Low-fat Control Research Articles

Targeting dopamine receptor D2 as a novel therapeutic strategy in endometrial cancer

BackgroundONC201 is a dopamine receptor D2 (DRD2) antagonist that inhibits tumor growth in preclinical models through ClpP activation to induce integrated stress response pathway and mitochondrial events related to inhibition of cell growth, which is being explored in clinical trials for solid tumors and hematological malignancies. In this study, we investigated the anti-tumorigenic effect of ONC201 in endometrial cancer cell lines and a genetically engineered mouse model of endometrial cancer.MethodsCell proliferation was assessed by MTT and colony formation assays. Cell cycle and apoptosis were evaluated by Cellometer. Invasion capacity was tested using adhesion, transwell and wound healing assays. LKB1fl/flp53fl/fl mouse model of endometrial cancer were fed a control low fat diet versus a high fat diet to mimic diet-induced obesity. Following tumor onset, mice were treated with placebo or ONC201. Metabolomics and lipidomics were used to identify the obesity-dependent effects of ONC201 in the mouse endometrial tumors. DRD2 expression was analyzed by immunohistochemistry in human endometrioid and serous carcinoma specimens. DRD2 mRNA expression from the Cancer Genome Atlas (TCGA) database was compared between the four molecular subtypes of endometrial cancer.ResultsIncreasing DRD2 expression in endometrial cancer was significantly associated with grade, serous histology and stage, as well as worse progression free survival and overall survival. Higher expression of DRD2 mRNA was found for the Copy Number High (CNH) subtype when compared to the other subtypes. ONC201 inhibited cell proliferation, induced cell cycle G1 arrest, caused cellular stress and apoptosis and reduced invasion in endometrial cancer cells. Diet-induced obesity promoted endometrial tumor growth while ONC201 exhibited anti-tumorigenic efficacy in the obese and lean LKB1fl/fl/p53fl/fl mice. Metabolomic analysis demonstrated that ONC201 reversed the obesity-driven upregulation of lipid biosynthesis and reduced protein biosynthesis in obese and lean mice.ConclusionONC201 has anti-tumorigenic effects in endometrial cancer cells and a transgenic mouse model of endometrial cancer, and DRD2 expression was documented in both human serous and endometrioid endometrial cancer. These studies support DRD2 antagonism via ONC201 as a promising therapeutic strategy for endometrial cancer that has already demonstrated pharmacodynamic activity and clinical benefit in both serous and endometrioid endometrial cancer patients.

Mediterranean Diet Maintained Platelet Count within a Healthy Range and Decreased Thrombocytopenia-Related Mortality Risk: A Randomized Controlled Trial.

There is little information on the dietary modulation of thrombosis-related risk factors such as platelet count. We aimed to assess the effects of Mediterranean diet (MedDiet) on platelet count and related outcomes in an older population at high cardiovascular risk. In participants of the PREDIMED (PREvención con DIeta MEDiterránea) study, we assessed whether an intervention with a MedDiet enriched with extra-virgin olive oil or nuts, relative to a low-fat control diet, modulated platelet count (n = 4189), the risk of developing thrombocytosis and thrombocytopenia (n = 3086), and the association between these alterations and all-cause mortality (median follow-up time: 3.0 years). Although platelet count increased over time (+0.98·109 units/L·year [95% confidence interval: 0.12; 1.84]), MedDiet interventions moderated this increase, particularly in individuals with near-high baseline count (both MedDiets combined: −3.20·109 units/L·year [−5.81; −0.59]). Thrombocytopenia incidence was lower in the MedDiet interventions (incidence rates: 2.23% in control diet, 0.91% in MedDiets combined; hazard ratio: 0.44 [0.23; 0.83]). Finally, thrombocytopenia was associated with a higher risk of all-cause mortality (hazard ratio: 4.71 [2.69; 8.24]), but this relationship was attenuated in those allocated to MedDiet (p-interaction = 0.018). In brief, MedDiet maintained platelet counts within a healthy range and attenuated platelet-related mortality in older adults at high cardiovascular risk.

Mediterranean diet and antihypertensive drug use: a randomized controlled trial.

