Control Chow Research Articles

Cafeteria diet and caloric restriction affect metabolic but not behavioral characteristics in male Wistar rats

This study aimed to evaluate the effects of a cafeteria diet and caloric restriction on behavioral and metabolic profiles of adult male Wistar rats. The rats were randomly divided into three groups (n = 12/group) and from 10 weeks of age fed either ad libitum standard rat chow (control group), ad libitum cafeteria diet in addition to standard chow (diet-induced obesity (DIO) group) or kept on caloric restriction (at 85% weight of controls; restricted group) for a period of 12 weeks. Body weight was assessed twice per week and glucose levels were measured at three times during the 12-week period. At week 11 the animals were behaviorally profiled using the multivariate concentric square field™ (MCSF) test. After 12 weeks of diet the animals were euthanized, blood collected, relative organ weights were assessed and plasma or serum levels of insulin, glucose, and lipid profile were measured. The DIO group gained 23% more weight than the control group (p < 0.001) and increased adipose tissue weight in comparison to the control (p < 0.001) and restricted (p < 0.001) groups. Glucose was significantly increased (p < 0.001) only during the second measurement at week 7 and insulin levels were elevated in the DIO group compared to controls and restricted groups (p < 0.01; p < 0.001, respectively). Plasma cholesterol levels were reduced for both DIO (p < 0.01) and restricted (p < 0.001) groups relative to controls. Adiponectin and leptin levels were higher for the DIO group in comparison to both the control (p < 0.001; p < 0.05) and restricted (p < 0.001; p < 0.001) groups. Thus, the two diets led to significant changes in body weight gain, adiposity, and metabolism. However, they did not alter the behavioral profiles in the MCSF test, suggesting that activity, exploration, risk assessment, risk taking or shelter seeking remained unaffected by the dietary interventions. The current findings suggest that an increase or reduction in energy intake resulted in no behavioral effects, despite the accompanying glycemic alterations potentially related to diabetes development.

Host Genetics Background Affects Intestinal Cancer Development Associated with High-Fat Diet-Induced Obesity and Type 2 Diabetes.

Obesity and type 2 diabetes (T2D) promote inflammation, increasing the risk of colorectal cancer (CRC). High-fat diet (HFD)-induced obesity is key to these diseases through biological mechanisms. This study examined the impact of genetic background on the multimorbidity of intestinal cancer, T2D, and inflammation due to HFD-induced obesity. A cohort of 357 Collaborative Cross (CC) mice from 15 lines was fed either a control chow diet (CHD) or HFD for 12 weeks. Body weight was tracked biweekly, and blood glucose was assessed at weeks 6 and 12 via intraperitoneal glucose tolerance tests (IPGTT). At the study's endpoint, intestinal polyps were counted, and cytokine profiles were analyzed to evaluate the inflammatory response. HFD significantly increased blood glucose levels and body weight, with males showing higher susceptibility to T2D and obesity. Genetic variation across CC lines influenced glucose metabolism, body weight, and polyp development. Mice on HFD developed more intestinal polyps, with males showing higher counts than females. Cytokine analysis revealed diet-induced variations in pro-inflammatory markers like IL-6, IL-17A, and TNF-α, differing by genetic background and sex. Host genetics plays a crucial role in susceptibility to HFD-induced obesity, T2D, CRC, and inflammation. Genetic differences across CC lines contributed to variability in disease outcomes, providing insight into the genetic underpinnings of multimorbidity. This study supports gene-mapping efforts to develop personalized prevention and treatment strategies for these diseases.

8480 TIX100, a Novel Small Molecule for the Treatment of MASLD/MASH

Abstract Disclosure: S. Jo: None. J. Chen: None. G. Jing: None. K.M. Habegger: None. A. Shalev: Other; Self; TIXiMED, Inc. Founder &amp; CSO. Metabolic dysfunction-associated steatotic liver disease (MASLD) and steatohepatitis (MASH) (aka NAFLD and NASH) affect over 1 billion people worldwide, yet no approved therapy is currently available. MASLD/MASH is often associated with diabetes and obesity and some diabetes medications have shown some beneficial effects in this context. However, beyond steatosis these drugs are in general unable to prevent all the problems associated with MASH, such as inflammation, hepatocyte injury and especially fibrosis. Interestingly, we have now discovered that the new antidiabetic small molecule thioredoxin-interacting protein (TXNIP) inhibitor, TIX100, effectively protects against high-fat diet-induced hepatic steatosis. Oral TIX100 also protected obese and diabetic db/db mice against steatosis and significantly reduced the elevated expression levels of lipogenic genes found in db/db livers, many of which have been shown to be elevated in human MASLD. In addition though, TIX100 also normalized the blood glucose of db/db mice to the levels observed in lean control animals. Therefore, to address the question of whether TIX100 would still be effective in the absence of diabetes, obesity and any genetic leptin-receptor deficiency, we used a validated choline-deficient, amino acid-defined, high-fat diet to induce MASH. Indeed, compared to control chow, this diet significantly increased plasma liver enzymes indicating liver injury without causing diabetes or obesity. It also led to elevated hepatic expression of inflammatory and fibrogenic markers and histology revealed lipid accumulation and steatosis as well as pronounced fibrosis and upregulation of liver macrophages. Excitingly, all these pathological changes were blunted or improved by oral TIX100 treatment. Taken together, oral TIX100 may provide an attractive and effective novel approach for the treatment of MASLD/MASH that goes beyond the correction of diabetes, obesity, and steatosis. Presentation: 6/3/2024

