Genomic Selection Signals in Andean Highlanders Reveal Adaptive Placental Metabolic Phenotypes That Are Disrupted in Preeclampsia.

BACKGROUND: The chronic hypoxia of high-altitude residence poses challenges for tissue oxygen supply and metabolism. Exposure to high altitude during pregnancy increases the incidence of hypertensive disorders of pregnancy and fetal growth restriction and alters placental metabolism. High-altitude ancestry protects against altitude-associated fetal growth restriction, indicating hypoxia tolerance that is genetic in nature. Yet, not all babies are protected and placental pathologies associated with fetal growth restriction occur in some Andean highlanders. METHODS: We examined placental metabolic function in 79 Andeans (18-45 years; 39 preeclamptic and 40 normotensive) living in La Paz, Bolivia (3600-4100 m) delivered by unlabored Cesarean section. Using a selection-nominated approach, we examined links between putatively adaptive genetic variation and phenotypes related to oxygen delivery or placental metabolism. RESULTS: Mitochondrial oxidative capacity was associated with fetal oxygen delivery in normotensive but not preeclamptic placenta and was also suppressed in term preeclamptic pregnancy. Maternal haplotypes in or within 200 kb of selection-nominated genes were associated with lower placental mitochondrial respiratory capacity (PTPRD [protein tyrosine phosphatase receptor-δ]), lower maternal plasma erythropoietin (CPT2 [carnitine palmitoyl transferase 2], proopiomelanocortin, and DNMT3 [DNA methyltransferase 3]), and lower VEGF (vascular endothelial growth factor) in umbilical venous plasma (TBX5 [T-box transcription factor 5]). A fetal haplotype within 200 kb of CPT2 was associated with increased placental mitochondrial complex II capacity, placental nitrotyrosine, and GLUT4 (glucose transporter type 4) protein expression. CONCLUSIONS: Our findings reveal novel associations between putatively adaptive gene regions and phenotypes linked to oxygen delivery and placental metabolic function in highland Andeans, suggesting that such effects may be of genetic origin. Our findings also demonstrate maladaptive metabolic mechanisms in the context of preeclampsia, including dysregulation of placental oxygen consumption.

Altered placental ion channel gene expression in preeclamptic high-altitude pregnancies.

High-altitude (>2,500 m) residence increases the risk of pregnancy vascular disorders such as fetal growth restriction and preeclampsia, each characterized by impaired placental function. Genetic attributes of highland ancestry confer relative protection against vascular disorders of pregnancy at high altitudes. Although ion channels have been implicated in placental function regulation, neither their expression in high-altitude placentas nor their relationship to high-altitude preeclampsia has been determined. Here, we measured the expression of 26 ion-channel genes in placentas from preeclampsia cases and normotensive controls in La Paz, Bolivia (3,850 m). In addition, we correlated gene transcription to maternal and infant ancestry proportions. Gene expression was assessed by PCR, genetic ancestry evaluated by ADMIXTURE, and ion channel proteins localized by immunofluorescence. In preeclamptic placentas, 11 genes were downregulated (ABCC9, ATP2A2, CACNA1C, KCNE1, KCNJ8, KCNK3, KCNMA1, KCNQ1, KCNQ4, PKD2, and TRPV6) and two were upregulated (KCNQ3 and SCNN1G). KCNE1 expression was positively correlated with high-altitude Amerindian ancestry and negatively correlated with non-high altitude. SCNN1G was negatively correlated with African ancestry, despite minimal African admixture. Most ion channels were localized in syncytiotrophoblasts (Cav1.2, TRPP2, TRPV6, and Kv7.1), whereas expression of Kv7.4 was primarily in microvillous membranes, Kir6.1 in chorionic plate and fetal vessels, and MinK in stromal cells. Our findings suggest a role for differential placental ion channel expression in the development of preeclampsia. Functional studies are needed to determine processes affected by these ion channels in the placenta and whether therapies directed at modulating their activity could influence the onset or severity of preeclampsia.

Neonatal intake of Omega-3 fatty acids enhances lipid oxidation in adipocyte precursors.

