RESUMEN
Perturbed maternal diet and prenatal exposure to air pollution (AP) affect the fetal brain, predisposing to postnatal neurobehavioral disorders. Glucose transporters (GLUTs) are key in fueling neurotransmission; deficiency of the neuronal isoform GLUT3 culminates in autism spectrum disorders. Along with the different neurotransmitters, serotonin (5-HT) and oxytocin (OXT) are critical for the development of neural connectivity. Serotonin transporter (SERT) modulates synaptic 5-HT levels, while the OXT receptor (OXTR) mediates OXT action. We hypothesized that perturbed brain GLUT1/GLUT3 regulated 5-HT-SERT imbalance, which serves as a contributing factor to postnatal neuropsychiatric phenotypes, with OXT/OXTR providing a counterbalance. Employing maternal diet restriction (intrauterine growth restriction [IUGR]), high-fat (HF) dietary modifications, and prenatal exposure to simulated AP, fetal (E19) murine brain 5-HT was assessed by ELISA with SERT and OXTR being localized by immunohistochemistry and measured by quantitative Western blot analysis. IUGR with lower head weights led to a 48% reduction in male and female fetal brain GLUT3 with no change in GLUT1, when compared to age- and sex-matched controls, with no significant change in OXTR. In addition, a â¼50% (p = 0.005) decrease in 5-HT and SERT concentrations was displayed in fetal IUGR brains. In contrast, despite emergence of microcephaly, exposure to a maternal HF diet or AP caused no significant changes. We conclude that in the IUGR during fetal brain development, reduced GLUT3 is associated with an imbalanced 5-HT-SERT axis. We speculate that these early changes may set the stage for altering the 5HT-SERT neural axis with postnatal emergence of associated neurodevelopmental disorders.
Asunto(s)
Contaminantes Atmosféricos , Transportador de Glucosa de Tipo 3 , Receptores de Oxitocina , Proteínas de Transporte de Serotonina en la Membrana Plasmática , Serotonina , Animales , Encéfalo , Dieta , Femenino , Proteínas Facilitadoras del Transporte de la Glucosa , Transportador de Glucosa de Tipo 1 , Masculino , Ratones , Oxitocina , EmbarazoRESUMEN
To understand the cellular basis for the neurodevelopmental effects of intrauterine growth restriction (IUGR), we examined the global and regional expression of various cell types within murine (Mus musculus) fetal brain. Our model employed maternal calorie restriction to 50% daily food intake from gestation day 10-19, producing IUGR offspring. Offspring had smaller head sizes with larger head:body ratios indicating a head sparing IUGR effect. IUGR fetuses at embryonic day 19 (E19) had reduced nestin (progenitors), ß-III tubulin (immature neurons), Glial fibrillary acidic protein (astrocytes), and O4 (oligodendrocytes) cell lineages via immunofluorescence quantification and a 30% reduction in cortical thickness. No difference was found in Bcl-2 or Bax (apoptosis) between controls and IUGR, though qualitatively, immunoreactivity of doublecortin (migration) and Ki67 (proliferation) was decreased. In the interest of examining a potential therapeutic peptide, we next investigated a novel pro-survival peptide, mouse Humanin (mHN). Ontogeny examination revealed highest mHN expression at E19, diminishing by postnatal day 15 (P15), and nearly absent in adult (3 months). Subanalysis by sex at E19 yielded higher mHN expression among males during fetal life, without significant difference between sexes postnatally. Furthermore, female IUGR mice at E19 had a greater increase in cortical mHN versus the male fetus over their respective controls. We conclude that maternal dietary restriction-associated IUGR interferes with neural progenitors differentiating into the various cellular components populating the cerebral cortex, and reduces cerebral cortical size. mHN expression is developmental stage and sex specific, with IUGR, particularly in the females, adaptively increasing its expression toward mediating a pro-survival approach against nutritional adversity.