7279 Survodutide and Semaglutide Both Induce Weight Loss but Show Different Effects on Food Preference and Dyslipidemia in the Free Choice Diet-induced Obese Hamster Model

Abstract Disclosure: F. Briand: Employee; Self; Physiogenix. R. Augustin: Employee; Self; Boehringer Ingelheim. K. Bleymehl: Employee; Self; Boehringer Ingelheim. T. Zimmermann: Employee; Self; Boehringer Ingelheim. T. Klöckener: Employee; Self; Boehringer Ingelheim. T. Klein: Employee; Self; Boehringer Ingelheim. Introduction: Semaglutide, a GLP-1 receptor agonist, and survodutide, a GCGR/GLP-1R dual agonist, have demonstrated significant weight loss in patients with obesity. To test whether this weight loss is associated with changes in food preference and improvement in dyslipidemia, we evaluated semaglutide and survodutide in the free choice diet-induced obese and dyslipidemic hamster, a model with a human-like lipoprotein metabolism. Methods: Obesity and dyslipidemia were induced with a 20-week free choice diet, which presents hamsters with a choice between control chow diet or high fat/cholesterol diet, and normal water or 10% fructose water. Obese hamsters were then kept on this free choice diet and treated subcutaneously once daily with vehicle, semaglutide 15 nmol/kg or survodutide 15 nmol/kg for 5 weeks. Results: Compared with vehicle, semaglutide and survodutide led to a 17% and 11% body weight reduction, respectively (both p&amp;lt;0.001 vs vehicle). Compared to vehicle, both chow diet and high fat diet intakes were significantly lower during the first week of semaglutide treatment, but then returned to baseline values. However, semaglutide significantly increased normal water intake and reduced fructose-enriched water intake during the whole treatment period (both p&amp;lt;0.001). Survodutide had a different effect, as it did not alter chow diet and normal water intakes, but significantly reduced high fat diet and fructose-enriched water intakes during the 5-week treatment period. Semaglutide and survodutide significantly reduced plasma insulin levels and the HOMA-IR index of insulin resistance (both p&amp;lt;0.01 vs. vehicle). Both semaglutide and survodutide significantly reduced plasma total cholesterol levels by 24% and 41%, respectively (both p&amp;lt;0.001). This hypocholesterolemic effect led to lower HDL-cholesterol levels (-25%, p&amp;lt;0.001) but unchanged LDL-cholesterol levels for hamsters treated with semaglutide. In contrast, in hamsters treated with survodutide, cholesterol lowering was observed in all lipoprotein fractions, including LDL-cholesterol (-39%, p&amp;lt;0.001). Conclusion: Semaglutide and survodutide both induce weight loss and lower HOMA-IR, but have different effects on food preference and dyslipidemia in the obese hamster model; such effects may be attributed to the GCGR agonism of survodutide. These preclinical data highlight the potential metabolic benefits of the GCGR/GLP-1R dual agonist survodutide for the treatment of obesity and related metabolic comorbidities. Presentation: 6/3/2024

6681 Age and Sex Disparity in NASH and the Potential Role of Estradiol Treatment for NASH

Abstract Disclosure: M. Tripathi: None. S. Sakthivel: None. A. Suzuki: None. B.K. Singh: None. P.M. Yen: None. Introduction: NAFLD (nonalcoholic fatty liver disease) and its more severe form, NASH (nonalcoholic steatohepatitis) are sexually dimorphic conditions that exhibit higher prevalence in men than women during reproductive ages but have similar prevalence after menopause. Reproductive age women are protected from hepatic fibrosis relative to men, although they lose this protection after menopause. These sex differences in NASH have been attributed, at least partly, to different circulating estrogen levels. However, the underlying mechanism(s) is not fully understood. We thus conducted animal experiments to investigate: 1) sex differences in the histologic severity of NASH, necroinflammatory and fibrogenic activities, and other relevant pathways in young and old male and female mice and 2) the effects of estradiol administration on these parameters in old male and female mice to evaluate the therapeutic potential of estradiol in NASH. Methodology: Young/reproductive age (12-weeks old) and old/post-reproductive age female and male (48∼50-weeks weeks old) C57BL/6J mice were fed control chow diet (NCD) or high-fat methionine/choline-deficient (MCD) diet for six weeks to induce NASH. The old mice were administered 200nM 17β-estradiol (physiological) in drinking water. We analyzed serum and hepatic NASH parameters, ER stress, and lysosome-autophagy defects. Results and Conclusion: Among reproductive age mice with NASH, females developed less severe histologic inflammation and fibrosis and decreased gene expression of Il1b, Il6, Tgfb, Col1a1, Ccl2, and Ccl5, and lower hepatic collagen than males. Females with NASH also had less inhibition of hepatic autophagy and lysosomal protein expression than males with NASH. Among old mice, post-reproductive age females fed NCD or MCD exhibited more severe hepatic inflammation and fibrosis and had higher gene expression of inflammatory and fibrogenic markers with comparable autophagic inhibition and lysosomal defects as similar age males. Physiological doses of estradiol significantly reduced hepatic gene expression of inflammatory and fibrogenic markers, ER stress and lysosomal-autophagic defects in post-reproductive age females. In conclusion, similar to clinical NASH, we observed marked sex and age-related differences in the severity of NASH. Furthermore, physiological doses of estradiol treatment improved the hepatic histology, lysosome-autophagy, inflammation, and fibrosis in post-reproductive age female mice with NASH and suggests that estradiol supplementation may decrease or reverse NASH progression in post-menopausal women with NASH. Presentation: 6/2/2024