Establishing metabolic programming begins during fetal and postnatal development, and early-life lipid exposures play a critical role during neonatal adipogenesis. We define how neonatal consumption of a low omega-6 to -3 fatty acid ratio (n6/n3 FA ratio) establishes FA oxidation in adipocyte precursor cells (APCs) before they become adipocytes. In vivo, APCs isolated from mouse pups exposed to the low n6/n3 FA ratio had superior FA oxidation capacity, elevated beige adipocyte mRNAs Ppargc1α, Ucp2, and Runx1, and increased nuclear receptor NR2F2 protein. In vitro, APC treatment with NR2F2 ligand-induced beige adipocyte mRNAs and increased mitochondrial potential but not mass. Single-cell RNA-sequencing analysis revealed low n6/n3 FA ratio yielded more mitochondrial-high APCs and linked APC NR2F2 levels with beige adipocyte signatures and FA oxidation. Establishing beige adipogenesis is of clinical relevance, because fat depots with energetically active, smaller, and more numerous adipocytes improve metabolism and delay metabolic dysfunction.

High altitude differentially modulates potassium channel-evoked vasodilatation in pregnant human myometrial arteries.

High-altitude (>2500 m or 8200 ft) residence reduces uterine artery blood flow during pregnancy, contributing to an increased incidence of preeclampsia and intrauterine growth restriction. However, not all pregnancies are affected by the chronic hypoxic conditions of high-altitude residence. K+ channels play important roles in the uterine vascular adaptation to pregnancy, promoting a reduction in myogenic tone and an increase in blood flow. We hypothesized that, in pregnancies with normal fetal growth at high altitude, K+ channel-dependent vasodilatation of myometrial arteries is increased compared to those from healthy pregnant women at a lower altitude (∼1700 m). Using pharmacological modulation of two K+ channels, ATP-sensitive (KATP ) and large-conductance Ca2+ -activated (BKCa ) K+ channels, we assessed the vasodilatation of myometrial arteries from appropriate for gestational age (AGA) pregnancies in women living at high or low altitudes. In addition, we evaluated the localization of these channels in the myometrial arteries using immunofluorescence. Our results showed an endothelium-dependent increase in KATP -dependent vasodilatation in myometrial arteries from high versus low altitude, whereas vasodilatation induced by BKCa activation was reduced in these vessels. Additionally, KATP channel co-localization with endothelial markers was reduced in the high-altitude myometrial arteries, which suggested that the functional increase in KATP activity may be by mechanisms other than regulation of channel localization. These observations highlight an important contribution of K+ channels to the human uterine vascular adaptation to pregnancy at high altitude serving to maintain normal fetal growth under conditions of chronic hypoxia. KEY POINTS: High-altitude (>2500 m or 8200 ft) residence reduces uterine blood flow during pregnancy and fetal growth. Animal models of high altitude/chronic hypoxia suggest that these reductions are partially due to reduced vascular K+. channel responses, such as those elicited by large conductance Ca2+ -activated (BKCa ) and ATP-sensitive (KATP ) K+ channel activation. We found that women residing at high versus low altitude during pregnancy showed diminished myometrial artery vasodilatory responses to endothelium-independent BKCa channel activation but greater responses to endothelium-dependent KATP channel activation. Our observations indicate that KATP channels play an adaptive role in maintaining myometrial artery vasodilator sensitivity under chronic hypoxic conditions during pregnancy. Thus, KATP channels represent potential therapeutic targets for augmenting uteroplacental blood flow and, in turn, preserving fetal growth in cases of uteroplacental hypoperfusion.

Vascular Disorders of Pregnancy Increase Susceptibility to Neonatal Pulmonary Hypertension in High-Altitude Populations.

BACKGROUND: Preeclampsia and fetal growth restriction increase cardiopulmonary disease risk for affected offspring and occur more frequently at high-altitude (≥2500 m). Retrospective studies indicate that birth to a preeclampsia woman at high altitude increases the risk of pulmonary hypertension (PH) in later life. This prospective study asked whether preeclampsia with or without fetal growth restriction exaggerated fetal hypoxia and impaired angiogenesis in the fetal lung, leading to neonatal cardiopulmonary circulation abnormalities and neonatal or infantile PH. METHODS AND RESULTS: We studied 79 maternal-infant pairs (39 preeclampsia, 40 controls) in Bolivia (3600-4100 m). Cord blood erythropoietin, hemoglobin, and umbilical artery and venous blood gases were measured as indices of fetal hypoxia. Maternal and cord plasma levels of angiogenic (VEGF [vascular endothelial growth factor]) and antiangiogenic (sFlt1 [soluble fms-like tyrosine kinase]) factors were determined. Postnatal echocardiography (1 week and 6-9 months) assessed pulmonary hemodynamics and PH. Preeclampsia augmented fetal hypoxia and increased the risk of PH in the neonate but not later in infancy. Pulmonary abnormalities were confined to preeclampsia cases with fetal growth restriction. Maternal and fetal plasma sFlt1 levels were higher in preeclampsia than controls and positively associated with PH. CONCLUSIONS: The effect of preeclampsia with fetal growth restriction to increase fetal hypoxia and sFlt1 levels may impede normal development of the pulmonary circulation at high altitude, leading to adverse neonatal pulmonary vascular outcomes. Our observations highlight important temporal windows for the prevention of pulmonary vascular disease among babies born to highland residents or those with exaggerated hypoxia in utero or newborn life.