Asunto(s)
Corteza Cerebral/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Fenómenos Fisiologicos Nutricionales Maternos/fisiología , Neuronas/metabolismo , Animales , Restricción Calórica , Femenino , Retardo del Crecimiento Fetal/metabolismo , Proteína Ácida Fibrilar de la Glía/metabolismo , Masculino , RatonesRESUMEN
We created a neural-specific conditional murine glut3 (Slc2A3) deletion (glut3flox/flox/nestin-Cre+) to examine the effect of a lack of Glut3 on neurodevelopment. Compared with age-matched glut3flox/flox = WT and heterozygotes (glut3flox/+/nestin-Cre+), we found that a >90% reduction in male and female brain Glut3 occurred by postnatal day 15 (PN15) in glut3flox/flox/nestin-Cre+ This genetic manipulation caused a diminution in brain weight and cortical thickness at PN15, a reduced number of dendritic spines, and fewer ultrasonic vocalizations. Patch-clamp recordings of cortical pyramidal neurons revealed increased frequency of bicuculline-induced paroxysmal discharges as well as reduced latency, attesting to a functional synaptic and cortical hyperexcitability. Concomitant stunting with lower glucose concentrations despite increased milk intake shortened the lifespan, failing rescue by a ketogenic diet. This led to creating glut3flox/flox/CaMK2α-Cre+ mice lacking Glut3 in the adult male limbic system. These mice had normal lifespan, displayed reduced IPSCs in cortical pyramidal neurons, less anxiety/fear, and lowered spatial memory and motor abilities but heightened exploratory and social responses. These distinct postnatal and adult phenotypes, based upon whether glut3 gene is globally or restrictively absent, have implications for humans who carry copy number variations and present with neurodevelopmental disorders.SIGNIFICANCE STATEMENT Lack of the key brain-specific glucose transporter 3 gene found in neurons during early postnatal life results in significant stunting, a reduction in dendritic spines found on neuronal processes and brain size, heightened neuronal excitability, along with a shortened lifespan. When occurring in the adult and limited to the limbic system alone, lack of this gene in neurons reduces the fear of spatial exploration and socialization but does not affect the lifespan. These features are distinct heralding differences between postnatal and adult phenotypes based upon whether the same gene is globally or restrictively lacking. These findings have implications for humans who carry copy number variations pertinent to this gene and have been described to present with neurodevelopmental disorders.
Asunto(s)
Encéfalo/metabolismo , Conducta Exploratoria/fisiología , Eliminación de Gen , Transportador de Glucosa de Tipo 3/deficiencia , Transportador de Glucosa de Tipo 3/genética , Fenotipo , Factores de Edad , Animales , Animales Recién Nacidos , Encéfalo/patología , Espinas Dendríticas/genética , Espinas Dendríticas/metabolismo , Espinas Dendríticas/patología , Femenino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Embarazo , Isoformas de Proteínas/deficiencia , Isoformas de Proteínas/genéticaRESUMEN
OBJECTIVE: We examined Red Blood Cell (RBC) Glucose Transporter isoform 1 (GLUT1) and White Blood Cell (WBC) Glucose Transporter isoform 3 (GLUT3) protein concentrations to assess their potential as surrogate biomarkers for the presence of hypoxic-ischemic encephalopathy (HIE) and response to therapeutic hypothermia (TH), with respect to the neurodevelopmental prognosis. STUDY DESIGN: A prospective feasibility study of 10 infants with HIE and 8 age-matched control subjects was undertaken. Following parental consent, blood samples were obtained at baseline before institution of TH (<6â¯h of life), during TH, at rewarming and post-TH in the HIE group with a baseline sample from the control group. GLUT1 and GLUT3 were measured by Enzyme-linked