Alternate-day fasting delays pubertal development in normal-weight mice but prevents high-fat diet-induced obesity and precocious puberty

Background/ObjectivesChildhood obesity, particularly in girls, is linked to early puberty onset, heightening risks for adult-onset diseases. Addressing childhood obesity and precocious puberty is vital to mitigate societal burdens. Despite existing costly and invasive medical interventions, introducing lifestyle-based alternatives is essential. Our study investigates alternate-day fasting’s (ADF) impact on pubertal development in normal-weight and high-fat diet (HFD)-induced obese female mice.MethodsFour groups of female mice were utilized, with dams initially fed control chow during and before pregnancy. Post-parturition, two groups continued on control chow, while two switched to an HFD. Offspring diets mirrored maternal exposure. One control and one HFD group were subjected to ADF. Morphometry and hormone analyses at various time points were performed.ResultsOur findings demonstrate that ADF in normal-weight mice led to reduced body length, weight, uterine, and ovarian weights, accompanied by delayed puberty and lower levels of sex hormones and growth hormone (GH). Remarkably, GH treatment effectively prevented ADF-induced growth reduction but did not prevent delayed puberty. Conversely, an HFD increased body length, induced obesity and precocious puberty, and altered sex hormones and leptin levels, which were counteracted by ADF regimen. Our data indicate ADF’s potential in managing childhood obesity and precocious puberty.ConclusionsADF reduced GH and sex hormone levels, contributing to reduced growth and delayed puberty, respectively. Therefore, parents of normal-weight children should be cautious about prolonged overnight fasting. ADF prevented HFD-induced obesity and precocious puberty, offering an alternative to medical approaches; nevertheless, further studies are needed for translation into clinical practice.

Liver proteomics identifies a disconnect between proteins associated with de novo lipogenesis and triglyceride storage

De novo lipogenesis (DNL) has been implicated in the development and progression of liver steatosis. Hepatic DNL is strongly influenced by dietary macronutrient composition with diets high in carbohydrate increasing DNL and while diets high in fat decrease DNL. The enzymes in the core DNL pathway have been well characterised, however less is known about other liver proteins that play accessory or regulatory roles. In the current study, we associate measured rates of hepatic DNL and fat content with liver proteomic analysis in mice to identify known and unknown proteins that may have a role in DNL. Male mice were fed either a standard chow diet, a semi-purified high starch or high fat diet. Both semi-purified diets resulted in increased body weight, fat mass and liver triglyceride content compared to chow controls and hepatic DNL was increased in the high starch and decreased in high fat fed mice. Proteomic analysis identified novel proteins associated with DNL that are involved in taurine metabolism, suggesting a link between these pathways. There was no relationship between proteins that associated with DNL and those associated with liver triglyceride content. Further analysis identified proteins that are differentially regulated when comparing a non-purified chow diet to either of the semi-purified diets which provide a set of proteins that are influenced by dietary complexity. Finally, we compared the liver proteome between 4- and 30-week diet-fed mice and found remarkable similarity suggesting metabolic remodelling of the liver occurs rapidly in response to differing dietary components.

Maternal high-fat high-sugar diet impairs bone quality and strength but not cartilage in aging mice.

Osteoarthritis (OA) is a prevalent aging disorder of synovial joints and recent work suggests that a parental high-fat diet increases OA severity following joint injury in offspring. We hypothesized that a maternal high-fat high-sugar (HFHS) diet would promote spontaneous osteoarthritis-related cartilage and bone changes in 1-year-old offspring. Female C57BL/6 J mice were placed on either a chow control or HFHS diet for 6 weeks before mating to a chow-fed C57BL/6 J male and maintained on their assigned diets throughout pregnancy and lactation. Male and female offspring were weaned onto a chow diet, raised to 1 year of age, and evaluated for cartilage and bone changes indicative of OA. However, offspring did not show early signs of OA as measured by histological Mankin scoring, mechanical testing of the pericellular matrix, histological synovitis scoring, or subchondral bone thickening as measured by microcomputed Tomography. On the other hand, male offspring from HFHS-fed dams had reduced trabecular bone quality in the tibial metaphysis and decreased cortical thickness. Although maternal HFHS diet did not impact trabecular or cortical bone quality in tibias of female offspring, the radii of these animals had decreased cortical thickness, increased medullary area, and impaired breaking strength compared to those of control-fed dams. Finally, we evaluated bone quality and strength in male and female F2 offspring and found that the grandmaternal diet modestly impacted radial bone geometry but not strength. Together these results suggest that maternal HFHS diet impairs F1 offspring skeletal integrity in a sex and bone site-specific manner.

