Parental obesity alters offspring blood pressure regulation and cardiovascular responses to stress: role of P2X7R and sex differences.

We examined the impact of parental obesity on offspring blood pressure (BP) regulation and cardiovascular responses to stress. Offspring from normal (N) diet-fed C57BL/6J parents were fed either N (NN) or a high-fat (H) diet (NH) from weaning until adulthood. Offspring from obese H diet-fed parents were also fed N (HN) or H diet (HH). Body weight, calorie intake, and fat mass were measured at 22 wk of age when cardiovascular phenotyping was performed. Male and female HH offspring were 15% heavier than NH and 70% heavier than NN offspring. Male HH and HN offspring had elevated BP (121 ± 2 and 115 ± 1 mmHg, by telemetry) compared with male NH and NN offspring (108 ± 6 and 107 ± 3 mmHg, respectively) and augmented BP responses to angiotensin II, losartan, and hexamethonium. Male HH and HN offspring also showed increased BP responses to air-jet stress (37 ± 2 and 38 ± 2 mmHg) compared with only 24 ± 3 and 25 ± 3 mmHg in NH and NN offspring. Baseline heart rate (HR) and HR responses to air-jet stress were similar among groups. In females, BP and cardiovascular responses to stress were similar among all offspring. Male H diet-fed offspring from obese H diet-fed purinoreceptor 7-deficient (HH-P2X7R-KO) parents had normal BP that was similar to control NN-P2X7R-KO offspring from lean parents. These results indicate that parental obesity leads to increased BP and augmented BP responses to stress in their offspring in a sex-dependent manner, and the impact of parental obesity on male offspring BP regulation is markedly attenuated in P2X7R-KO mice.

Dimethyl fumarate preserves left ventricular infarct integrity following myocardial infarction via modulation of cardiac macrophage and fibroblast oxidative metabolism.

Myocardial infarction (MI) is one of the leading causes of mortality and cardiovascular disease worldwide. MI is characterized by a substantial inflammatory response in the infarcted left ventricle (LV), followed by transition of quiescent fibroblasts to active myofibroblasts, which deposit collagen to form the reparative scar. Metabolic shifting between glycolysis and mitochondrial oxidative phosphorylation (OXPHOS) is an important mechanism by which these cell types transition towards reparative phenotypes. Thus, we hypothesized that dimethyl fumarate (DMF), a clinically approved anti-inflammatory agent with metabolic actions, would improve post-MI remodeling via modulation of macrophage and fibroblast metabolism. Adult male C57BL/6J mice were treated with DMF (10 mg/kg) for 3-7 days after MI. DMF attenuated LV infarct and non-infarct wall thinning at 3 and 7 days post-MI, and decreased LV dilation and pulmonary congestion at day 7. DMF improved LV infarct collagen deposition, myofibroblast activation, and angiogenesis at day 7. DMF also decreased pro-inflammatory cytokine expression (Tnf) 3 days after MI, and decreased inflammatory markers in macrophages isolated from the infarcted heart (Hif1a, Il1b). In fibroblasts extracted from the infarcted heart at day 3, RNA-Seq analysis demonstrated that DMF promoted an anti-inflammatory/pro-reparative phenotype. By Seahorse analysis, DMF did not affect glycolysis in either macrophages or fibroblasts at day 3, but enhanced macrophage OXPHOS while impairing fibroblast OXPHOS. Our results indicate that DMF differentially affects macrophage and fibroblast metabolism, and promotes anti-inflammatory/pro-reparative actions. In conclusion, targeting cellular metabolism in the infarcted heart may be a promising therapeutic strategy.

Sex differences in the impact of parental obesity on offspring cardiac SIRT3 expression, mitochondrial efficiency, and diastolic function early in life.