immunosorbent assay (ELISA) with brain biomarkers, Neuron-Specific Enolase (NSE) and Glial Fibrillary Acidic Protein (GFAP). Novel "HIE-high risk" and "Neurological" scores were developed to help identify HIE and to assess severity and prognosis, respectively. RESULTS: RBC GLUT1 concentrations were increased at the baseline pre-TH time point in HIE versus control subjects (pâ¯=â¯.006), normalizing after TH (pâ¯=â¯.05). An association between GLUT1 and NSE concentrations (which was reflective of the HIE-high risk and the Neuro-scores) in controls and HIE pre-TH was seen (R2â¯=â¯0.36, pâ¯=â¯.008), with GLUT1 demonstrating 90% sensitivity and 88% specificity for presence of HIE identified by Sarnat Staging. WBC GLUT3 concentrations were low and no different in HIE versus control, and GFAP concentrations trended higher during re-warming (pâ¯=â¯.11) and post-TH (pâ¯=â¯.16). We demonstrated a significant difference between HIE and controls for both the "HIE-high risk" and the "Neurological" Scores. The latter score revealing the severity of clinical neurological illness correlated with the corresponding RBC GLUT1 (R2 valueâ¯=â¯0.39; pâ¯=â¯.006). CONCLUSION: Circulating RBC GLUT1 concentrations with NSE demonstrate a significant potential in reflecting the severity of HIE pre-TH and gauging effectiveness of TH. In contrast, the low neonatal WBC GLUT3 concentrations make discerning differences between degrees of HIE as well as assessing effectiveness of TH difficult. The HIE-high risk and Neurological scores may extend the "Sarnat staging" towards assessing severity and neuro-developmental prognosis of HIE.
Asunto(s)
Transportador de Glucosa de Tipo 1/metabolismo , Transportador de Glucosa de Tipo 3/metabolismo , Hipoxia-Isquemia Encefálica/diagnóstico , Biomarcadores , Eritrocitos/metabolismo , Estudios de Factibilidad , Femenino , Proteína Ácida Fibrilar de la Glía/metabolismo , Humanos , Recién Nacido , Unidades de Cuidado Intensivo Neonatal , Masculino , Fosfopiruvato Hidratasa/metabolismo , Proyectos Piloto , Pronóstico , Estudios Prospectivos , Sensibilidad y Especificidad , Índice de Severidad de la EnfermedadRESUMEN
Intrauterine growth restriction leads to the development of adult onset obesity/metabolic syndrome, diabetes mellitus, cardiovascular disease, hypertension, stroke, dyslipidemia, and non-alcoholic fatty liver disease/steatohepatitis. Continued postnatal growth restriction has been shown to ameliorate many of these sequelae. To further our understanding of the mechanism of how intrauterine and early postnatal growth affects adult health we have employed Affymetrix microarray-based expression profiling to characterize hepatic gene expression of male offspring in a rat model of maternal nutrient restriction in early and late life. At day 21 of life (p21) combined intrauterine and postnatal calorie restriction treatment led to expression changes in circadian, metabolic, and insulin-like growth factor genes as part of a larger transcriptional response that encompasses 144 genes. Independent and controlled experiments at p21 confirm the early life circadian, metabolic, and growth factor perturbations. In contrast to the p21 transcriptional response, at day 450 of life (d450) only seven genes, largely uncharacterized, were differentially expressed. This lack of a transcriptional response identifies non-transcriptional mechanisms mediating the adult sequelae of intrauterine growth restriction. Independent experiments at d450 identify a circadian defect as well as validate expression changes to four of the genes identified by the microarray screen which have a novel association with growth restriction. Emerging from this rich dataset is a portrait of how the liver responds to growth restriction through circadian dysregulation, energy/substrate management, and growth factor modulation.