Breast Cancer Endocrine Therapy Promotes Weight Gain With Distinct Adipose Tissue Effects in Lean and Obese Female Mice.

Breast cancer survivors treated with tamoxifen and aromatase inhibitors report weight gain and have an elevated risk of type 2 diabetes, especially if they have obesity. These patient experiences are inconsistent with, preclinical studies using high doses of tamoxifen which reported acute weight loss. We investigated the impact of breast cancer endocrine therapies in a preclinical model of obesity and in a small group of breast adipose tissue samples from women taking tamoxifen to understand the clinical findings. Mature female mice were housed at thermoneutrality and fed either a low-fat/low-sucrose (LFLS) or a high-fat/high-sucrose (HFHS) diet. Consistent with the high expression of Esr1 observed in mesenchymal stem cells from adipose tissue, endocrine therapy was associated with adipose accumulation and more preadipocytes compared with estrogen-treated control mice but resulted in fewer adipocyte progenitors only in the context of HFHS. Analysis of subcutaneous adipose stromal cells revealed diet- and treatment-dependent effects of endocrine therapies on various cell types and genes, illustrating the complexity of adipose tissue estrogen receptor signaling. Breast cancer therapies supported adipocyte hypertrophy and associated with hepatic steatosis, hyperinsulinemia, and glucose intolerance, particularly in obese females. Current tamoxifen use associated with larger breast adipocyte diameter only in women with obesity. Our translational studies suggest that endocrine therapies may disrupt adipocyte progenitors and support adipocyte hypertrophy, potentially leading to ectopic lipid deposition that may be linked to a greater type 2 diabetes risk. Monitoring glucose tolerance and potential interventions that target insulin action should be considered for some women receiving life-saving endocrine therapies for breast cancer.

Lipoprotein Lipase Overexpression in Skeletal Muscle Attenuates Weight Regain by Potentiating Energy Expenditure.

Moderate weight loss improves numerous risk factors for cardiometabolic disease; however, long-term weight loss maintenance (WLM) is often thwarted by metabolic adaptations that suppress energy expenditure and facilitate weight regain. Skeletal muscle has a prominent role in energy homeostasis; therefore, we investigated the effect of WLM and weight regain on skeletal muscle in rodents. In skeletal muscle of obesity-prone rats, WLM reduced fat oxidative capacity and downregulated genes involved in fat metabolism. Interestingly, even after weight was regained, genes involved in fat metabolism were also reduced. We then subjected mice with skeletal muscle lipoprotein lipase overexpression (mCK-hLPL), which augments fat metabolism, to WLM and weight regain and found that mCK-hLPL attenuates weight regain by potentiating energy expenditure. Irrespective of genotype, weight regain suppressed dietary fat oxidation and downregulated genes involved in fat metabolism in skeletal muscle. However, mCK-hLPL mice oxidized more fat throughout weight regain and had greater expression of genes involved in fat metabolism and lower expression of genes involved in carbohydrate metabolism during WLM and regain. In summary, these results suggest that skeletal muscle fat oxidation is reduced during WLM and regain, and therapies that improve skeletal muscle fat metabolism may attenuate rapid weight regain.

High altitude regulates the expression of AMPK pathways in human placenta.