To examine in older individuals at high cardiovascular risk whether following a Mediterranean diet decreased the necessity of antihypertensive drugs and modulated their associated cardiovascular risk. In the PREvención con DIeta MEDiterránea study, we assessed whether volunteers randomly allocated to an intervention with a Mediterranean diet enriched with extra-virgin olive oil or nuts (relative to a low-fat control diet) disclosed differences in the risk of: initiating antihypertensive medication in nonusers at baseline (n = 2188); and escalating therapy in participants using one, two, or three drugs at baseline (n = 2361, n = 1579, and n = 554, respectively). We also assessed whether allocation to Mediterranean diet modified the association between antihypertensive drug use and incident cardiovascular events. Participants allocated to Mediterranean diet interventions were associated with lower risk of initiating antihypertensive therapy [5-year incidence rates: 47.1% in the control diet, 43.0% in MedDiets; hazard ratio = 0.84, 95% CI (0.74--0.97), in a model adjusted for age, sex, and recruitment site]. Volunteers using two drugs at baseline in the Mediterranean diet intervention enriched with extra-virgin olive oil decreased their risk of therapy escalation [5-year incidence rates: 22.9% in the control diet, 20.1% in the MedDiet; hazard ratio = 0.77, 95% CI (0.60--0.99)]. Allocation to Mediterranean diet interventions attenuated the association between antihypertensive therapy at baseline and incidence of major adverse cardiovascular events (P interaction = 0.003). In an older population at high cardiovascular risk, following a Mediterranean diet reduced the risk of initiating or escalating antihypertensive medication and attenuated cardiovascular risk in antihypertensive drug users.

Weight gain and neuroadaptations elicited by high fat diet depend on fatty acid composition

Overconsumption of food is a major health concern in the western world. Palatable food has been shown to alter the activity of neural circuits, and obesity has been linked to alterations in the connectivity between the hypothalamus and cortical regions involved in decision-making and reward processing, putatively modulating the incentive value of food. Outlining neurophysiological adaptations induced by dietary intake of high fat diets (HFD) is thus valuable to establish how the diet by itself may promote overeating. To this end, C57BL/6 mice were fed HFD rich in either saturated fatty acids (HFD-S) or polyunsaturated fatty acids (HFD-P), or a low-fat control diet (LFD) for four weeks. Food and energy intake were monitored and ex vivo electrophysiology was employed to assess neuroadaptations in lateral hypothalamus (LH) and corticostriatal circuits, previously associated with food intake. In addition, the effects of dietary saturated and polyunsaturated fatty acids on the gene expression of NMDA, AMPA and GABAA receptor subunits in the hypothalamus were investigated. Our data shows that mice fed HFD-P had increased daily food and energy intake compared with mice fed HFD-S or LFD. However, this increase in energy intake had no obesogenic effects. Electrophysiological recordings demonstrated that HFD-P had a selective effect on glutamatergic neurotransmission in the LH, which was concomitant with a change in mRNA expression of AMPA receptor subtypes Gria1, Gria3 and Gria4, with no effect on the mRNA expression of NMDA receptor subtypes or GABAA receptor subtypes. Furthermore, while synaptic output from corticostriatal subregions was not significantly modulated by diet, synaptic plasticity in the form of long-term depression (LTD) was impaired in the dorsomedial striatum of mice fed HFD-S. In conclusion, this study suggests that the composition of fatty acids in the diet not only affects weight gain, but may also modulate neuronal function and plasticity in brain regions involved in food intake.

Mediterranean Diet Decreases the Initiation of Use of Vitamin K Epoxide Reductase Inhibitors and Their Associated Cardiovascular Risk: A Randomized Controlled Trial.

Our aim is to assess whether following a Mediterranean Diet (MedDiet) decreases the risk of initiating antithrombotic therapies and the cardiovascular risk associated with its use in older individuals at high cardiovascular risk. We evaluate whether participants of the PREvención con DIeta MEDiterránea (PREDIMED) study allocated to a MedDiet enriched in extra-virgin olive oil or nuts (versus a low-fat control intervention) disclose differences in the risk of initiation of: (1) vitamin K epoxide reductase inhibitors (acenocumarol/warfarin; n = 6772); (2) acetylsalicylic acid as antiplatelet agent (n = 5662); and (3) other antiplatelet drugs (cilostazol/clopidogrel/dipyridamole/ditazol/ticlopidine/triflusal; n = 6768). We also assess whether MedDiet modifies the association between the antithrombotic drug baseline use and incident cardiovascular events. The MedDiet intervention enriched with extra-virgin olive oil decreased the risk of initiating the use of vitamin K epoxide reductase inhibitors relative to control diet (HR: 0.68 [0.46–0.998]). Their use was also more strongly associated with an increased risk of cardiovascular disease in participants not allocated to MedDiet interventions (HRcontrol diet: 4.22 [1.92–9.30], HRMedDiets: 1.71 [0.83–3.52], p-interaction = 0.052). In conclusion, in an older population at high cardiovascular risk, following a MedDiet decreases the initiation of antithrombotic therapies and the risk of suffering major cardiovascular events among users of vitamin K epoxide reductase inhibitors.