Asunto(s)
Restricción Calórica/efectos adversos , Retardo del Crecimiento Fetal/genética , Perfilación de la Expresión Génica/métodos , Hígado/crecimiento & desarrollo , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Animales , Animales Recién Nacidos/crecimiento & desarrollo , Peso Corporal , Ritmo Circadiano , Femenino , Retardo del Crecimiento Fetal/etiología , Regulación del Desarrollo de la Expresión Génica , Masculino , Embarazo , Efectos Tardíos de la Exposición Prenatal/genética , Ratas , Ratas Sprague-DawleyRESUMEN
Pre- and postnatal calorie restriction is associated with postnatal growth restriction, reduced circulating leptin concentrations, and perturbed energy balance. Hypothalamic regulation of energy balance demonstrates enhanced orexigenic (NPY, AgRP) and diminished anorexigenic (POMC, CART) neuropeptide expression (PN21), setting the stage for subsequent development of obesity in female Sprague-Dawley rats. Leptin replenishment during the early postnatal period (PN2-PN8) led to reversal of the hypothalamic orexigenic:anorexigenic neuropeptide ratio at PN21 by reducing only the orexigenic (NPY, AgRP), without affecting the anorexigenic (POMC, CART) neuropeptide expression. This hypothalamic effect was mediated via enhanced leptin receptor (ObRb) signaling that involved increased pSTAT3/STAT3 but reduced PTP1B. This was further confirmed by an increase in body weight at PN21 in response to intracerebroventricular administration of antisense ObRb oligonucleotides (PN2-PN8). The change in the hypothalamic neuropeptide balance in response to leptin administration was associated with increased oxygen consumption, carbon dioxide production, and physical activity, which resulted in increased milk intake (PN14) with no change in body weight. This is in contrast to the reduction in milk intake with no effect on energy expenditure and physical activity observed in controls. We conclude that pre- and postnatal calorie restriction perturbs hypothalamic neuropeptide regulation of energy balance, setting the stage for hyperphagia and reduced energy expenditure, hallmarks of obesity. Leptin in turn reverses this phenotype by increasing hypothalamic ObRb signaling (sensitivity) and affecting only the orexigenic arm of the neuropeptide balance.
Asunto(s)
Restricción Calórica , Ingestión de Energía/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Hipotálamo/efectos de los fármacos , Leptina/farmacología , Neuropéptidos/metabolismo , Efectos Tardíos de la Exposición Prenatal/patología , Factores de Edad , Proteína Relacionada con Agouti/metabolismo , Animales , Animales Recién Nacidos , Femenino , Hipotálamo/metabolismo , Masculino , Proteínas del Tejido Nervioso , Neuropéptido Y/metabolismo , Embarazo , Efectos Tardíos de la Exposición Prenatal/metabolismo , Proopiomelanocortina/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
Glucose is the primary energy source for most mammalian cells and its transport is affected by a family of facilitative glucose transporters (GLUTs) encoded by the SLC2 gene. GLUT1 and GLUT3, highly expressed isoforms in the blood-brain barrier and neuronal membranes, respectively, are associated with multiple neurodevelopmental disorders including epilepsy, dyslexia, ADHD, and autism spectrum disorder (ASD). Dietary therapies, such as the ketogenic diet, are widely accepted treatments for patients with the GLUT1 deficiency syndrome, while ameliorating certain symptoms associated with GLUT3 deficiency in animal models. A ketogenic diet, high-fat diet, and calorie/energy restriction during prenatal and postnatal stages can also alter the placental and brain GLUTs expression with long-term consequences on neurobehavior. This review focuses primarily on the role of diet/energy perturbations upon GLUT isoform-mediated emergence of neurodevelopmental and neurodegenerative disorders.