INTRODUCTION: High-altitude (>2500 m) residence augments the risk of intrauterine growth restriction (IUGR) and preeclampsia likely due, in part, to uteroplacental hypoperfusion. Previous genomic and transcriptomic studies in humans and functional studies in mice and humans suggest a role for AMP-activated protein kinase (AMPK) pathway in protecting against hypoxia-associated IUGR. AMPK is a metabolic sensor activated by hypoxia that is ubiquitously expressed in vascular beds and placenta. METHODS: We measured gene expression and protein levels of AMPK and its upstream regulators and downstream targets in human placentas from high (>2500 m) vs. moderate (~1700 m) and low (~100 m) altitude. RESULTS: We found that phosphorylated AMPK protein levels and its downstream target TSC2 were increased in placentas from high and moderate vs. low altitude, whereas the phosphorylated form of the downstream target translation repressor protein 4E-BP1 was increased in high compared to moderate as well as low altitude placentas. Mean birth weights progressively fell with increasing altitude but no infants, by study design, were clinically growth-restricted. Gene expression analysis showed moderate increases in PRKAG2, encoding the AMPK γ2 subunit, and mechanistic target of rapamycin, MTOR, expression. DISCUSSION: These results highlight a differential regulation of placental AMPK pathway activation in women residing at low, moderate or high altitude during pregnancy, suggesting AMPK may be serving as a metabolic regulator for integrating hypoxic stimuli with placental function.

Hypoxia-induced inhibition of mTORC1 activity in the developing lung: a possible mechanism for the developmental programming of pulmonary hypertension.

Perinatal hypoxia induces permanent structural and functional changes in the lung and its pulmonary circulation that are associated with the development of pulmonary hypertension (PH) in later life. The mechanistic target of the rapamycin (mTOR) pathway is vital for fetal lung development and is implicated in hypoxia-associated PH, yet its involvement in the developmental programming of PH remains unclear. Pregnant C57/BL6 dams were placed in hyperbaric (760 mmHg) or hypobaric chambers during gestation (505 mmHg, day 15 through postnatal day 4) or from weaning through adulthood (420 mmHg, postnatal day 21 through 8 wk). Pulmonary hemodynamics and right ventricular systolic pressure (RVSP) were measured at 8 wk. mTOR pathway proteins were assessed in fetal (day 18.5) and adult lung (8 wk). Perinatal hypoxia induced PH during adulthood, even in the absence of a sustained secondary hypoxic exposure, as indicated by reduced pulmonary artery acceleration time (PAAT) and peak flow velocity through the pulmonary valve, as well as greater RVSP, right ventricular (RV) wall thickness, and RV/left ventricular (LV) weight. Such effects were independent of increased blood viscosity. In fetal lung homogenates, hypoxia reduced the expression of critical downstream mTOR targets, most prominently total and phosphorylated translation repressor protein (4EBP1), as well as vascular endothelial growth factor, a central regulator of angiogenesis in the fetal lung. In contrast, adult offspring of hypoxic dams tended to have elevated p4EBP1 compared with controls. Our data suggest that inhibition of mTORC1 activity in the fetal lung as a result of gestational hypoxia may interrupt pulmonary vascular development and thereby contribute to the developmental programming of PH.NEW & NOTEWORTHY We describe the first study to evaluate a role for the mTOR pathway in the developmental programming of pulmonary hypertension. Our findings suggest that gestational hypoxia impairs mTORC1 activation in the fetal lung and may impede pulmonary vascular development, setting the stage for pulmonary vascular disease in later life.

AMP-activated protein kinase activator AICAR attenuates hypoxia-induced murine fetal growth restriction in part by improving uterine artery blood flow.