Comparison of human breast milk vs commercial formula-induced early trophic enteral nutrition during postoperative prolonged starvation in an animal model

The present study aimed to characterize the changes in macromolecular composition and structure in ileal tissue induced by postoperative prolonged starvation (PS), human breast milk feeding (HM) and commercial formula feeding (CF) for 48 and 72 h (h). Forty-two Wistar albino rats underwent an ileal transection and primary anastomosis and were then divided into six subgroups. Two groups of seven rats were food-deprived for 48 and 72 h with free access to water only in metabolic cages (48 h PS, 72 h PS). Then, two groups of seven rats received early enteral trophic nutrition (EEN) either using HM, and CF at 48 h post-operation (48 h HM, 48 h CF). The other two groups of seven rats received the same trophic enteral nutrition at 72 h post-operation (72 h HM, 72 h CF). An additional seven rats were fed normal rat chow (control), after which the ileal tissues were harvested and freeze-dried overnight. Then sample spectra were recorded by Fourier transform infrared (FTIR) spectroscopy. PS at 48 and 72 h resulted in an increase in the concentration of lipids and a decrease in the concentration of proteins. CF and HM trophic feeding induced a decrease in membrane fluidity and an increase in lipid order. Ileal tissues showed similar compositional and structural changes in lipids and proteins in the PS and CF groups after 48 and 72 h. A marked decrease in nucleic acid concentration was seen in CF at 48 h compared to HM. The human milk feeding groups did not induce any significant alterations and showed compositional and structural data similar to the controls. In conclusion, EEN application seems to be safer when introduced at 48 h rather than 72 h and time of this nutrition is crucial to maintain ileum structure and therefore immunity and well-being. HM-induced trophic nutrition is seen to protect the ileal tissue from significant alterations within lipid and protein compositions, whereas CF caused notable changes. HM is absolutely the best nutritional source for gut health in this animal model.

Abstract P452: Cell Populations and Transcriptomes Differ Across Blood Pressure and Sex in Perivascular Adipose Tissue: A Single-Nuclei RNA Sequencing Study

The perivascular adipose tissue (PVAT), fat that surrounds many blood vessels, is capable of modulating vascular tone through multiple modalities. Despite increasing research interest, PVAT remains less understood compared to other vascular layers. We hypothesized that the transcriptome and cell population of thoracic aorta PVAT (taPVAT) differ between hypertensive and normotensive rats. To test this hypothesis, we performed single-nucleus RNA-sequencing (snRNA-seq) on twenty-five taPVAT samples from male and female Dahl SS rats, fed either control (Con) chow or a high-fat diet (HFD) for 8 or 24 weeks. Mean arterial pressure (MAP) measurements showed elevated values in female rats fed a HFD for 24 weeks with a trend towards increasing blood pressure in male rats (Fig. 1a). Our analysis included at least three taPVAT samples per treatment group, resulting in 71,813 high-quality sequenced nuclei, representing eight major cell types (Fig. 1b) and twenty-eight cell sub-types. Within each cell type, the differences in population and gene expression across diet, diet duration, and sex were examined. The most substantial population proportion differences were observed across diet duration, with fewer differences noted across diet and sex (Fig. 1c). The total number of differentially expressed genes (DEGs) (adjusted p-value ≤ 0.05, |log 2 (fold change)| ≥ 1) across diet, diet duration, and sex were 191, 424, and 119, respectively. Interestingly, in the taPVAT from HFD fed rats, the gene Csmd1 (CUB and Sushi multiple domains 1) was consistently upregulated across nearly all cell types. Whereas Scd (stearoyl-CoA desaturase), which is involved in fatty acid biosynthesis, was frequently the most downregulated gene. Pathway enrichment analysis revealed that the DEGs related to the HFD were associated with a decrease in lipid and fatty acid biosynthetic processes. DEGs related to rats fed a HFD for twenty-four weeks versus eight weeks were linked to an enrichment in inflammatory response and extracellular matrix organization pathways, and in HFD fed male rats compared to females, there was enrichment for leukocyte aggregation and peptidyl-cysteine modification pathways. In conclusion, our findings help paint a more detailed picture of the molecular and cellular changes in taPVAT associated with hypertension.

Intravenous lipid emulsions designed to meet preterm infant requirements increase plasma and tissue levels of docosahexaenoic acid and arachidonic acid in mice

Intravenous lipid emulsions used in preterm infants contain insufficient docosahexaenoic acid (DHA) and arachidonic acid (ARA) to support normal development, resulting in deficiencies that contribute to complications of prematurity and cognitive delay. We sought to investigate the effects of new intravenous lipid emulsions designed to contain sufficient DHA and ARA to meet preterm needs, while avoiding liver toxicity. Three new lipid emulsions (NLE A-C) were laboratory-generated using high pressure homogenization. First, a long-term experiment evaluated the impact on plasma, liver, and frontal cortex fatty acid composition compared to commercially available lipid emulsions. Lipid emulsions were administered via daily orogastric gavage to four-week-old C57Bl/6J mice. Next, liver toxicity was evaluated in a murine model of parenteral nutrition-induced hepatosteatosis. Mice were provided an ad lib fat-free high carbohydrate diet, with intravenous lipid emulsion administration every other day for 19 days. Administration of commercially available lipid emulsions (soybean oil, mixed oil, or fish oil) resulted in decreased plasma and tissue levels of DHA and/or ARA compared to a chow control. The new lipid emulsions demonstrated a dose-response effect in plasma and tissue concentration of DHA and ARA. NLE C (with an approximately even DHA:ARA ratio), compared to chow, maintained similar DHA (19.2±0.3 vs. 19.3±0.3%, P=1.00) and ARA (10.4±0.2 vs. 9.9±0.2% ARA, P=0.75) content in frontal cortex tissue. All three new lipid emulsions prevented biochemical liver injury and pathologist-assessed hepatosteatosis; soybean oil lipid emulsion and mixed oil lipid emulsion treatment resulted in hepatosteatosis in both experiments. Long-term treatment with the new lipid emulsions in juvenile mice resulted in increased plasma and tissue DHA and/or ARA content compared to currently available lipid emulsions. The new lipid emulsions also prevented hepatosteatosis and biochemical liver injury with enteral and parenteral administration.