Asunto(s)
Encéfalo , Transportador de Glucosa de Tipo 1 , Transportador de Glucosa de Tipo 3 , Trastornos del Neurodesarrollo , Placenta , Transportador de Glucosa de Tipo 3/metabolismo , Transportador de Glucosa de Tipo 3/genética , Humanos , Embarazo , Encéfalo/metabolismo , Placenta/metabolismo , Femenino , Transportador de Glucosa de Tipo 1/metabolismo , Trastornos del Neurodesarrollo/etiología , Animales , Fenómenos Fisiologicos Nutricionales Maternos , Dieta Cetogénica , Dieta Alta en Grasa/efectos adversos , Enfermedades Neurodegenerativas/etiología , Enfermedades Neurodegenerativas/metabolismo , Glucosa/metabolismoRESUMEN
Introduction: Serotonin (5-HT) is critical for neurodevelopment and the serotonin transporter (SERT) modulates serotonin levels. Perturbed prenatal and postnatal dietary exposures affect the developing offspring predisposing to neurobehavioral disorders in the adult. We hypothesized that the postnatal brain 5-HT-SERT imbalance associated with gut dysbiosis forms the contributing gut-brain axis dependent mechanism responsible for such ultimate phenotypes. Methods: Employing maternal diet restricted (IUGR, n=8) and high fat+high fructose (HFhf, n=6) dietary modifications, rodent brain serotonin was assessed temporally by ELISA and SERT by quantitative Western blot analysis. Simultaneously, colonic microbiome studies were performed. Results: At early postnatal (P) day 2 no changes in the IUGR, but a ~24% reduction in serotonin (p = 0.00005) in the HFhf group occurred, particularly in the males (p = 0.000007) revealing a male versus female difference (p = 0.006). No such changes in SERT concentrations emerged. At late P21 the IUGR group reared on HFhf (IUGR/HFhf, (n = 4) diet revealed increased serotonin by ~53% in males (p = 0.0001) and 36% in females (p = 0.023). While only females demonstrated a ~40% decrease in serotonin (p = 0.010), the males only trended lower without a significant change within the HFhf group (p = 0.146). SERT on the other hand was no different in HFhf or IUGR/RC, with only the female IUGR/HFhf revealing a 28% decrease (p = 0.036). In colonic microbiome studies, serotonin-producing Bacteriodes increased with decreased Lactobacillus at P2, while the serotonin-producing Streptococcus species increased in IUGR/HFhf at P21. Sex-specific changes emerged in association with brain serotonin or SERT in the case of Alistipase, Anaeroplasma, Blautia, Doria, Lactococcus, Proteus, and Roseburia genera. Discussion: We conclude that an imbalanced 5-HT-SERT axis during postnatal brain development is sex-specific and induced by maternal dietary modifications related to postnatal gut dysbiosis. We speculate that these early changes albeit transient may permanently alter critical neural maturational processes affecting circuitry formation, thereby perturbing the neuropsychiatric equipoise.
RESUMEN
Air pollution (AP) is detrimental to pregnancies including increasing risk factors of gestational diabetes mellitus. We hypothesized that exposure to AP causes cardiovascular and metabolic disruption thereby altering placental gene expression, which in turn affects the placental phenotype and thereby embryonic/fetal development. To test this hypothesis, we investigated the impact of intra-nasal instilled AP upon gestational day 16-19 maternal mouse cardiovascular and metabolic status, placental nutrient transporters, and placental-fetal size and morphology. To further unravel mechanisms, we also examined placental total DNA 5'-hydroxymethylation and bulk RNA sequenced gene expression profiles. AP exposed pregnant mice and fetuses were tachycardic with a reduction in maternal left ventricular fractional shortening and increased uterine artery with decreased umbilical artery systolic peak velocities. In addition, they were hyperglycemic, glucose intolerant and insulin resistant, with changes in placental glucose (Glut3) and fatty acid (Fatp1 & Cd36) transporters, and a spatial disruption of cells expressing Glut10 that imports L-dehydroascorbic acid in protecting against oxidative stress. Placentas revealed inflammatory cellular infiltration with associated cellular edema and necrosis, with dilated vascular spaces and hemorrhage. Placental and fetal body weights decreased in mid-gestation with a reduction in brain cortical thickness emerging in late gestation. Placental total DNA 5'-hydroxymethylation was 2.5-fold higher, with perturbed gene expression profiles involving key metabolic, inflammatory, transcriptional, cellular polarizing and processing genes and pathways. We conclude that gestational exposure to AP incites a maternal inflammatory response resulting in features mimicking maternal gestational diabetes mellitus with altered placental DNA 5'-hydroxymethylation, gene expression, and associated injury.