Previous studies suggest that parental obesity may adversely impact long-term metabolic health of the offspring. We tested the hypothesis that parental (paternal + maternal) obesity impairs cardiac function in the offspring early in life. Within 1-3 days after weaning, offspring from obese rats fed a high-fat diet (HFD-Offs) and age-matched offspring from lean rats (ND-Offs) were submitted to echocardiography and cardiac catheterization for assessment of pressure-volume relationships. Then, hearts were digested and isolated cardiomyocytes were used to determine contractile function, calcium transients, proteins related to calcium signaling, and mitochondrial bioenergetics. Female and male HFD-Offs were heavier (72 ± 2 and 61 ± 4 g vs. 57 ± 2 and 49 ± 1 g), hyperglycemic (112 ± 8 and 115 ± 12 mg/dL vs. 92 ± 10 and 96 ± 8 mg/dL) with higher plasma insulin and leptin concentrations compared with female and male ND-Offs. When compared with male controls, male HFD-Offs exhibited similar systolic function but impaired diastolic function as indicated by increased IVRT (22 ± 1 vs. 17 ± 1 ms), E/E' ratio (29 ± 2 vs. 23 ± 1), and tau (5.7 ± 0.2 vs. 4.8 ± 0.2). The impaired diastolic function was associated with reduced resting free Ca2+ levels and phospholamban protein expression, increased activated matrix metalloproteinase 2, and reduced SIRT3 protein expression, mitochondrial ATP reserve, and ATP-linked respiration. These results indicate that male and female Offs from obese parents have multiple metabolic abnormalities early in life (1-3 days after weaning) and that male, but not female, Offs have impaired diastolic function as well as reductions in cardiac SIRT3, resting free Ca2+ levels, and mitochondrial biogenesis.NEW & NOTEWORTHY Parental obesity contributes to diastolic dysfunction in young offspring (1-3 days after weaning) in a sex-dependent manner, as well as reduced cardiac SIRT3 expression and altered mitochondrial bioenergetics, resting Ca2+ levels, and reduced phospholamban protein levels.

Impact of leptin deficiency compared with neuronal-specific leptin receptor deletion on cardiometabolic regulation.

The main goal of this study was to compare the impact of total body leptin deficiency with neuronal-specific leptin receptor (LR) deletion on metabolic and cardiovascular regulation. Liver fat, diacylglycerol acyltransferase-2 (DGTA2), and CD36 protein content were measured in wild-type (WT), nervous system LR-deficient (LR/Nestin-Cre), and leptin deficient (ob/ob) mice. Blood pressure (BP) and heart rate (HR) were recorded by telemetry, and motor activity (MA) and oxygen consumption (V̇o2) were monitored at 24 wk of age. Female and male LR/Nestin-Cre and ob/ob mice were heavier than WT mice (62 ± 5 and 61 ± 3 vs. 31 ± 1 g) and hyperphagic (6.2 ± 0.5 and 6.1 ± 0.7 vs. 3.5 ± 1.0 g/day), with reduced V̇o2 (27 ± 1 and 33 ± 1 vs 49 ± 3 ml·kg-1·min-1) and decreased MA (3 ± 1 and 7 ± 2 vs 676 ± 105 cm/h). They were also hyperinsulinemic and hyperglycemic compared with WT mice. LR/Nestin-Cre mice had high levels of plasma leptin, while ob/ob mice had undetectable leptin levels. Despite comparable obesity, LR/Nestin-Cre mice had lower liver fat content, DGTA2, and CD36 protein levels than ob/ob mice. Male WT, LR/Nestin-Cre, and ob/ob mice exhibited similar BP (111 ± 3, 110 ± 1 and 109 ± 2 mmHg). Female LR/Nestin-Cre and ob/ob mice, however, had higher BP than WT females despite similar metabolic phenotypes compared with male LR/Nestin-Cre and ob/ob mice. These results indicate that although nervous system LRs play a crucial role in regulating body weight and glucose homeostasis, peripheral LRs regulate liver fat deposition. In addition, our results suggest potential sex differences in the impact of obesity on BP regulation.