KEY POINTS: Pregnancy at high altitude is associated with a greater incidence of fetal growth restriction due, in part, to lesser uterine artery blood flow. AMP-activated protein kinase (AMPK) activation vasodilates arteries and may increase uterine artery blood flow. In this study, pharmacological activation of AMPK by the drug AICAR improved fetal growth and elevated uterine artery blood flow. These results suggest that AMPK activation is a potential strategy for improving fetal growth and raising uterine artery blood flow in pregnancy, which may be important in pregnancy disorders characterized by uteroplacental ischaemia and/or fetal hypoxia. ABSTRACT: Uteroplacental hypoxia is associated with pregnancy disorders such as intrauterine growth restriction and preeclampsia, which are characterized by uteroplacental ischaemia and/or fetal hypoxia. Activation of AMP-activated protein kinase (AMPK) results in vasodilatation and is therefore a potential therapeutic strategy for restoring uteroplacental perfusion in pregnancy disorders. In this study, C57Bl/6 mice were treated with subcutaneous pellets containing vehicle, the AMPK activator AICAR (200 mg kg-1 day-1 ), or the AMPK inhibitor Compound C (20 mg kg-1 day-1 ) beginning on gestational day 13.5, and were exposed to hypoxia starting on gestational day 14.5 that induced intrauterine growth restriction. Pharmacological AMPK activation by AICAR partially prevented hypoxia-induced fetal growth restriction (P < 0.01), due in part to increased uterine artery blood flow (P < 0.0001). The proportion of total cardiac output flowing through the uterine artery was increased with AICAR in hypoxic mice (P < 0.001), suggesting that the vasodilator effect of AICAR was selective for the uterine circulation. Further, pharmacological inhibition of AMPK with Compound C reduced uterine artery diameter and increased uterine artery contractility in normoxic mice, providing evidence that physiological levels of AMPK activation are necessary for vasodilatation in healthy pregnancy. Two-way ANOVA analyses indicated that hypoxia reduced AMPK activation in the uterine artery and placenta, and AICAR increased AMPK activation in these tissues compared to vehicle. These findings provide support for further investigation into the utility of pharmacological AMPK activation for treatment of fetal growth restriction.

AMPK activation in pregnant human myometrial arteries from high-altitude and intrauterine growth-restricted pregnancies.

High-altitude (>2,500 m) residence increases the incidence of intrauterine growth restriction (IUGR) due, in part, to reduced uterine artery blood flow and impaired myometrial artery (MA) vasodilator response. A role for the AMP-activated protein kinase (AMPK) pathway in protecting against hypoxia-associated IUGR is suggested by genomic and transcriptomic studies in humans and functional studies in mice. AMPK is a hypoxia-sensitive metabolic sensor with vasodilatory properties. Here we hypothesized that AMPK-dependent vasodilation was increased in MAs from high versus low-altitude (<1,700 m) Colorado women with appropriate for gestational age (AGA) pregnancies and reduced in IUGR pregnancies regardless of altitude. Vasoreactivity studies showed that, in AGA pregnancies, MAs from high-altitude women were more sensitive to vasodilation by activation of AMPK with A769662 due chiefly to increased endothelial nitric oxide production, whereas MA responses to AMPK activation in the low-altitude women were endothelium independent. MAs from IUGR compared with AGA pregnancies had blunted vasodilator responses to acetylcholine at high altitude. We concluded that 1) blunted vasodilator responses in IUGR pregnancies confirm the importance of MA vasodilation for normal fetal growth and 2) the increased sensitivity to AMPK activation in AGA pregnancies at high altitude suggests that AMPK activation helped maintain MA vasodilation and fetal growth. These results highlight a novel mechanism for vasodilation of MAs under conditions of chronic hypoxia and suggest that AMPK activation could provide a therapy for increasing uteroplacental blood flow and improving fetal growth in IUGR pregnancies.NEW & NOTEWORTHY Intrauterine growth restriction (IUGR) impairs infant well- being and increases susceptibility to later-in-life diseases for mother and child. Our study reveals a novel role for AMPK in vasodilating the myometrial artery (MA) from women residing at high altitude (>2,500 m) with appropriate for gestational age pregnancies but not in IUGR pregnancies at any altitude.

Kynurenic Acid Protects Against Ischemia/Reperfusion-Induced Retinal Ganglion Cell Death in Mice.

BACKGROUND: Glaucoma is an optic neuropathy and involves the progressive degeneration of retinal ganglion cells (RGCs), which leads to blindness in patients. We investigated the role of the neuroprotective kynurenic acid (KYNA) in RGC death against retinal ischemia/reperfusion (I/R) injury. METHODS: We injected KYNA intravenously or intravitreally to mice. We generated a knockout mouse strain of kynurenine 3-monooxygenase (KMO), an enzyme in the kynurenine pathway that produces neurotoxic 3-hydroxykynurenine. To test the effect of mild hyperglycemia on RGC protection, we used streptozotocin (STZ) induced diabetic mice. Retinal I/R injury was induced by increasing intraocular pressure for 60 min followed by reperfusion and RGC numbers were counted in the retinal flat mounts. RESULTS: Intravenous or intravitreal administration of KYNA protected RGCs against I/R injury. The I/R injury caused a greater loss of RGCs in wild type than in KMO knockout mice. KMO knockout mice had mildly higher levels of fasting blood glucose than wild type mice. Diabetic mice showed significantly lower loss of RGCs when compared with non-diabetic mice subjected to I/R injury. CONCLUSION: Together, our study suggests that the absence of KMO protects RGCs against I/R injury, through mechanisms that likely involve higher levels of KYNA and glucose.