Asunto(s)
Contaminantes Atmosféricos , Placenta , Femenino , Embarazo , Animales , Placenta/metabolismo , Placenta/efectos de los fármacos , Contaminantes Atmosféricos/toxicidad , Exposición Materna/efectos adversos , Fenotipo , Ratones Endogámicos C57BL , Metilación de ADN/efectos de los fármacos , Ratones , 5-Metilcitosina/análogos & derivados , 5-Metilcitosina/metabolismo , Feto/efectos de los fármacos , Feto/metabolismo , Desarrollo Fetal/efectos de los fármacosRESUMEN
Infants born with intrauterine growth retardation (IUGR) are at increased risk of adverse pulmonary outcomes at birth, including meconium aspiration and persistent pulmonary hypertension. Preterm infants with IUGR are at especially high risk of developing bronchopulmonary dysplasia (BPD), a disease hallmarked by alveolar hypoplasia. Although vitamin A supplementation has been shown to decrease the incidence of BPD or death in preterm very low birth weight infants, its potential to reduce BPD or death in preterm infants with IUGR remains unknown. We used a well-characterized rat model of caloric restriction to mimic IUGR and determine the impact of IUGR on lung development. We hypothesized that retinoic acid treatment would preserve alveolar formation through increases in key signaling molecules of the retinoic acid signaling pathway. Our results showed that alveolar hypoplasia caused by caloric restriction can be reversed with refeeding, and that retinoic acid prevents the alveolar hypoplasia coincident with the increased expression of elastin and retinoic acid receptor-α and decreased transforming growth factor-ß activity in developing rat lungs. These findings suggest that alveolar hypoplasia attributable to caloric restriction is reversible, and raises the possibility that retinoic acid therapy may prove a useful strategy to prevent adverse pulmonary sequelae such as BPD in preterm infants with IUGR.
Asunto(s)
Restricción Calórica , Hiperplasia/prevención & control , Pulmón/embriología , Exposición Materna , Alveolos Pulmonares/efectos de los fármacos , Tretinoina/farmacología , Animales , Elastina/metabolismo , Femenino , Embarazo , Alveolos Pulmonares/metabolismo , Alveolos Pulmonares/patología , Ratas , Ratas Sprague-Dawley , Receptores de Ácido Retinoico/metabolismo , Receptor alfa de Ácido Retinoico , Transducción de Señal , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
To investigate in vivo transcription of the facilitative glucose transporter isoform-GLUT3 gene, we created GLUT3-firefly luciferase transgenic mouse lines that demonstrate tissue-specific [adult: brain > testis ≥ skeletal muscle > placenta; postnatal (PN): skeletal muscle > brain = skin], temporal, and spatial distribution of the reporter gene/enzyme activity that is unique from endogenous GLUT3 mRNA/protein. In this mouse model, luciferase expression/activity serving as a readout of in vivo transcription peaked at 12 days gestation along with proliferating cell nuclear antigen (cell replication) in placenta and embryonic brain preceding peak GLUT3 protein expression at 18-19 days gestation. In contrast, a postnatal increase in brain luciferase mRNA peaked with endogenous GLUT3 mRNA, but after that of NeuroD6 protein (neurogenesis) at PN7. Luciferase activity paralleled GLUT3 protein expression with Na(+)-K(+)-ATPase (membrane expansion) and synaptophysin (synaptogenesis) proteins, peaking at PN14 and lasting until 60 days in the adult. Thus GLUT3 transcription in placenta and embryonic brain coincided with cell proliferation and in postnatal brain with synaptogenesis. Longitudinal noninvasive bioluminescence (BLI) monitoring of in vivo brain GLUT3 transcription reflected cross-sectional ex vivo brain luciferase activity only between PN7 and PN21. Hypoxia/reoxygenation at PN7 revealed transcriptional increase in brain GLUT3 expression reflected by in vivo BLI and ex vivo luciferase activity. These observations collectively support a temporal contribution by transcription toward ensuring adequate tissue-specific, developmental (placenta and embryonic brain), and postnatal hypoxic brain GLUT3 expression.