The beneficial impact of ketogenic diets on chemically-induced colitis in mice depends on the diet's lipid composition

Previously, we showed that restrictive diets, including ketogenic diet (KD), have an anti-inflammatory impact on the healthy gastrointestinal tract of mice. Afterward, we found that energy-restricting diets mitigate inflammation in the dextran sodium sulfate (DSS) colitis mouse model. The current study aimed to verify the impact of KD on DSS colitis and assess if the diet's fat composition influences the outcomes of the intervention.Mice with mild chronic colitis were fed control chow, KD composed of long-chain triglycerides (KD LCT) or a KD containing a mix of LCT and medium-chain triglycerides (KD LCT/MCT).KDs did not reverse DSS-enhanced gut permeability and shortening of the colon. Both KDs had a similar impact on liver, cecum, and spleen weight, villi and colon length, the thickness of muscularis externa, and expression of ZO-1 and occludin. On the contrary, body weight, glutathione (GSH) and taurine-GSH levels, GSH-S transferase (GST), and myeloperoxidase (MPO) activity, as well as an abundance of several fecal bacteria, all were differentially affected by the two types of KDs. When compared to the DSS control diet, reduction in colon mucosa cytokines expression was stronger in KD LCT than in the KD LCT/MCT group.We conclude that the outcomes of the KD interventions in terms of potential therapeutical applications depend on lipid composition. KD LCT showed a strong positive impact on gut inflammation. A potential contribution of GSH to KD outcomes and a correlation between MPO activity and microbiota composition was identified.

Abstract Mo111: Comparison of Pre-Clinical Models of Heart Failure with Preserved Ejection Fraction Using Multimodal Approaches

There are various pre-clinical models of heart failure with preserved ejection fraction (HFpEF) designed to model different clinical phenotypes of HFpEF. However, detailed assessment and head to head comparison of diastolic function by invasive and non-invasive techniques in these models is lacking. Our goal was to comprehensively evaluate cardiac structure and function in three HFpEF mouse models – deoxycortisterone acetate (DOCA)-salt, Nω-nitro-l-arginine methyl ester high fat diet (LNAME+HFD), and aged female mice fed HFD with angiotensin infusion (Ang+HFD) – using both non-invasive and invasive methods. We employed pulse wave and tissue Doppler echocardiography, cardiac magnetic resonance imaging (CMR), and left ventricular (LV) catheterization in the commonly used C57BL/6J mouse, enabling direct comparisons between models. Cardiac hypertrophy was assessed by heart weight (HW) to tibia length (TL) ratio. The Mann-Whitney test was used for statistical testing within each model. We found that on echocardiography, all models demonstrated diastolic dysfunction with significant differences (p &lt; 0.05) or trends for significance (p &lt; 0.1) on tissue doppler peak mitral annular velocity during early filling (e’), ratio of peak early mitral flow velocity (E) to e’ (E/e’), ratio of peak early mitral flow velocity (E) to e’ (E/e’), isovolumetric relaxation time (IVRT), and E pressure half time (E PHT). CMR was notable for findings of reduced right ventricular ejection function in the LNAME+HFD group (36.1±2.7% vs 49.8±1.4% in chow controls, p&lt;0.01). On invasive hemodynamics, only DOCA-salt had a higher LV diastolic time constant (t, 11.7±1.4 vs 6.9±0.8 ms in sham controls, p=0.02) and only Ang+HFD had a higher minimum rate of pressure change in the ventricle (dP/dt min , -3506±342 vs -5268±227 in chow fed, vehicle infusion controls, p&lt;0.01). No model showed a difference in LV end diastolic pressure. For cardiac hypertrophy, we found that both DOCA-salt (8.1±0.2 vs 10.3±0.5, p&lt;0.01) and Ang+HFD (9.2±0.3 vs 11.5±0.4, p&lt;0.01) models demonstrated increased HW to TL ratio, while LNAME+HFD did not. These results highlight differences in cardiac structure and function in HFpEF mouse models and the need for multi-modal assessment in pre-clinical studies.

