fatty Zucker rats, but was generally similar across dietary groups. Physical activity, oxygen consumption, food intake and weight gain were also similar amongst the various fatty Zucker groups.ConclusionsIncreasing the consumption of a food naturally enriched with CLAt10,c12 significantly worsens glucose and insulin tolerance in a diabetes-prone rodent model. This outcome is not explained by changes in tissue insulin signaling, physical activity, energy expenditure, food intake or body mass.

Abstract P636: Sex Differences in Cardioprotective AT2R Expression in Diabetic Rats and Its Correlation with Myocardial Damage

Diabetes mellitus (DM) is an independent risk factor for cardiovascular disease (CVD). Healthy, young women are protected from CVD, while diabetic women are more susceptible to CVD compared to age-matched diabetic men and non-diabetic women. Underlying mechanisms for this sex difference in CVD are not fully elucidated. The angiotensin II type 2 receptor (AT2R) is a member of the protective, vasodilative arm of the renin angiotensin system. The Agtr2 gene that codes for AT2R is X-linked, and increased Agtr2 expression is reported in female vasculature of rodent models. We hypothesized that a sex difference might exist in DM-associated regulation of cardiac AT2R expression. To test this, we used hyperglycemic, male and female Zucker diabetic fatty (ZDF) rats and age- and sex-matched normoglycemic Zucker lean (ZL) rats. The male ZDF (ZDF-M) rat is an established model of type 2 DM. We have reported previously that hyperglycemic, female ZDF (ZDF-F) rats had the highest body fat and lowest lean muscle mass compared to male and female lean rats (ZL-M, ZL-F) and ZDF-M. Cardiac Agtr2 expression was measured by qRT-PCR at 5-months, cardiac function by echocardiography was compared at 3- and 5-months, and histopathology of cardiac tissue was assessed at 5-months. ZL-F had a nearly 2-fold increase of Agtr2 compared to ZL-M (p&lt;0.01). Relative to lean controls, ZDF-M had no significant change in Agtr2, while ZDF-F exhibited ~60% suppression (Rq=0.42) of Agtr2 (p&lt;0.001). Echocardiography data revealed evidence of compensated systolic function in all groups since fractional shortening was &gt;50% at both ages, while heart rate and stroke volume were similar. However, diastolic dysfunction was observed in both ZDF-F and ZDF-M, relative to their lean counterparts, due to increased isovolumic relaxation time and decreased early:late ventricular filling ratio (E/A). ZDF-F exhibited the highest cardiomyocyte hypertrophy (≥35% over ZL-F, ZL-M and ZDF-M). Both ZDF and ZDF-M showed mitochondrial clustering and disrupted spatial orientation of mitochondria relative to the sarcomere (assessed by TEM). Based on our results, we propose that myocardial remodeling, diastolic dysfunction and loss of cardioprotective AT2R may underlie greater susceptibility of diabetic females to CVD.

Sex Differences in the Progression of Diabetes‐associated Cardiac Pathology

Type 2 Diabetes Mellitus (T2DM) is a major contributor to cardiovascular disease (CVD). We hypothesized that there are significant sex‐related differences in the progression of T2DM‐induced cardiac pathology in pre‐menopausal diabetic female rats and age‐matched diabetic male rats. These differences were assessed in 3 study groups: ZDF male rats (ZDF‐M) and lean female rats (L‐F) on Purina 5008 diet, and ZDF female (ZDF‐F) rats on Research Diets D12468 (to induce T2DM) from the age of 6‐20 weeks. ZDF‐M went from having a ~26 fold higher fasting plasma insulin (INS) than L‐F at 9‐weeks, to &lt;2 fold at 20‐weeks (p&lt;0.05). In contrast, in ZDF‐F, INS was ~ 64 fold higher than L‐F at 15‐weeks and was still ~50 fold higher than L‐F and ~21 fold higher than ZDF‐M at 20‐weeks (p&lt;0.05). Hyperglycemia was lower in ZDF‐F than ZDF‐M at all ages, but ZDF‐F had lower plasma HDL and higher LDL levels and higher % body fat and lower % lean muscle mass (as determined by EchoMRI) compared to ZDF‐M and L‐F (p&lt;0.05). Cardiac echocardiography showed heart rate was significantly different among all three groups with ZDF‐F being greatest and ZDF‐M lowest. However, only ZDF‐M displayed cardiac diastolic dysfunction, indicated by dilatory remodeling, increased isovolumic relaxation time, increased late diastolic filling (A) and decreased E/A and E'/A' ratios. ZDF‐M had the highest 24‐hr urine clearance of Na+, K+, glucose and protein (p&lt;0.05).ConclusionT2DM had different effects on ZDF‐F and ZDF‐M. ZDF‐F had a quicker loss of lean muscle mass and HDL despite longer hyperinsulinemic stage of T2DM, whereas ZDF‐M had higher HDL, but worse cardiac dysfunction.Funding: LSME, UM‐Columbia (LP), NIH 1R01HL118376‐01(LP), R01HL112998 (CAE).