Improvement of Low Fat Kashkaval Cheese Quality by Using Transglutaminase: b- The Impact on some Rheological Characteristics

Improvement of low fat Kashkaval cheese quality by using different percentages of Transglutaminase has been studied. Low fat Kashkaval cheese was made from mixture of buffalo's and cow s milk (1:1) standardized to 1% fat, treated by adding Transglutaminase (TGase) at the rate of 0.3, 0.5 and 0.7 g/Liter respectively. The resultant cheese was stored for 45 days at 10°C and 80% relative humidity. Data showed that using Transglutaminase in low fat cheese increased moisture %, TN % and fat %, while decreased acidity % as compared to low fat control cheese. Using Transglutaminase had significant positive effect on the rheological characteristic of low fat kashkaval cheese. A linear relationship was found between the concentration of TGase used and the improvement of cheese rheological properties. It could be concluded that low fat Kashkaval cheese can be made from milk treated with 0.7 g TGase/L in order to obtain cheese of acceptable texture and good body.

The Beneficial Effects of Apical Sodium-Dependent Bile Acid Transporter Inactivation Depend on Dietary Fat Composition.

ScopeThe apical sodium‐dependent bile acid transporter (ASBT, SLC10A2) is important in the enterohepatic cycling of bile acids and thereby in the intestinal absorption of lipids. ASBT inhibition has been shown to improve aspects of the metabolic syndrome, but the underlying mechanisms have remained unclear. Here, the effect of ASBT inhibition on the uptake of specific fatty acids and its consequences for diet‐induced obesity and non‐alcoholic fatty liver disease (NAFLD) are investigated.MethodsIntestinal fat absorption is determined in mice receiving an ASBT inhibitor and in Asbt−/− mice. Metabolic disease development is determined in Asbt−/− mice receiving a low‐fat control diet (LFD) or high‐fat diet (HFD) rich in saturated fatty acids (SFAs) or PUFAs.ResultsBoth ASBT inhibition and Asbt gene inactivation reduce total fat absorption, particularly of SFAs. Asbt gene inactivation lowers bodyweight gain, improves insulin sensitivity, and decreases the NAFLD activity score upon feeding a HFD rich in SFAs, but not in PUFAs.ConclusionsThe beneficial metabolic effects of ASBT inactivation on diet‐induced obesity depend on decreased intestinal absorption of SFAs, and thus on the dietary fatty acid composition. These findings highlight the importance of dietary fatty acid composition in the therapeutic effects of ASBT inhibition.

TAMI-12. CANCER STEM CELL ENRICHMENT AND METABOLIC SUBSTRATE ADAPTABILITY ARE DRIVEN BY HYDROGEN SULFIDE SUPPRESSION IN GLIOBLASTOMA

Abstract Glioblastoma (GBM) remains among the deadliest of human malignancies. The emergence of the cancer stem cell (CSC) phenotype represents a major challenge to disease management and durable treatment response. The extrinsic, environmental, and lifestyle factors that result in CSC enrichment are not well understood. The CSC state endows cells with a fluid metabolic profile, enabling the utilization of multiple nutrient sources. Therefore, to test the impact of diet on CSC enrichment, we evaluated disease progression in tumor-bearing mice fed an obesity-inducing high-fat diet (HFD) versus an energy-balanced, low-fat control diet. HFD consumption resulted in hyper-aggressive disease that was accompanied by CSC enrichment and shortened survival. HFD consumption also drove intracerebral accumulation of saturated fats, which in turn inhibited the production and signaling of the gasotransmitter hydrogen sulfide (H2S). H2S is an endogenously produced bio-active metabolite derived from sulfur amino acid catabolism. It functions principally through protein S-sulfhydration and regulates a variety of programs including mitochondrial bioenergetics and cellular metabolism. Inhibition of H2S synthesis resulted in increased proliferation and chemotherapy resistance, whereas treatment with H2S donors led to cytotoxicity and death of cultured GBM cells. Compared to non-cancerous controls, patient GBM specimens were reduced in overall protein S-sulfhydration, which was primarily lost from proteins regulating cellular metabolism. These findings support the hypothesis that diet-regulated H2S signaling serves to suppress GBM by restricting metabolic adaptability, while its loss triggers CSC enrichment and disease acceleration. Interventions augmenting H2S bioavailability concurrent with GBM standard of care may improve outcomes for GBM patients.