Peroxisome proliferator-activated receptor gamma blunts endothelin-1-mediated contraction of the uterine artery in a murine model of high-altitude pregnancy.

The environmental hypoxia of high altitude (HA) increases the incidence of intrauterine growth restriction (IUGR) approximately threefold. The peroxisome proliferator-activated receptor γ (PPAR-γ), a ligand-activated nuclear receptor that promotes vasorelaxation by increasing nitric oxide and downregulating endothelin-1 (ET-1) production, has been implicated in IUGR. Based on our prior work indicating that pharmacologic activation of the PPARγ pathway protects against hypoxia-associated IUGR, we used an experimental murine model to determine whether such effects may be attributed to vasodilatory effects in the uteroplacental circulation. Using wire myography, ex vivo vasoreactivity studies were conducted in uterine arteries (UtA) isolated from pregnant mice exposed to hypoxia or normoxia from gestational day 14.5 to 18.5. Exposure to troglitazone, a high-affinity PPARγ agonist-induced vasorelaxation in UtA preconstricted with phenylephrine, with HA-UtA showing increased sensitivity. Troglitazone blunted ET-1-induced contraction of UtA in hypoxic and normoxic dams equivalently. Immunohistological analysis revealed enhanced staining for ET-1 receptors in the placental labyrinthine zone in hypoxic compared to normoxic dams. Our results suggest that pharmacologic PPAR-γ activation, via its vasoactive properties, may protect the fetal growth under hypoxic conditions by improving uteroplacental perfusion and thereby justify further investigation into PPARγ as a therapeutic target for IUGR in pregnancies complicated by hypoxia.

Regular exercise potentiates energetically expensive hepatic de novo lipogenesis during early weight regain.

Exercise is a potent facilitator of long-term weight loss maintenance (WLM), whereby it decreases appetite and increases energy expenditure beyond the cost of the exercise bout. We have previously shown that exercise may amplify energy expenditure through energetically expensive nutrient deposition. Therefore, we investigated the effect of exercise on hepatic de novo lipogenesis (DNL) during WLM and relapse to obesity. Obese rats were calorically restricted with (EX) or without (SED) treadmill exercise (1 h/day, 6 days/wk, 15 m/min) to induce and maintain weight loss. After 6 wk of WLM, subsets of WLM-SED and WLM-EX rats were allowed ad libitum access to food for 1 day to promote relapse (REL). An energy gap-matched group of sedentary, relapsing rats (REL-GM) were provided a diet matched to the positive energy imbalance of the REL-EX rats. During relapse, exercise increased enrichment of hepatic DN-derived lipids and induced hepatic molecular adaptations favoring DNL compared with the gap-matched controls. In the liver, compared with both REL-SED and REL-GM rats, REL-EX rats had lower hepatic expression of genes required for cholesterol biosynthesis; greater hepatic expression of genes that mediate very low-density lipoprotein synthesis and secretion; and greater mRNA expression of Cyp27a1, which encodes an enzyme involved in the biosynthesis of bile acids. Altogether, these data provide compelling evidence that the liver has an active role in exercise-mediated potentiation of energy expenditure during early relapse.

Impact of Exercise and Activity on Weight Regain and Musculoskeletal Health Post-Ovariectomy.