Abstract 629: Gender Differences In The Expression Of Cardioprotective Myeloid Cell Leukemia 1 (Mcl-1) In Diabetic Male And Female Rats

Diabetes Mellitus (DM) is an important risk factor for cardiovascular disease (CVD) and young, diabetic women are more susceptible to CVD compared to age-matched men and non-diabetic women. It is unclear how the progression of DM differentially affects cardiac pathology in males and females. In this context, recent studies have shown that myeloid cell leukemia 1 (MCL-1) plays an important role in cardiac protection and cardiac-specific ablation of Mcl-1 causes fatal dilated cardiomyopathy. We hypothesized that there is a gender difference in the expression of cardiac MCL-1 in young, diabetic males and females. To test this, we compared cardiac Mcl-1 expression in young rats (ages 11-12 weeks): diabetic males [ZDF obese (n=8)], non-diabetic females [ZDF lean (n=5)], and non-diabetic males [ZDF lean (n=8)] on Purina 5008 diet to young, diabetic female rats [ZDF obese (n=5)] on Research Diets D12468. Body composition analysis by EchoMRI showed that both male and female lean rats had increased muscle mass compared to their obese, diabetic counterparts, whereas obese, diabetic male and female rats had &gt;5-fold increase in body fat than the corresponding lean controls (p&lt;0.05). Fasting plasma insulin levels in lean female rats were lower than that seen in lean male rats; there was a 49-fold increase in diabetic females, and only a 12-fold increase in diabetic males, compared to lean counterparts (p&lt;0.05). Fasting plasma glucose levels increased 2-3 fold in diabetic males and females, compared to lean controls (p&lt;0.05). Quantitative RT-PCR analysis of mRNA isolated from cardiac tissues of lean and diabetic male and female rats showed that there was a 45% decrease in Mcl-1 mRNA in diabetic males compared to lean males, whereas there was an over 90% suppression of Mcl-1 mRNA in diabetic females compared to lean females (p&lt;0.05). Previously, we have shown that insulin improves MCL-1 expression in female mouse cardiomyocytes. Data presented here suggest that although diabetic female rats have more severe hyperinsulinemia compared to diabetic male rats, loss of cardiac MCL-1 expression is more severe in diabetic females compared to diabetic males.This may underlie the loss of cardioprotective benefits in diabetic females.

A rosemary extract rich in carnosic acid selectively modulates caecum microbiota and inhibits β-glucosidase activity, altering fiber and short chain fatty acids fecal excretion in lean and obese female rats.

BackgroundCarnosic acid (CA) and rosemary extracts (RE) show body-weight, energy metabolism and inflammation regulatory properties in animal models but the mechanisms are not yet understood. Gut microbiota plays an important role in the host metabolism and inflammatory status and is modulated by the diet. The aim of this research was to investigate whether a RE enriched in CA affected caecum microbiota composition and activity in a rat model of genetic obesity.Methods and Principal FindingsA RE (40% CA) was administered with the diet (0.5% w/w) to lean (fa/+) and obese (fa/fa) female Zucker rats for 64 days. Changes in the microbiota composition and β-glucosidase activity in the caecum and in the levels of macronutrients and short chain fatty acids (SCFA) in feces were examined. The RE increased the Blautia coccoides and Bacteroides/Prevotella groups and reduced the Lactobacillus/Leuconostoc/Pediococccus group in both types of animals. Clostridium leptum was significantly decreased and Bifidobacterium increased only in the lean rats. β-Glucosidase activity was significantly reduced and fecal fiber excretion increased in the two genotypes. The RE also increased the main SCFA excreted in the feces of the obese rats but decreased them in the lean rats reflecting important differences in the uptake and metabolism of these molecules between the two genotypes.ConclusionsOur results indicate that the consumption of a RE enriched in CA modifies microbiota composition and decreases β-glucosidase activity in the caecum of female Zucker rats while it increases fiber fecal elimination. These results may contribute to explain the body weight gain reducing effects of the RE. The mutated leptin receptor of the obese animals significantly affects the microbiota composition, the SCFA fecal excretion and the host response to the RE intake.