A Novel Combination of Fruits and Vegetables Prevents Diet-Induced Hepatic Steatosis and Metabolic Dysfunction in Mice

Epidemiological studies suggest that higher fruits and vegetables (F&V) consumption correlates with reduced risk of hepatic steatosis, yet evidence for causality and the underlying mechanisms is lacking. We aimed to determine the causal relation between F&V consumption and improved metabolic disorders in mice fed high-fat (HF) (Experiment-1) or normal-fat (Experiment-2) diets and its underlying mechanisms. Six-week-old male C57BL/6J mice were randomly grouped and fed diets supplemented at 0%-15% (wt:wt) with a freeze-dried powder composed of 24 commonly consumed F&V (human equivalent of 0-9 servings/d) for 20 wk. In Experiment-1, mice were fed an HF (45%kcal fat) diet with 0% (HF0), 5%, 10%, or 15% (HF15) F&V or a matched low-fat control diet (10%kcal fat). In Experiment-2, mice were fed an AIN-93 diet (basal) (B, 16%kcal fat) with 0% (B0), 5%, 10%, or 15% (B15) F&V supplementation. Body weight and composition, food intake, hepatic steatosis, inflammation, ceramide levels, sphingomyelinase activity, and gut microbiota were assessed. In Experiment-1, mice fed the HF15 diet had lower weight gain (17.9%), hepatic steatosis (48.4%), adipose tissue inflammation, blood (24.6%) and liver (33.9%) ceramide concentrations, and sphingomyelinase activity (38.8%) than HF0 mice (P <0.05 for all). In Experiment-2, mice fed the B15 diet had no significant changes in weight gain but showed less hepatic steatosis (28.5%), blood and adipose tissue inflammation, and lower blood (30.0%) ceramide concentrations than B0 mice (P<0.05 for all). These F&V effects were associated with favorable microbiota changes. These findings represent the first evidence for a causal role of high F&V intake in mitigating hepatic steatosis in mice. These beneficial effects may be mediated through changes in ceramide and/or gut microbiota, and suggest that higher than currently recommended servings of F&V may be needed to achieve maximum health benefits.

Mediterranean Diet and Atherothrombosis Biomarkers: A Randomized Controlled Trial

To assess whether following a Mediterranean diet (MedDiet) improves atherothrombosis biomarkers in high cardiovascular risk individuals. In 358 random volunteers from the PREvención con DIeta MEDiterránea trial, the 1-year effects on atherothrombosis markers of an intervention with MedDiet, enriched with virgin olive oil (MedDiet-VOO; n = 120) or nuts (MedDiet-Nuts; n = 119) versus a low-fat control diet (n = 119), and whether large increments in MedDiet adherence (≥3 score points, versus compliance decreases) and intake changes in key food items are associated with 1-year differences in biomarkers. Differences are observed between 1-year changes in the MedDiet-VOO intervention and control diet on the activity of platelet activating factor acetylhydrolase in high-density lipoproteins (HDLs) (+7.5% [95% confidence interval: 0.17; 14.8]) and HDL-bound α1 -antitrypsin levels (-6.1% [-11.8; -0.29]), and between the MedDiet-Nuts intervention and the control arm on non-esterified fatty acid concentrations (-9.3% [-18.1; -0.53]). Large MedDiet adherence increments are associated with less fibrinogen (-9.5% [-18.3; -0.60]) and non-esterified fatty acid concentrations (-16.7% [-31.7; -1.74]). Increases in nut, fruit, vegetable, and fatty fish consumption, and decreases in processed meat intake are linked to enhancements in biomarkers. MedDiet improves atherothrombosis biomarkers in high cardiovascular risk individuals.

Abstract P220: Differential Cardiac Geometry During Pregnancy In Lean Versus Obese Mice

Objectives: Women with a history of pregnancy complications are at increased risk for subsequent cardiovascular disease (CVD), but mechanisms linking these conditions are unclear. Obesity is associated with pregnancy complications, and is an independent risk factor for CVD. We hypothesized that obesity during pregnancy in mice would promote impaired cardiac function and/or adverse cardiac remodeling. Methods/Results: Female C57BL/6J mice (8 weeks old) were fed a high fat (HF; 60% kcal from fat) or a control low fat (LF; 10% kcal from fat) diet for 8 weeks, then were either crossed with male mice to become pregnant (P) or remained non-pregnant (NP) controls. At gestational day 18, cardiac function was quantified by echocardiography in LF- and HF-fed P and NP females. On gestational 19 day, mice were euthanized for tissue collection. HF-fed females had significantly increased body weight compared to LF-fed controls (P&lt;0.001), and body weight was increased in P compared to NP mice (P&lt;0.001; LF NP: 22.3 + 0.8 [n=20], LF P: 34.8 + 1.3 [n=10], HF NP: 32.7 + 0.9 [n=18], HF P: 39.2 + 1.1 [n=12] g). In response to pregnancy, LF-, but not HF-fed, mice had significantly increased LV mass (LF NP: 60.0 + 2.4, LF P: 75.5 + 3.1, HF NP: 70.3 + 2.0, HF P: 75.1 + 3.8 mg; P&lt;0.01) and LV end-diastolic diameter (LVED; LF NP: 3.60 + 0.05, LF P: 3.95 + 0.07, HF NP: 3.67 + 0.05, HF P: 3.73 + 0.01 mm; P&lt;0.01) compared to NP controls. In contrast, HF-fed pregnant mice had smaller LVED (P&lt;0.01) and increased relative wall thickness (RWT; LF NP: 0.36 + 0.01, LF P: 0.35 + 0.01, HF NP: 0.39 + 0.01, HF P: 0.40 + 0.02 [2* LV posterior wall thickness/LV end-diastolic diameter], P&lt;0.05) compared to LF-fed pregnant mice. Preliminary gene expression analysis indicate mRNA abundance of TGFb1 is elevated by 50% (P&lt;0.05) hearts of HF-fed compared to LF-fed pregnant mice. Conclusions: These data suggest obesity during pregnancy promotes concentric remodeling, versus eccentric remodeling in lean mice. These data are clinically meaningful, as they suggest that obesity during pregnancy may promote pathological cardiac remodeling, which may contribute to subsequent maternal risk for cardiovascular disease.