Kidney outcomes are altered by preconception weight modulation in rodent mothers with obesity

Obesity increases the risk of chronic kidney disease. We have previously demonstrated the benefits of preconception maternal weight loss on fertility and pregnancy outcomes in a mouse model of maternal obesity. Here, we elucidate if preconception weight loss, either by diet modification or the glucose-like peptide 1 agonist liraglutide, used in the treatment of diabetes and obesity, improves maternal kidney outcomes in late gestation. C57BL/6 female mice were fed either a high-fat-diet (HFD) or a chow (control) diet for 8 weeks. To induce pre-pregnancy weight loss, HFD-fed dams were switched to chow diet (HFD-C) or administered liraglutide (0.3 mg/kg subcutaneous) whilst continuing on HFD (HFD-L). Liraglutide was discontinued one week prior to mating. HFD-V mice continued on HFD, with saline injections. A group of HFD-fed dams were ‘diet switched’ to chow after conception (post-conception, HFD-PC). Maternal body weight and glucose tolerance were measured: (1) preconception and (2) during late gestation followed by blood, urine and kidney collection. Serum creatinine, urinary creatinine and albumin, kidney tissue gene expression and protein were measured. In the preconception period, HFD-L and HFD-C mothers have lower urine albumin:creatinine ratios (UACR) and fatty acid synthase (FAS) protein expression (P < 0.005 vs. HFD-V). At late gestation, kidneys of HFD-V and HFD-PC dams have increased gene expression of insulin receptor and FAS (P < 0.05) and higher UACR compared to controls (P < 0.01). In the HFD-PC group, kidneys show increased mRNA and protein expression of metabolic and oxidative stress markers (FAS, 8-OHdG vs. control, P < 0.05, P < 0.0001 respectively). The preconception intervention groups with liraglutide, or diet change show reduced oxidative stress (protein expression of 8-OHdG, P < 0.05 vs. HFD), mRNA and protein expression of FAS (P < 0.05 vs. HFD), protein expression of fibrosis markers (collagen IV, fibronectin vs. HFD, P < 0.05), and UACR (P < 0.05 vs. HFD). This study suggests that preconception weight loss benefits maternal kidney health during pregnancy, superior to diet intervention once already pregnant.

Maternal obesogenic diet operates at the tumor cell of origin to increase incidence and decrease latency of Neurofibromatosis Type 1 optic pathway glioma.

Pediatric low-grade glioma incidence has been rising in the U.S., mirroring the rising rates of pediatric and maternal obesity. Recently, children of obese mothers were demonstrated to develop brain tumors at higher rates. Importantly, obesity in the U.S. is largely driven by diet, given the prevalence of high fat and high sugar (HFHS) food choices. Since high-fat diet exposure can increase embryonic neuroglial progenitor cell (NPC) proliferation, the potential cells of origin for low-grade glioma, we hypothesized that in utero exposure to an obesogenic diet would modify pediatric brain penetrance and latency by affecting the tumor cell of origin. We employed several murine models of the Neurofibromatosis type 1 (NF1) pediatric brain tumor predisposition syndrome, in which optic pathway gliomas (Nf1-OPGs) arise from NPCs in the embryonic third ventricular zone (TVZ). We exposed dams and offspring to an obesogenic HFHS diet or control chow and analysed fetal neurodevelopment at E19.5 and tumor formation at 6w-3mo. Progeny from HFHS diet-exposed dams demonstrated increased TVZ NPC proliferation and glial differentiation. Dietary switch cohorts confirmed that these effects were dependent upon maternal diet, rather than maternal weight. Obesogenic diet (Ob) similarly accelerated glioma formation in a high-penetrance Nf1-OPG strain and increased glioma penetrance in two low-penetrance Nf1-OPG strains. In contrast, Ob exposure in the postnatal period alone did not recapitulate these effects. These findings establish maternal obesogenic diet as a risk factor for murine Nf1-OPG formation, acting in part through in utero effects on the tumor cell of origin.

P-011 Maternal gestational methyl donor deficiency affects reproductive development and fertility of mice offspring

Abstract Study question Is maternal methyl donor deficiency during gestation affects offspring’s reproductive health at adulthood? Summary answer Maternal gestational methyl donor deficiency affects reproductive development and fertility of F1 offspring and also leads to increased post implantation loss What is known already Reproductive disorders and their exact mechanisms are scarce in “The developmental origins of health and disease” (DOHaD) hypothesis which well explains the metabolic complications such as obesity, type 2 diabetes and hypertension that may take root during early development. The maternal nutritional environment during early development has important implications on offspring’s health later in life including reproductive health. The gestational period is of critical importance during which major epigenetic reprogramming occurs. The maternal methyl donor deficiency during gestational period may affect this process and epigenetic signatures of developing gametes and the fertility potential Study design, size, duration Pregnant female OCT4-GFP mice, fed on control chow diet (CCD) &amp; 40% methyl-donors (Folate, Methionine &amp; Vitamin B12) deficient diet (MDD) from gestation day (GD) 5 to GD 20. The gonadal development of F1 offsprings was studied on gestation day GD 18, post-natal day (PND) 22 and PND 60. Fertility study was done by mating adult F1 male offspring with adult female mice fed on standard diet Participants/materials, setting, methods Homocysteine levels in dams (F0) at GD18 were estimated by ELISA. Testosterone levels in F1 offsprings at PND 60 were estimated by ELISA. Furthermore, Serum glucose levels in F1 offsprings were measured at PND 60 and PND 75 by glucometer. Testicular histology, quantitative histomorphometry, immunofluorescence and TUNEL assay was performed at PND 60. Main results and the role of chance Fetal weight was reduced at GD13 and GD18 but postnatal body weight at PND 22 and 60 was not significantly affected in MDD group. Teratological defects were observed in pups of 40% MDD group as compared to CDD. Testosterone levels were significantly reduced in F1 offsprings of 40% MDD group. Glucose levels were similar in both the groups at PND 60 but at PND 75 it is significantly higher in case of F1 offsprings of 40% MDD. Maternal serum homocysteine levels and global DNA methylation levels of fetal gonads were affected. Testicular histology was significantly impaired in 40% MDD group at GD18, PND22 and PND60. Quantitative histomorphometry and immunofluorescence study showed the loss of spermatogonia and spermatocytes with significant increase in germ cells sloughing and seminiferous tubules atrophy. Apoptosis is also observed in seminiferous tubules. Copulation index, sperm count and motility was significantly reduced in case of 40% MDD group at adulthood. The F1 offspring of this group were sub-fertile with significant post implantation loss. Limitations, reasons for caution Although mice share ∼90% genome with humans, they may not resemble all aspect of physiology or pathology in humans hence it could limit the findings to extrapolate to humans. Wider implications of the findings The knowledge gained from this work could be helpful in highlighting the need of gestational micronutrient interventions specially methyl donors, which could help improving maternal and child health as well as reducing the future disease burden of reproductive disorders Trial registration number not applicable