Effect of the estrous cycle and surgical ovariectomy on energy balance, fuel utilization, and physical activity in lean and obese female rats

This study presents an in-depth analysis of the effects of obesity on energy balance (EB) and fuel utilization in adult female rats, over the estrous cycle and immediately after surgical ovariectomy (OVX), to model pre- and postmenopausal states, respectively. Female Wistar rats were fed a high-fat (46%) diet for 16 wk to produce mature lean and obese animals. Stage of estrous was identified by daily vaginal lavage, while energy intake (EI), total energy expenditure (TEE), and fuel utilization were monitored in a multichamber indirect calorimeter and activity was monitored by infrared beam breaks. Metabolic monitoring studies were repeated during the 3-wk period of rapid OVX-induced weight gain. Component analysis of TEE was performed to determine the nonresting and resting portions of energy expenditure. Obesity was associated with a greater fluctuation in EB across the estrous cycle. Cycling obese rats were less active, expended more energy per movement, and oxidized more carbohydrate than lean rats. The changes in EB over the cycle in lean and obese rats were driven by changes in EI. Finally, OVX induced a large positive energy imbalance in obese and lean rats. This resulted primarily from an increase in EI in both groups, with little change in TEE following OVX. These observations reveal a dominant effect of obesity on EB, fuel utilization, and activity levels in cycling rats, which has implications for studies focused on obesity and EB in female rodents.

Energy Restriction Is Associated with Lower Bone Mineral Density of the Tibia and Femur in Lean but Not Obese Female Rats1–3

Energy restriction decreases bone mineral density (BMD), and epidemiological studies suggest that the risk of weight loss-induced bone loss is greater in lean than in heavier individuals. Our goal in this study was to determine how bone density and geometry respond to energy restriction in mature obese rats compared with lean rats. At 6 mo of age, 36 diet-induced obese and lean female Sprague-Dawley rats were allocated to control (CTL; ad libitum; n = 18) and energy-restricted (EnR; 40% restriction; n = 18) diets. After 10 wk of dietary intervention, obese EnR rats lost more weight (−61 ± 14 g) than lean EnR rats (−91 ± 34 g) (P < 0.02), whereas body weight did not change significantly in the 2 CTL groups (14 ± 23 g). Only the lean EnR (and not obese EnR) rats showed lower BMD compared with CTL rats at the tibia, distal, and proximal femur and femoral neck, and trabecular bone volume (P < 0.05). Serum estradiol declined in lean EnR rats compared with baseline (P < 0.05) but not in the obese EnR rats. In addition, the final serum 25-hydroxyvitamin D (25OHD) concentration was higher (P < 0.05) in obese than in lean EnR rats. Serum parathyroid hormone decreased (P < 0.05) from baseline to final in lean and obese CTL, but not EnR rats. These data support the hypothesis that energy restriction in lean rats compared with obese rats is more detrimental to bone, and it is possible that the greater decline in estrogen and lower levels of 25OHD contribute to this effect.