PCSK9 Deficiency Modulates Glucose Homeostasis and Insulin Secretion Via Pancreatic Ldl Receptors And Cholesterol Accumulation

Intracellular lipid accumulation contributes to β-cell dysfunction in type 2 diabetes. How PCSK9 interacts with lipid metabolism to alter insulin availability/resistance is not well studied. Genetic deficiency of PCSK9 mice was fed a high fat (HFD; 45% Kcal from fat) for 14 weeks, and the metabolic phenotype including glucose, insulin, GTT, ITT, and cholesterol was examined. In addition, the expression of PCSK9 and LDLR was were examined by RT-qPCR, and immunohistochemistry. We provide data supporting a role for PCSK9 in pancreatic islets. In HFD mice, PCSK9 deletion led to impaired glucose tolerance and decreased insulin secretion. Pancreatic islets of HFD PCSK9-/ mice showed impaired insulin production and increased LDLR mRNA. Cholesterol was higher in islets of both HFD-fed WT and PCSK9 KO mice compared to low-fat diet controls. Glucose intolerance in PCSK9 deficiency was reversed when the LDLR was also lacking. Further, islets isolated from PCSK-/-LDR-/- mice showed improved insulin secretion. Our findings support a tissue-specific role for PCSK9 in pancreatic cholesterol homeostasis such that PCSK deficiency may accentuate LDLR-dependent β-cell dysfunction.

Effect of shengmai san on insulin resistance, tumor necrosis factor-alpha and oxidative stress in rats fed on a high-fat diet

Shengmai San (SMS) is a well-known traditional Chinese medicine formula prescribed for patients with coronary heart disease (CHD). Insulin resistance is the most likely explanation for the development and progression of this disease. To investigate the effect of SMS on high-fat diet-induced insulin resistance, inflammatory cytokine production and oxidative stress, male Wistar rats were fed with low-fat control diet, high-fat diet and high-fat diet supplemented with 4% SMS for eight weeks. An oral glucose tolerance test was conducted in the seventh week. Rats fed the SMS diet had significantly lower plasma fructosamine concentration (p < 0.05) and tended to have lower (p < 0.1) AUC values of plasma glucose and insulin concentrations after glucose challenge, compared to those fed the high-fat diet. Moreover, SMS reduced TNF-α and lipid peroxidation levels in the liver and heart. However, SMS had no effect on fasting plasma concentrations of glucose, insulin, triglyceride, non-esterified fatty acids, total cholesterol, HDL-cholesterol and insulin resistance index (homeostasis model assessment; HOMA) in rats. Our results show that SMS may have little or no significant effect on reducing insulin resistance, but display anti-oxidative and anti-inflammatory properties.

Effects of diets high in corn oil or in extra virgin olive oil on oxidative stress in an experimental model of breast cancer.

Experimental evidence highlights the importance of dietetic factors on breast cancer. In this work we aimed to analyze the effects two oils, corn oil (rich in n-6 polyunsaturated fatty acids -PUFA-) and extra virgin olive oil (EVOO), on oxidative stress in an animal model of breast carcinogenesis. Female rats were fed a low-fat control, a high-corn oil, or a high-EVOO diet from weaning or after induction with 7,12-dimethylbenz[a]anthracene at 53 days. Animals were euthanized at 36, 51, 100 and 246 days of age. We analyzed antioxidant enzymes (mRNA and activity of superoxide dismutase, glutathione peroxidase and catalase), non-enzymatic capacity (oxidized and reduced glutathione) and DNA damage (8-oxo-dG) in tumors and mammary gland at different ages. We also analyzed lipid peroxidation (isoprostanes in serum and lipofuscin in liver). Results indicated a decrease in the enzymatic antioxidant capacity and increased oxidative stress in mammary gland of healthy young animals after a short period of high-fat diets intake, followed by an adaptation to chronic dietary intervention. After induction both diets, especially the one high in n-6 PUFA, increased the oxidized glutathione. In tumors no clear effects of the high-fat diets were observed, although in the long-term lipofuscin and 8-oxo-dG suggested greater oxidative damage by effect of the n-6 PUFA-rich diet. Considering the differential effects of these diets on mammary carcinogenesis that we have previously reported, this study suggests that these high-fat diets could have an effect on oxidative stress that would lead to different signaling pathways.