The purpose of this study was to determine whether obesity and/or exercise training alters weight regain and musculoskeletal health after ovariectomy (OVX). Female rats were fed high-fat diet (HFD) to reveal obesity-prone (OP) and obesity-resistant (OR) phenotypes. The OP and OR exercising (EX) and sedentary (SED) rats were calorically restricted to lose 15% of body weight using medium-fat diet. Rats were then maintained in energy balance for 8 wk before OVX. After OVX and a brief calorically limited phase, rats were allowed to eat ad libitum until body weight plateaued. Starting at weight loss, EX ran 1 h·d, 6 d·wk, 15 m·min. Energy intake, spontaneous physical activity (SPA), and total energy expenditure were evaluated at the end of weight maintenance pre-OVX, and at three time points post-OVX: before weight regain, during early regain, and after regain. Data are presented as mean ± SE. Exercise attenuated weight regain after OVX in OP only (OP-EX, 123 ± 10 g; OP-SED, 165 ± 12 g; OR-EX, 121 ± 6 g; OR-SED, 116 ± 6 g), which was primarily an attenuation of fat gain. The early post-OVX increase in energy intake explained much of the weight regain, and was similar across groups. Exercising improved bone strength, as did maintaining SPA. Group differences in muscle mitochondrial respiration were not significant. The large decrease in SPA due to OVX was persistent, but early weight regain was dependent on decreased SPA. In conclusion, leanness and exercise do not necessarily protect from OVX-induced weight gain. Exercise prevented weight gain in obese rats, but loss of SPA was the greatest contributor to post-OVX weight gain. Thus, understanding the mechanisms resulting in reduction in SPA after ovarian hormone loss is critical in the prevention of menopause-associated metabolic dysfunction.

Compensation for cold-induced thermogenesis during weight loss maintenance and regain.

Prevalence of obesity is exacerbated by low rates of successful long-term weight loss maintenance (WLM). In part, relapse from WLM to obesity is due to a reduction in energy expenditure (EE) that persists throughout WLM and relapse. Thus, interventions that increase EE might facilitate WLM. In obese mice that were calorically restricted to reduce body weight by ~20%, we manipulated EE throughout WLM and early relapse using intermittent cold exposure (ICE; 4°C, 90 min/day, 5 days/wk, within the last 3 h of the light cycle). EE, energy intake, and spontaneous physical activity were measured during the obese, WLM, and relapse phases. During WLM and relapse, the ICE group expended more energy during the light cycle because of cold exposure but expended less energy in the dark cycle, which led to no overall difference in total daily EE. The compensation in EE appeared to be mediated by activity, whereby the ICE group was more active during the light cycle because of cold exposure but less active during the dark cycle, which led to no overall effect on total daily activity during WLM and relapse. In brown adipose tissue of relapsing mice, the ICE group had greater mRNA expression of Dio2 and protein expression of UCP1 but lower mRNA expression of Prdm16. In summary, these findings indicate that despite robust increases in EE during cold exposures, ICE is unable to alter total daily EE during WLM or early relapse, likely due to compensatory behaviors in activity.

Low Neonatal Plasma n-6/n-3 PUFA Ratios Regulate Offspring Adipogenic Potential and Condition Adult Obesity Resistance.

Adipose tissue expansion progresses rapidly during postnatal life, influenced by both prenatal maternal factors and postnatal developmental cues. The ratio of omega-6 (n-6) relative to n-3 polyunsaturated fatty acids (PUFAs) is believed to regulate perinatal adipogenesis, but the cellular mechanisms and long-term effects are not well understood. We lowered the fetal and postnatal n-6/n-3 PUFA ratio exposure in wild-type offspring under standard maternal dietary fat amounts to test the effects of low n-6/n-3 ratios on offspring adipogenesis and adipogenic potential. Relative to wild-type pups receiving high perinatal n-6/n-3 ratios, subcutaneous adipose tissue in 14-day-old wild-type pups receiving low n-6/n-3 ratios had more adipocytes that were smaller in size; decreased Pparγ2, Fabp4, and Plin1; several lipid metabolism mRNAs; coincident hypermethylation of the PPARγ2 proximal promoter; and elevated circulating adiponectin. As adults, offspring that received low perinatal n-6/n-3 ratios were diet-induced obesity (DIO) resistant and had a lower positive energy balance and energy intake, greater lipid fuel preference and non-resting energy expenditure, one-half the body fat, and better glucose clearance. Together, the findings support a model in which low early-life n-6/n-3 ratios remodel adipose morphology to increase circulating adiponectin, resulting in a persistent adult phenotype with improved metabolic flexibility that prevents DIO.

Prior weight loss exacerbates the biological drive to gain weight after the loss of ovarian function.