Effects of vitamin A deficiency on plasma glucose, insulin and leptin in rats

We have observed that vitamin A and its catalytic metabolites synergize with insulin to induce hepatic glucokinase gene expression. It has been known that retinoic acid (RA) can stimulate the expression of phosphoenolpyruvate carboxyl kinase gene. To better understand the role of vitamin A in glucose and lipid homeostasis, Zucker lean (female) rats were fed with vitamin A deficient (VAD) diet (TestDiet, #5822) and vitamin A sufficient (VAS) control diet (TestDiet, #5755) after weaning. When body weight gain stops in VAD group (a sign of vitamin A deficiency), animals were used to access the effect of vitamin A deficiency on glucose and lipid metabolism. All animals were fasted for six hours before their plasma and tissue samples were collected for analysis. To this end, levels of plasma glucose (153 ± 35.1 mg/dl), insulin (0.6 ±0.35 ng/ml), and leptin (1.3 ± 0.64 ng/ml) of VAD rats were all significantly lower than those (glucose, 176 ± 23.1 mg/dl; insulin, 1.3 ± 0.6 ng/ml; 4.0 ± 2.1 ng/ml, all P&lt;0.04)) of VAS fed rats. When VAD rats were fed with VAS diet, the plasma glucose level (150 ± 40 mg/dl) of VAD rats was significantly increased at three days (207 ± 62 mg/dl) and seven days (221 ± 45 mg/dl) after initiation of VAS diet (all P&lt;0.05). At the same time, plasma cholesterol level (49.6 ± 4.4 mg/dl) of VAD rats was also significantly increased at three days (74.6 ± 8.8 mg/dl) and seven days (83.9 ± 9.2 mg/dl) after resuming the VAS diet (all P&lt;0.004). These results demonstrated that vitamin A deficiency can result in the changes of plasma glucose and lipid homeostasis. Further work is warranted to understand the underlying molecular mechanisms.

Effects of Long‐Term Soy vs Casein Protein Intake and Obesity on Serum Insulin‐Like Growth Factor 1 (IGF‐1) and IGFBP‐3 Levels using Obese Female Zucker Rat Model.

Elevated serum IGF‐1 is an increased risk for breast cancer development. The main objective of this study was to investigate the effects of high‐isoflavone soy protein diet (hi‐isof soy) and obesity on serum IGF‐1 and binding proteins (BP) IGFBP‐3 levels. Lean (n= 54) and obese female (n= 45) rats were maintained on either casein or hi‐isof soy protein diet for 155 days. All rats were orally gavaged at age 50 days with 65 mg/kg DMBA. Rats were weighed once a week and were killed 155 days post‐DMBA treatment. Blood was drawn at different ages (43, 99 and 197 d) to measure serum IGF‐1 and IGFBP‐3 by ELISA. Obese soy‐fed rats gained significantly (P&lt;0.001) more weight than obese casein rats. At age 43 d, no differences were found on serum IGF‐1 levels among the groups but IGFBP‐3 was increased by hi‐isof soy diet in lean group compared to casein fed lean rats. At the age 99 d, obesity increased (P&lt;0.05) serum IGF‐1 for both casein and high‐isof soy diet compared to the lean group and this increase was maintained for the rest of the experiment. At age 99 d ,obesity increased IGFBP‐3 levels in the casein but not hi‐isof soy diet group (P&lt;0.001). At end of the Exp. (197 d); obesity increased IGFBP‐3 (P&lt;0.05) both in casein and soy fed rats. Our results suggest that dietary protein source can affect serum IGF‐1 and IGFBP‐3 in obese female Zucker rats (Supported by USDA and ABI to RH).

Effects of soy vs casein protein diet and obesity on DMBA‐induced mammary tumor development

We reported that intact obese Zucker rats fed a standard chow diet had increased susceptibility to DMBA‐induced mammary tumors compared to lean Zucker rats. The objective of this study was to investigate the effects of long‐term high‐isoflavone soy (high‐isof soy) vs casein protein intake and obesity on mammary tumor development. Five‐week‐old female Zucker rats (45 obese fa/fa and 54 lean) were used. After one week of acclimation (age 42 days), rats were randomly assigned to the following diet groups; 1) Lean, casein; 2) Obese, casein; 3) Lean, high‐isof soy and 4) Obese, high‐isof soy. Rats had ad lib access to water and diet with either casein or high‐isof soy protein isolate. All rats were orally gavaged at age 50 days with 65 mg/kg DMBA. Rats were weighed and palpated twice weekly and sacrificed 155 days post‐DMBA treatment. Obese soy‐fed rats gain significantly (P&lt;0.001) more weight than obese casein rats. The lean high‐isof soy‐fed rats had slightly higher (P = 0.057) body weight than lean casein‐fed rats. At end of the experiment, 69% of the Lean casein rats developed mammary tumors compared to 50% in lean high‐isof soy group (P&lt;0.08). In the obese group, 76% of the high‐isof soy‐fed rats developed mammary tumors compared to 15% (P&lt;0.05) of obese casein‐ fed rats. These results suggest that intact obese versus lean Zucker female rats fed semi‐purified diets are protected against DMBA‐induced mammary tumor on the casein but not high‐isof soy diet. (Supported by USDA and ABI to RH)