Long-term Supplementation of Deep-fried Oil Consumption Impairs Oxidative Stress, Colon Histology and Increases Neurodegeneration.

Sesame oil and sunflower oil are popular cooking oils in southern India. Deep-frying is a frequent method of food preparation. Deep-frying at high temperatures has been linked with several disorders, including cancer, diabetes, and unknown metabolic problems. There have been no long-term investigations on the influence of deep-fried oils on PUFA metabolism and pathogenesis. As a result, the current study aimed to explore the effect of deep-fried frying oil on Wistar rats by continuous treatment. Furthermore, the pathophysiology of MSG-induced neurotoxicity in Wistar rats was investigated. Wistar rats weighing 200-260 g were used in this study. Female rats were divided into five groups fed with (1) standard chow (control group), (2) unheated sesame oil (UHSO) along with standard chow, and (3) reheated sesame oil (RHSO) along with standard chow, (4) unheated sunflower oil (UHSFO) along with standard chow, and (5) reheated sunflower oil (RHSFO) along with standard chow and continued up to F1 generation. Furthermore, F1 male rats were treated with MSG of 2 g/kg body weight for 10 alternative days and were sacrificed for major tissues. We found that rats treated with RHSO and RHSFO showed increased body weight. Deep-fried oil-fed rats (RHSO and RHSFO) showed a significant increase in total cholesterol- 100 mg/dl, LDL- 23 mg/dl, & TAG-100 mg/dl, when compared to unheated oil rats. Liver function tests revealed that AST and ALT levels were significantly elevated in RHSO and RHSFO when compared to unheated oils and the control group. Inflammatory markers revealed that Hs-CRP (0.35 mg/dl) and LDH levels (6000 U/L) were significantly elevated in RHSO and RHSFO when compared to the unheated oils and control group. RT-PCR results showed significant elevation in the antioxidant genes SOD (twofold) and GPX (3-fold) when compared to UHSO and UHSFO groups. Liver and colon histology showed significant damage in the cell structure of RHSO and RHSFO-treated rats. Further, rats treated with unheated oils and MSG showed statistically significantly higher mRNA expression of neuroplasticity genes CREB, BDNF and reduced NMDA levels (UHSO, UHSFO) when compared to reheated oil groups (RHSO & RHSFO). Proinflammatory marker TNF-α expression was significantly elevated in RHSFO-treated rats when compared to control. Brain histology showed focal damage in glial cell degeneration in rats treated with RHSO and RHSFO when compared to other groups. The results from the present study proved that continuous supplementation deep-fried reheated oil consumption increased serum TGL and oxidative stress markers. Impaired liver metabolism and the involvement of the gut-liver-brain axis increased the risk of neurodegeneration.

Initiation of metformin in early pregnancy results in fetal bioaccumulation, growth restriction, and renal dysmorphology in a primate model