Role of SOCS3 in POMC neurons in metabolic and cardiovascular regulation.

Suppressor of cytokine signaling 3 (SOCS3) is a negative regulator of leptin signaling. We previously showed that the chronic effects of leptin on blood pressure (BP) and glucose regulation are mediated by stimulation of proopiomelanocortin (POMC) neurons. In this study we examined the importance of endogenous SOCS3 in POMC neurons in control of metabolic and cardiovascular function and potential sex differences. Male and female SOCS3flox/flox/POMC-Cre mice in which SOCS3 was selectively deleted in POMC neurons and control SOCS3flox/flox mice were studied during a control diet (CD) or a high-fat diet (HFD) and during chronic leptin infusion. Body weight was lower in male and female SOCS3flox/flox/POMC-Cre than control mice fed the CD, despite similar food intake. Male SOCS3flox/flox/POMC-Cre mice exhibited increased energy expenditure. BP and heart rate were similar in male and female SOCS3flox/flox/POMC-Cre and control mice fed the CD. HFD-fed male and female SOCS3flox/flox/POMC-Cre mice showed attenuated weight gain. HFD-induced elevations in baseline BP and BP responses to an air-jet stress test were greater in female SOCS3flox/flox/POMC-Cre than control mice. Chronic leptin infusion produced similar responses for food intake, body weight, oxygen consumption, blood glucose, BP, and heart rate in all groups. Thus SOCS3 deficiency in POMC neurons influences body weight regulation in the setting of CD and HFD and differentially affects BP and energy balance in a sex-specific manner but does not amplify the dietary, glycemic, or cardiovascular effects of leptin.

Role of autonomic nervous system in chronic CNS-mediated antidiabetic action of leptin.

This study tested whether ganglionic blockade or hepatic vagotomy attenuates the chronic central nervous system (CNS)-mediated antidiabetic and cardiovascular effects of leptin. Male Sprague-Dawley rats were instrumented with telemetry probes and arterial and venous catheters for determination of blood pressure (BP), heart rate (HR), blood sampling, and intravenous (iv) infusions. An intracerebroventricular (ICV) cannula was placed into the brain lateral ventricle for infusion of leptin or vehicle. After control measurements, streptozotocin (STZ) was injected iv (50 mg/kg) to induce diabetes, and 5 days later leptin (n = 6) or saline vehicle (n = 5) was infused ICV for 12 days via osmotic pumps. Beginning on day 6 of leptin treatment, the ganglionic blocker hexamethonium (15 mg·kg-1·day-1 iv) was infused, while leptin infusion was continued, to assess the role of the autonomic nervous system. Induction of diabetes was associated with increases in blood glucose (98 ± 7 to 350 ± 19 mg/dl), food intake (23 ± 3 to 43 ± 3 g/day), decreases in HR (-70 ± 11 beats/min), polyuria, and increased water consumption, which were all completely normalized by ICV leptin infusion. Although hexamethonium attenuated leptin's effect on HR, it failed to impair leptin's ability to restore euglycemia or to prevent the polyuria or increased water intake in STZ-diabetic rats. We also found that after pretreatment with hexamethonium (n = 8), ICV leptin infusion, during continued ganglionic blockade, completely normalized blood glucose in diabetic rats. In addition, selective hepatic vagotomy did not attenuate leptin's ability to restore euglycemia in diabetic rats. These results suggest that leptin's powerful chronic CNS antidiabetic actions are mediated primarily via nonautonomic mechanisms.

Temporal changes in glucose metabolism reflect polarization in resident and monocyte-derived macrophages after myocardial infarction.