Early life stress reduces voluntary exercise and its prevention of diet-induced obesity and metabolic dysfunction in mice.

The development of obesity-related metabolic syndrome (MetS) involves a complex interaction of genetic and environmental factors. One environmental factor found to be significantly associated with MetS is early life stress (ELS). We have previously reported on our mouse model of ELS, induced by neonatal maternal separation (NMS), that displays altered regulation of the hypothalamic-pituitary-adrenal (HPA) axis and increased sensitivity in the urogenital organs, which was attenuated by voluntary wheel running. Here, we are using our NMS model to determine if ELS-induced changes in the HPA axis also influence weight gain and MetS. Naïve (non-stressed) and NMS male mice were given free access to a running wheel and a low-fat control diet at 4-weeks of age. At 16-weeks of age, half of the mice were transitioned to a high fat/sucrose (HFS) diet to investigate if NMS influences the effectiveness of voluntary exercise to prevent diet-induced obesity and MetS. Overall, we observed a greater impact of voluntary exercise on prevention of HFS diet-induced outcomes in naïve mice, compared to NMS mice. Although body weight and fat mass were still significantly higher, exercise attenuated fasting insulin levels and mRNA levels of inflammatory markers in epididymal adipose tissue in HFS diet-fed naïve mice. Only moderate changes were observed in exercised NMS mice on a HFS diet, although this could partially be explained by reduced running distance within this group. Interestingly, sedentary NMS mice on a control diet displayed impaired glucose homeostasis and moderately increased pro-inflammatory mRNA levels in epididymal adipose, suggesting that early life stress alone impairs metabolic function and negatively impacts the therapeutic effect of voluntary exercise.

1820-P: A Direct AMPK Activator Reduces Liver Steatosis in a Mouse Model of NASH

Nonalcoholic fatty liver disease (NAFLD), is associated with metabolic disease and estimated to affect 25% of adults. NAFLD can progress to nonalcoholic steatohepatitis (NASH) which may lead to cirrhosis and hepatocellular carcinoma. There is no current treatment or cure for NASH. AMPK (5’AMP-activated protein kinase) activators have shown potential for treating NAFLD/NASH due to their effects on fatty acid inhibition and cholesterol synthesis. To evaluate this potential, an AMPK activator (herein known as C455) was selected for a preclinical NASH study. C455 demonstrated potent and selective activation of AMPK (cellular EC50= 85 nM). X-ray crystallography showed the compound binds at the ADaM site, and pharmacokinetics determined a bioavailability of 45% and half-life of 3h. Mice (male ob/ob on Amylin-NASH diet for 14-weeks or low-fat control) were dosed orally with vehicle or C455 at 3 or 30mg/kg daily. After 6 weeks of treatment, plasma ALT and terminal liver weight were decreased 32% and 22% respectively in mice treated with 30mg/kg C455 vs. vehicle. This dose also decreased liver lipid from 33% to 23%. Fibrosis via collagen staining did not improve with treatment, but significant decreases in mRNA transcripts for Col1a1(62%), Col1a2(59%), Col4a1(56%), and Timp1(55%) were seen with the high dose. Histology showed dose dependent increases in heart weight relative to body weight, and increased expression of genes associated with cardiac hypertrophy. Mice given 30mg/kg C455 also had increased cardiac glycogen storage compared to vehicle treated or LFD control mice. Our study results show that systemic activation of AMPK in a diet-induced mouse model of NASH reduces plasma ALT, decreases liver lipid, and suppressed hepatic collagen gene expression but shows no histological improvement in fibrosis. Coupled with increased heart weight and glycogen storage, the data suggests that current small molecule activators of AMPK may not be viable therapies for the treatment of NASH-related fibrotic disease. Disclosure K.M. Mather: None. M.L. Boland: None. E. Rivers: None. A. Srivastava: None. M. Schimpl: Employee; Self; AstraZeneca. Stock/Shareholder; Self; AstraZeneca. J. Robinson: None. P.T.C. Wan: None. J.L. Hansen: None. J.A. Read: Employee; Self; AstraZeneca. Stock/Shareholder; Self; AstraZeneca. J.L. Trevaskis: Employee; Self; Gilead Sciences, Inc. D.M. Smith: Employee; Self; AstraZeneca.