BackgroundIn recent years, pragmatic metformin use in pregnancy has stretched to include prediabetes, type 2 diabetes, gestational diabetes and (most recently) pre-eclampsia. With its expanded use, however, concerns of unintended harm have been raised. ObjectiveWe developed an experimental primate model and applied triple-quadruple pole LC mass spectrometry (UHPLC-QQQ) for direct quantitation of maternal and fetal tissue metformin levels with detailed fetal biometry and histopathology. Study designWithin 30 days of confirmed conception (defined as early pregnancy), n=13 time-bred (TMB) Rhesus dams with gestations designated for fetal necropsy were initiated on twice daily human dose-equivalent 10 mg/kg metformin or vehicle control. Pregnant dams were maintained as pairs and fed either a control chow or 36% fat Western-style diet (WSD). Metformin or placebo vehicle control were delivered in a variety of treats while animals were separated via a slide. A Cesarean was performed at G145, and amniotic fluid and blood were collected and the fetus and placenta were delivered. The fetus was immediately necropsied by trained primate center personnel. All fetal organs were dissected, measured, sectioned, and processed per clinical standards. Fluid and tissue metformin levels were assayed using validated UHPLC-QQQ in SRM against standard curves. ResultsAmong the n=13 G145 pregnancies with fetal necropsy, n=1 dam and its fetal tissues had detectable metformin levels despite being allocated to the vehicle control group (>1 μM metformin/kg maternal weight or fetal/placental tissue), while a second fetus allocated to the vehicle control group had severe fetal growth restriction (birthweight 248.32 g, <1%) and was suspected of having a fetal congenital condition. After excluding these two fetal gestations from further analyses, 11 fetuses from dams initiated on either vehicle control (n=4, 3 female, 1 male fetuses) or 10 mg/kg metformin (n=7, 5 female, 2 male fetuses) were available for analyses. Among dams initiated on metformin by G30 (regardless of maternal diet), we observed significant bioaccumulation within the fetal kidney (0.78-6.06 μmol/kg, mean 2.48 μmol/kg) , liver (0.16-0.73 μmol/kg, mean 0.38 μmol/kg), fetal gut (0.28-1.22 μmol/kg, mean 0.70 μmol/kg), amniotic fluid (0.43-3.33 μmol/L, mean 1.88 μmol/L), placenta (0.16-1.0 μmol/kg , mean 0.50 μmol/kg) and fetal serum (0 -0.66 μmol/L , mean 0.23 μmol/L ), and fetal urine (4.1-174.1 μmol/L mean 38.5 μmol/L ), with fetal levels near biomolar equivalent to maternal levels (maternal serum 0.18-0.86 μmol/L , mean 0.46 μmol/L; maternal urine 42.6-254.0 μmol/L , mean 149.3 μmol/L). WSD feeding neither accelerated nor reduced metformin bioaccumulations in maternal or fetal serum, urine, amniotic fluid, placenta nor fetal tissues. In these 11 animals, fetal bioaccumulation of metformin was associated with less fetal skeletal muscle (57% lower cross-sectional area of gastrocnemius) and decreased liver, heart, and retroperitoneal fat masses (p<0.05), collectively driving lower delivery weight (p<0.0001) without changing the crown-rump length. Sagittal sections of fetal kidneys demonstrated delayed maturation, with disorganized glomerular generations and increased cortical thickness; this renal dysmorphology was not accompanied by structural nor functional changes indicative of renal insufficiency. ConclusionsWe demonstrate fetal bioaccumulation of metformin with associated fetal growth restriction and renal dysmorphology following maternal initiation of the drug within 30 days of conception in primates. Given these results and the prevalence of metformin use during pregnancy, additional investigation of any potential immediate and enduring effects of prenatal metformin use is warranted.

Effect of hypercholesterolemia on circulating and cardiomyocyte-derived extracellular vesicles

Hypercholesterolemia (HC) induces, propagates and exacerbates cardiovascular diseases via various mechanisms that are yet not properly understood. Extracellular vesicles (EVs) are involved in the pathomechanism of these diseases. To understand how circulating or cardiac-derived EVs could affect myocardial functions, we analyzed the metabolomic profile of circulating EVs, and we performed an in-depth analysis of cardiomyocyte (CM)-derived EVs in HC. Circulating EVs were isolated with Vezics technology from male Wistar rats fed with high-cholesterol or control chow. AC16 human CMs were treated with Remembrane HC supplement and EVs were isolated from cell culture supernatant. The biophysical properties and the protein composition of CM EVs were analyzed. THP1-ASC-GFP cells were treated with CM EVs, and monocyte activation was measured. HC diet reduced the amount of certain phosphatidylcholines in circulating EVs, independently of their plasma level. HC treatment significantly increased EV secretion of CMs and greatly modified CM EV proteome, enriching several proteins involved in tissue remodeling. Regardless of the treatment, CM EVs did not induce the activation of THP1 monocytes. In conclusion, HC strongly affects the metabolome of circulating EVs and dysregulates CM EVs, which might contribute to HC-induced cardiac derangements.

Histological and biochemical effects of an ethanolic extract of Myrtus communis leaf on the pancreases of rats fed high fat diets

ABSTRACT We investigated the effects of an ethanolic extract of Myrtus communis subsp. communis (MC) leaves on the pancreases of rats fed with a high fat diet (HFD). Wistar albino rats were fed either with standard lab chow (Control group) or with a 45% fat diet (HFD and HFD+MC groups) for 4 months, with the MC extract (100 mg/kg) being administered by orogastric gavage to rats in the HFD+MC group during the last month. Blood and pancreas samples were collected from all experimental groups at the end of the study. Insulin and leptin levels, and the lipid profile, were analyzed in the blood serum. Pancreatic injury was assessed histologically. Insulin, nuclear factor kappa beta (NF-κB), and alpha-smooth muscle actin (α-SMA) were assessed using immunohistochemistry. Apoptosis was assessed using terminal deoxynucleotidyl transferase dUTP nick end-labeling (TUNEL) immunohistochemistry. In addition, oxidant/antioxidant activity was analyzed by biochemical methods. Increased body weight, serum insulin and leptin levels, blood glucose level and pancreatic tissue malondialdehyde (MDA), 8-hydroxy-2’-deoxyguanosine (8-OHdG) levels, myeloperoxidase (MPO) activity and decreased tissue glutathione (GSH) level were observed in the HFD group compared to the Control group, in addition to dyslipidemia. An increased histopathological damage score, pancreatic islet area, insulin, TUNEL, NF-κB and α-SMA immunoreactivity were seen in animals from the HFD group compared to the Control group. However, such pathological changes were reduced in the HFD+MC group. Our data indicate further investigation of MC extract as a therapeutic adjuvant for HFD-induced pancreatic injury, acting via anti-inflammatory and antioxidant mechanisms, is worth carrying out.