Both the history of obesity and weight loss may change how menopause affects metabolic health. The purpose was to determine whether obesity and/or weight loss status alters energy balance (EB) and subsequent weight gain after the loss of ovarian function. Female lean and obese Wistar rats were randomized to 15% weight loss (WL) or ad libitum fed controls (CON). After the weight loss period, WL rats were kept in EB at the reduced weight for 8 weeks prior to ovariectomy (OVX). After OVX, all rats were allowed to eat ad libitum until weight plateaued. Energy intake (EI), spontaneous physical activity, and total energy expenditure (TEE) were measured with indirect calorimetry before OVX, immediately after OVX, and after weight plateau. Changes in energy intake (EI), TEE, and weight gain immediately after OVX were similar between lean and obese rats. However, obese rats gained more total weight and fat mass than lean rats over the full regain period. Post-OVX, EI increased more (P ≤ 0.03) in WL rats (58.9 ± 3.5 kcal/d) than CON rats (8.5 ± 5.2 kcal/d), and EI partially normalized (change from preOVX: 20.5 ± 4.2 vs. 1.5 ± 4.9 kcal/day) by the end of the study. As a result, WL rats gained weight (week 1:44 ± 20 vs. 7 ± 25 g) more rapidly (mean = 44 ± 20 vs. 7 ± 25 g/week; P < 0.001) than CON Prior obesity did not affect changes in EB or weight regain following OVX, whereas a history of weight loss prior to OVX augmented disruptions in EB after OVX, resulting in more rapid weight regain.

Metformin Accumulation Correlates with Organic Cation Transporter 2 Protein Expression and Predicts Mammary Tumor Regression In Vivo.

Several epidemiologic studies have associated metformin treatment with a reduction in breast cancer incidence in prediabetic and type II diabetic populations. Uncertainty exists regarding which patient populations and/or tumor subtypes will benefit from metformin treatment, and most preclinical in vivo studies have given little attention to the cellular pharmacology of intratumoral metformin uptake. Epidemiologic reports consistently link western-style high fat diets (HFD), which drive overweight and obesity, with increased risk of breast cancer. We used a rat model of HFD-induced overweight and mammary carcinogenesis to define intratumoral factors that confer metformin sensitivity. Mammary tumors were initiated with 1-methyl-1-nitrosourea, and rats were randomized into metformin-treated (2 mg/mL drinking water) or control groups (water only) for 8 weeks. Two-thirds of existing mammary tumors responded to metformin treatment with decreased tumor volumes (P < 0.05), reduced proliferative index (P < 0.01), and activated AMPK (P < 0.05). Highly responsive tumors accumulated 3-fold greater metformin amounts (P < 0.05) that were positively correlated with organic cation transporter-2 (OCT2) protein expression (r = 0.57; P = 0.038). Importantly, intratumoral metformin concentration negatively associated with tumor volume (P = 0.03), and each 10 pmol increase in intratumoral metformin predicted >0.11 cm3 reduction in tumor volume. Metformin treatment also decreased proinflammatory arachidonic acid >1.5-fold in responsive tumors (P = 0.023). Collectively, these preclinical data provide evidence for a direct effect of metformin in vivo and suggest that OCT2 expression may predict metformin uptake and tumor response. Cancer Prev Res; 10(3); 198-207. ©2017 AACR.

Attenuated Pik3r1 expression prevents insulin resistance and adipose tissue macrophage accumulation in diet-induced obese mice.

Obese white adipose tissue (AT) is characterized by large-scale infiltration of proinflammatory macrophages, in parallel with systemic insulin resistance; however, the cellular stimulus that initiates this signaling cascade and chemokine release is still unknown. The objective of this study was to determine the role of the phosphoinositide 3-kinase (PI3K) regulatory subunits on AT macrophage (ATM) infiltration in obesity. Here, we find that the Pik3r1 regulatory subunits (i.e., p85α/p55α/p50α) are highly induced in AT from high-fat diet-fed obese mice, concurrent with insulin resistance. Global heterozygous deletion of the Pik3r1 regulatory subunits (αHZ), but not knockout of Pik3r2 (p85ß), preserves whole-body, AT, and skeletal muscle insulin sensitivity, despite severe obesity. Moreover, ATM accumulation, proinflammatory gene expression, and ex vivo chemokine secretion in obese αHZ mice are markedly reduced despite endoplasmic reticulum (ER) stress, hypoxia, adipocyte hypertrophy, and Jun NH(2)-terminal kinase activation. Furthermore, bone marrow transplant studies reveal that these improvements in obese αHZ mice are independent of reduced Pik3r1 expression in the hematopoietic compartment. Taken together, these studies demonstrate that Pik3r1 expression plays a critical role in mediating AT insulin sensitivity and, more so, suggest that reduced PI3K activity is a key step in the initiation and propagation of the inflammatory response in obese AT.