Herring Milt and Herring Milt Protein Hydrolysate Reduce Weight Gain and Improve Insulin Sensitivity and Pancreatic Beta-Cell Function in Diet-Induced Obese Mice

Abstract Objectives The present study was designed to examine the effect of herring milt dry powder (HMDP) on glucose homeostasis and related metabolic phenotypes and compare its efficacy with herring milt protein hydrolysate (HMPH) in diet-induced obese and insulin resistant mice. Methods Male C57BL/6 J mice were pretreated with a high-fat diet for 7 weeks were divided into 3 groups where one group continued on the high-fat diet and used as the obese and insulin resistant control (HFC) and the other two groups were fed a modified HFC diet where 70% of casein was replaced with an equal percentage of protein derived from HMDP or HMPH. A group of mice fed a low-fat diet all the time was used as the normal or low-fat control (LFC). Body weight was obtained weekly and food intake was recorded daily. Semi-fating (4–6 hr) blood glucose was measured every other week using a glucometer using the blood from tail vein. Oral glucose tolerance was measured twice during weeks 5 and 9, respectively, and insulin tolerance was determined during week 7 of the treatment. At the end of the experiment, serum was obtained following overnight fasting for the measurement of fasting insulin, leptin, free fatty acids and lipids as well as other glucose metabolism-related biomarkers. Results During the 9-week treatment period, mice on the high-fat diet maintained significantly higher body weight and semi-fasting blood glucose levels and exhibited impaired oral glucose tolerance and insulin resistance relative to mice on the low-fat diet. At the end of the study, the analysis of fasting blood samples revealed that mice on the high-fat diet had increases in serum insulin, leptin, free fatty acids and cholesterol levels. Mice fed the high-fat diet also showed an increase in insulin resistance index and a decrease in β-cell function index. Compared to mice on the high-fat diet, the 70% replacement of dietary casein with an equal percentage of protein derived from HMDP or HMPH reversed or markedly improved these parameters, and HMDP and HMPH showed similar effects. Conclusions The results demonstrate that replacing dietary casein with the same amount of protein derived from either HMDP or HMPH prevents and improves high-fat-diet-induced obesity and insulin resistance. Funding Sources Atlantic Canada Opportunity Agency through the Atlantic Innovation Fund grant (no. 193,594) and National Research Council of Canada – NHP program.

Dietary β-Hydroxy-β-Methylbutyrate Supplementation Promotes PDAC Response to Gemcitabine and Immunotherapy Response in Obese Mice

Abstract Objectives Late diagnosis, aggressive underlying biology, and limited treatment options contribute towards the exceptionally poor survival of pancreatic ductal adenocarcinoma (PDAC). Obesity promotes both incidence and progression of PDAC via chronic systemic inflammation, generation of an immunosuppressive tumor microenvironment, and promotion of tumor fibrosis. β-Hydroxy-β-Methylbutyrate (HMB) reduces cancer associated cachexia and promotes modest reductions in tumor growth in animal models of cancer. The objective of this study was to determine if HMB supplementation would alter therapeutic response to either gemcitabine or anti-PD1 immunotherapy. Methods C57BL/6 mice were fed either a diet induced obesity high fat diet or a matched low fat control. Following PANC02 tumor induction, animals were treated with HMB alone or in combination with gemcitabine or anti-PD1 immunotherapy. Tumor transcriptomic analysis was preformed using Affymetrix microarray, with subsequent gene set enrichment analysis. Immunohistochemistry was performed for CD3 (a T cell marker). C57BL/6 mice were fed either a diet induced obesity high fat diet or a matched low fat control. Following PANC02 tumor induction, animals were treated with HMB alone or in combination with gemcitabine or anti-PD1 immunotherapy. Tumor transcriptomic analysis was preformed using Affymetrix microarray, with subsequent gene set enrichment analysis. Immunohistochemistry was performed for CD3 (a T cell marker). Results DIO-induced immune suppression was partially reversed by HMB, with reduced tumor growth, increased T cell markers and enhanced efficacy of gemcitabine following HMB treatment in obese mice. Separately, HMB enhanced the efficacy of anti-PD1 immunotherapy. Conclusions HMB enhanced PDAC immune surveillance, augmenting both cytotoxic chemotherapy and immunotherapy. HMB-induced suppression of obesity driven PDAC tumor growth, and promotion of immune surveillance may provide extend the therapeutic index of both chemotherapies and immunotherapies in PDAC. Funding Sources This study was supported by a grant from the National Cancer Institute (R35 CA197627) to SDH.