Introduction: Metabolic reprogramming from glycolysis to the mitochondrial tricarboxylic acid (TCA) cycle and oxidative phosphorylation may mediate macrophage polarization from the pro-inflammatory M1 to the anti-inflammatory M2 phenotype. We hypothesized that changes in cardiac macrophage glucose metabolism would reflect polarization status after myocardial infarction (MI), ranging from the early inflammatory phase to the later wound healing phase. Methods: MI was induced by permanent ligation of the left coronary artery in adult male C57BL/6J mice for 1 (D1), 3 (D3), or 7 (D7) days. Infarct macrophages were subjected to metabolic flux analysis or gene expression analysis. Monocyte versus resident cardiac macrophage metabolism was assessed using mice lacking the Ccr2 gene (CCR2 KO). Results: By flow cytometry and RT-PCR, D1 macrophages exhibited an M1 phenotype while D7 macrophages exhibited an M2 phenotype. Macrophage glycolysis (extracellular acidification rate) was increased at D1 and D3, returning to basal levels at D7. Glucose oxidation (oxygen consumption rate) was decreased at D3, returning to basal levels at D7. At D1, glycolytic genes were elevated (Gapdh, Ldha, Pkm2), while TCA cycle genes were elevated at D3 (Idh1 and Idh2) and D7 (Pdha1, Idh1/2, Sdha/b). Surprisingly, Slc2a1 and Hk1/2 were increased at D7, as well as pentose phosphate pathway (PPP) genes (G6pdx, G6pd2, Pgd, Rpia, Taldo1), indicating increased PPP activity. Macrophages from CCR2 KO mice showed decreased glycolysis and increased glucose oxidation at D3, and decreases in Ldha and Pkm2 expression. Administration of dichloroacetate, a pyruvate dehydrogenase kinase inhibitor, robustly decreased pyruvate dehydrogenase phosphorylation in the non-infarcted remote zone, but did not affect macrophage phenotype or metabolism in the infarct zone. Discussion: Our results indicate that changes in glucose metabolism and the PPP underlie macrophage polarization following MI, and that metabolic reprogramming is a key feature of monocyte-derived but not resident macrophages.

Restoration of Cardiac Function After Myocardial Infarction by Long-Term Activation of the CNS Leptin-Melanocortin System.

Heart failure has a high mortality rate, and current therapies offer limited benefits. The authors demonstrate that activation of the central nervous system leptin-melanocortin pathway confers remarkable protection against progressive heart failure following severe myocardial infarction. The beneficial cardiac-protective actions of leptin require activation of brain melanocortin-4 receptors and elicit improvements in cardiac substrate oxidation, cardiomyocyte contractility, Ca2+ coupling, and mitochondrial efficiency. These findings highlight a potentially novel therapeutic approach for myocardial infarction and heart failure.

Interaction of Obesity and Hypertension on Cardiac Metabolic Remodeling and Survival Following Myocardial Infarction.

Background Obesity and hypertension are risk factors for myocardial infarction (MI); however, their potential interactions on post-MI outcomes are unclear. We examined interactions of obesity and hypertensionon post-MI function, remodeling, metabolic changes, and recovery. Methods and Results Male and female C57BL/6J mice were provided standard chow or high-fat/fructose diet for 8 weeks and then infused with angiotensin II for 2 weeks to induce hypertension. MI was then induced by surgical ligation of the left coronary artery for 7 days. Obesity alone did not cause cardiac injury or exacerbate hypertension-induced cardiac dysfunction. After MI, however, obese-normotensive mice had lower survival rates compared with chow-fed mice (56% versus 89% males; 54% versus 75% females), which were further decreased by hypertension (29% males; and 35% females). Surviving obese-normotensive males displayed less left ventricular dilation and pulmonary congestion compared with chow-fed males after MI; hypertension reversed left ventricular dilation because of high-fat/fructose diet and promoted significant pulmonary congestion compared with chow-fed controls. Obese-normotensive males displayed higher left ventricular α-MHC (alpha-myosin heavy chain) protein, phosphorylated Akt (protein kinase B) and AMPK (adenosine-monophosphate activated kinase), PPAR-γ (peroxisome proliferator activated receptor gamma), and plasma adiponectin levels after MI, indicating favorable contractile and metabolic changes. However, these favorable contractile and metabolic changes were attenuated by hypertension. Obese-hypertensive males also had lower levels of collagen in the infarcted region, indicating decreased ability to promote an adaptive wound healing response to MI. Conclusions Obesity reduces post-MI survival but is associated with improved post-MI cardiac function and metabolism in surviving normotensive mice. When hypertension accompanies obesity, favorable metabolic pathways associated with obesity are attenuated and post-MI cardiac function and remodeling are adversely impacted.