Obesity-Associated Leptin Signaling Promotes Chemotherapy Resistance in Basal-Like Breast Cancer: The Role of Tumor-Associated Macrophages

Abstract Objectives Obesity is associated with a reduced response to cytotoxic chemotherapies. Given that the adipokine leptin has been shown to promote macrophage production of pro-tumor cytokines, we hypothesized that leptin-induced pro-tumor macrophage polarization is a key mediator of obesity-associated docetaxel resistance. Methods Wild-type C57BL/6 mice were fed a low-fat control (10% kcal from fat) or high-fat diet-induced obesity (DIO; 60% kcal from fat) diet for 15 weeks, then orthotopically injected with 2 mouse mammary tumor cell lines: EWnt-S (scrambled shRNA) and EWnt-L (shRNA to the leptin receptor) in the 9th and 4th mammary glands, respectively. Mice in each group were then randomized to vehicle or docetaxel (20 mg/kg/week IV for 3 weeks). The transcriptomes of both tumors (n = 6/group) were analyzed by microarray, and cell type enrichment analysis performed using the webtool xCell. The in vitro effects of leptin +/− docetaxel on macrophage polarization as well as cancer cell cytotoxicity and markers of invasive capacity are currently being assessed. Results The DIO mice had greater body weight and % body fat at euthanization versus control mice (P &amp;lt; 0.01 for both). Docetaxel treatment in DIO mice reduced tumor growth rate in EWnt-L tumors (P &amp;lt; 0.05), but not EWnt-S tumors. In contrast, docetaxel treatment reduced the EWnt-L and EWnt-S tumor growth rate in control mice (P &amp;lt; 0.05 for both). xCell analysis indicated that docetaxel treatment increased M2 macrophage levels in the DIO mouse tumors, but not the controls. Leptin receptor knockdown attenuated this effect. In addition, key genes related to “macrophage markers”, “cytokines and inflammatory response”, and “interleukin (IL)-6 signaling” were differentially expressed in docetaxel-treated EWnt-S tumors from DIO versus control mice. Conclusions These results indicate that leptin signaling mediates obesity-induced docetaxel resistance in the E-Wnt model of basal-like breast cancer. They also suggest that docetaxel interacts with leptin signaling to promote an increase in tumor-promoting M2 macrophages and subsequent docetaxel resistance. This hypothesis is currently being explored through our in vitro studies. Funding Sources This work was supported by a grant from the Developmental Funding Program, Lineberger Comprehensive Cancer Center, UNC Chapel Hill.

Exercise-stimulated resolvin biosynthesis in adipose tissue is abrogated by obesity-induced adrenergic dysfunction

Abstract Adipose tissue (AT) inflammation is associated with obesity-induced insulin resistance. Exercise (Exe) prevents the development of chronic inflammatory diseases and insulin resistance largely through unknown mechanisms. We provide data that specialized proresolving lipid mediators (SPM) contribute to the anti-inflammatory effect of Exe. Compared with sedentary controls (Sed), mice exposed to 4 wk of Exe showed elevated AT expression of SPM biosynthetic enzyme Alox15 (Sed 1.12±0.2 vs Exe 1.68±0.1 relative expression, n=5–7), SPM levels (Sed vs Exe; RvD1 156.7±12.0 vs 223.0±24.7; 17R-RvD1 354.0±35.9 vs 534.4±98.9; RvD4 31.31±6.9 vs 123.1±30.7; and 17R-RvD3 2.92±1.1 vs 8.20±1.7 pg/g, n=5–7), and M2 macrophages (F4/80+CD301+; Sed 17.5±3.2 vs Exe 40.9±4.5 %F4/80+, n=4–6). These changes were diet-dependent as high fat (HFD) abrogated this Exe effect in AT when compared with low fat (LFD) controls. The exercise-stimulated Alox15 expression was restricted to AT macrophages (F4/80+ cells; LFD Sed 0.11±0.03 vs LFD Exe 0.23±0.03; HFD Sed 0.10±0.04 vs HFD Exe 0.08±0.02 relative expression, n=3–6), as adipocytes isolated from Exe mice showed no difference compared with Sed controls. That epinephrine (Epi) is released with exercise and stimulates macrophage Alox15 expression and RvD1 production, we questioned if Epi biosynthesis is affected by HFD. HFD feeding diminished expression of phenylethanolamine N-methyltransferase compared with controls (LFD 1.0±0.25 vs HFD 0.36±0.15, n=4). Similarly, obese-diabetic mice (db/db) also showed reduced catecholamine production and biosynthetic enzyme levels. These results suggest that obesity-induced adrenergic dysfunction abrogates exercise-stimulated resolvin production in AT.