Role of melanocortin 4 receptor in hypertension induced by chronic intermittent hypoxia.

AIM: We previously demonstrated that central nervous system (CNS) melanocortin 4 receptors (MC4R) play a key role in regulating blood pressure (BP) in some conditions associated with increased SNS activity, including obesity. In this study, we examined whether activation of CNS MC4R contributes to chronic intermittent hypoxia (CIH)-induced hypertension and ventilatory responses to hypercapnia. METHODS: Rats were instrumented with an intracerebroventricular (ICV) cannula in the lateral cerebral ventricle for continuous infusion of MC4R antagonist (SHU-9119) and telemetry probes for measuring mean arterial pressure (MAP) and heart rate (HR). Untreated and SHU-9119-treated rats as well as obese and lean MC4R-deficient rats were exposed to CIH for 7-18 consecutive days. RESULTS: Chronic intermittent hypoxia reduced cumulative food intake by 18 ± 5 g while MAP and HR increased by 10 ± 3 mm Hg and 9 ± 5 bpm in untreated rats. SHU-9119 increased food intake (from 15 ± 1 to 46 ± 3 g) and prevented CIH-induced reduction in food intake. CIH-induced hypertension was not attenuated by MC4R antagonism (average increase of 10 ± 1 vs 9 ± 1 mm Hg for untreated and SHU-9119 treated rats). In obese MC4R-deficient rats, CIH for 7 days raised BP by 11 ± 4 mm Hg. However, when MC4R-deficient rats were food restricted to prevent obesity, CIH-induced hypertension was attenuated by 32%. We also found that MC4R deficiency was associated with impaired ventilatory responses to hypercapnia independently of obesity. CONCLUSION: These results show that obesity and the CNS melanocortin system interact in complex ways to elevate BP during CIH and that MC4R may be important in the ventilatory responses to hypercapnia.

Increased sleep time and reduced energy expenditure contribute to obesity after ovariectomy and a high fat diet.

In this study we examined if sleep time, caloric intake and energy expenditure are important contributors to development of ovariectomy-induced obesity in mice fed control or high fat diet (HFD). Twelve female mice at 6 weeks of age were divided into 2 groups: Sham (n = 5) and ovariectomized (OVX, n = 7). Mice were fed control diet for 9 weeks and shifted to HFD for additional 9 weeks. Food intake and body weight were measured daily and body composition was measured weekly by EchoMRI. Energy expenditure (EE), oxygen consumption (VO2), motor activity (MA) and sleep time were monitored at week 9 during control diet and HFD. OVX did not alter caloric intake, body weight or body composition, MA, sleep time or fasting blood glucose, but slightly reduced EE compared to Sham mice on control diet. After HFD feeding, OXV mice had similar caloric intake, lean mass, MA, and blood glucose levels but had significantly greater weight gain (8.2 ±â€¯1.0 vs. 4.8 ±â€¯1.2 g, p < 0.05), increased fat mass and sleep time, and reduced EE (3.3 ±â€¯0.4 vs. 5.5 ±â€¯0.2 kcal/h) and VO2 (1.12 ±â€¯0.01 vs. 1.83 ±â€¯0.05 ml/min) compared to Sham group. Daytime blood pressure was higher while nighttime heart rate was lower in OVX group. These results suggest that OVX may not substantially alter body weight or body composition in mice fed a normal diet, but when combined with HFD it increases sleep time and reduces EE, leading to greater weight gain and adiposity without altering food intake.

Neuronal Suppressor of Cytokine Signaling 3: Role in Modulating Chronic Metabolic and Cardiovascular Effects of Leptin.

We determined whether deficiency of neuronal SOCS3 (suppressor of cytokine signaling 3)-a potential negative regulator of leptin signaling-amplifies the chronic effects of leptin on food intake, energy expenditure, glucose, and blood pressure (BP) and protects against adverse cardiometabolic effects of obesity. BP and heart rate were recorded by telemetry, and oxygen consumption (VO2) was monitored in 22-week-old mice with nervous system SOCS3 deficiency (SOCS3-Nestin-Cre) and control mice (SOCS3flox/flox) fed normal or high-fat-high-fructose diet from 6 to 22 weeks of age. Compared with controls, SOCS3-Nestin-Cre mice had lower plasma glucose (124±7 versus 146±10 mg/dL), consumed less food (3.0±0.4 versus 3.6±0.2 g/d), and had similar VO2 (77±6 versus 73±3 mL/kg per minute) and BP (103±3 versus 107±3 mm Hg) but higher heart rate (666±15 versus 602±17 bpm). In mice fed the normal diet, leptin infusion for 7 days caused similar reductions in food intake (2.3±0.1 versus 2.4±0.2 g) but greater increases in BP (15±3 versus 7±2 mm Hg) in SOCS3-Nestin-Cre compared with controls. Leptin reduced blood glucose concentrations in both groups. Male or female SOCS3-Nestin-Cre fed high-fat-high-fructose diet exhibited less weight gain, body fat, and liver steatosis and greater energy expenditure and heart rate compared with controls. Female SOCS3-Nestin-Cre mice fed high-fat-high-fructose diet had higher BP compared with controls. Thus, neuronal SOCS3 seems to play an important role in cardiometabolic regulation because neuronal SOCS3 deficiency reduced body weight and food intake while amplifying leptin's effects on appetite and BP and attenuating the adverse metabolic effects of high-fat-high-fructose diet.

Regulation of Blood Pressure, Appetite, and Glucose by CNS Melanocortin System in Hyperandrogenemic Female SHR.

BACKGROUND: Hyperandrogenemia in females may be associated with sympathetic nervous system (SNS) activation and increased blood pressure (BP). However the importance of hyperandrogenemia in causing hypertension in females and the mechanisms involved are still unclear. We tested whether chronic hyperandrogenemia exacerbates hypertension in young female spontaneously hypertensive rats (SHR) and whether endogenous melanocortin-3/4 receptor (MC3/4R) activation contributes to the elevated BP. METHODS: Cardiovascular and metabolic effects of chronic MC3/4R antagonism were assessed in female SHR treated with dihydrotestosterone (DHT, beginning at 5 weeks of age) and placebo-treated female SHR. BP and heart rate (HR) were measured by telemetry and an intracerebroventricular (ICV) cannula was placed in the lateral ventricle for infusions. After control measurements, the MC3/4R antagonist (SHU-9119) was infused for 10 days (1 nmol/hour, ICV, at 15 weeks of age) followed by a 5-day recovery period. RESULTS: MC3/4R antagonism increased food intake and body weight in DHT-treated SHR (14±1 to 35±1g/day and 244±3 to 298±8g) and controls (14±1 to 34±2g/day and 207±4 to 269±8g). Compared to untreated SHR, DHT-treated SHR had similar BP but lower HR (146±3 vs. 142±4mm Hg and 316±2 vs. 363±4 bpm). Chronic SHU-9119 infusion reduced BP and HR in DHT-treated SHR (-12±2mm Hg and -14±4 bpm) and control female SHR (-19±2mm Hg and -21±6 bpm). CONCLUSION: These results indicate that hyperandrogenemia does not exacerbate hypertension in female SHR. MC3/4R antagonism reduces BP and HR despite marked increases in food intake and body weight in hyperandrogenemic and control female SHR.