Asunto(s)
Encéfalo/metabolismo , Transportador de Glucosa de Tipo 3/metabolismo , Placenta/metabolismo , Activación Transcripcional/fisiología , Transgenes/fisiología , Animales , Femenino , Genes Reporteros/fisiología , Transportador de Glucosa de Tipo 3/genética , Luciferasas/fisiología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Transgénicos , Embarazo , Distribución TisularRESUMEN
We have shown in vitro a hypoxia-induced time-dependent increase in facilitative glucose transporter isoform 3 (GLUT3) expression in N2A murine neuroblasts. This increase in GLUT3 expression is partially reliant on a transcriptional increase noted in actinomycin D and cycloheximide pretreatment experiments. Transient transfection assays in N2A neuroblasts using murine glut3-luciferase reporter constructs mapped the hypoxia-induced enhancer activities to -857- to -573-bp and -203- to -177-bp regions. Hypoxia-exposed N2A nuclear extracts demonstrated an increase in HIF-1α and p-Creb binding to HRE (-828 to -824 bp) and AP-1 (-187 to -180 bp) cis-elements, respectively, in electromobility shift and supershift assays, which was confirmed by chromatin immunoprecipitation assays. In addition, the interaction of CBP with Creb and HIF-1α and CREST with CBP in hypoxia was detected by coimmunoprecipitation. Furthermore, small interference (si)RNA targeting Creb in these cells decreased endogenous Creb concentrations that reduced by twofold hypoxia-induced glut3 gene transcription. Thus, in N2A neuroblasts, phosphorylated HIF-1α and Creb mediated the hypoxia-induced increase in glut3 transcription. Coactivation by the Caâºâº-dependent CREST and CBP proteins may enhance cross-talk between p-Creb-AP-1 and HIF-1α/HRE of the glut3 gene. Collectively, these processes can facilitate an adaptive response to hypoxic energy depletion targeted at enhancing glucose transport and minimizing injury while fueling the proliferative potential of neuroblasts.
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Proteína de Unión a CREB/metabolismo , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Transportador de Glucosa de Tipo 3/biosíntesis , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Células-Madre Neurales/metabolismo , Transactivadores/metabolismo , Activación Transcripcional , Animales , Hipoxia de la Célula , Línea Celular , Núcleo Celular/metabolismo , Células Cultivadas , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/genética , Genes Reporteros , Transportador de Glucosa de Tipo 3/genética , Transportador de Glucosa de Tipo 3/metabolismo , Ratones , Ratones Endogámicos BALB C , Proteínas del Tejido Nervioso/biosíntesis , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Células-Madre Neurales/citología , Neuronas/citología , Neuronas/metabolismo , Transporte de Proteínas , Interferencia de ARN , Proteínas Recombinantes de Fusión/biosíntesis , Proteínas Recombinantes de Fusión/metabolismo , Transducción de Señal , Regulación hacia ArribaRESUMEN
Intra-Uterine Growth Restriction (IUGR) is a risk factor for many adult-onset chronic diseases, such as diabetes and obesity. These diseases are associated with intestinal microbiome perturbations (dysbiosis). The establishment of an intestinal microbiome begins in utero and continues postnatally (PN). Hypercaloric diet-induced dysbiosis is a major driver of childhood obesity. We hypothesized that different postnatal diets superimposed on IUGR will alter the postnatal intestinal microbiome. We compared four experimental rat groups: (1) Ad lib fed regular chow diet pre- and postnatally (CON), (2-3) IUGR induced by maternal caloric restriction prenatally followed postnatally (PN) by either (2) the control diet (IUGR-RC) or (3) High-Fat-high-fructose (IUGR-HFhf) diet, and lastly (4) HFhf ad lib pre- and postnatally (HFhf). Fecal samples were collected from dams and male and female rat offspring at postnatal day 2, 21, and adult day 180 for 16S rRNA gene sequencing. Maternal diet induced IUGR led to dysbiosis of the intestinal microbiome at PN21. Postnatal HFhf diet significantly reduced microbial diversity and worsened dysbiosis reflected by an increased Gammaproteobacteria/Clostridia ratio. Dysbiosis arising from a mismatch between IUGR and a postnatal HFhf diet may contribute to increased risk of the IUGR offspring for subsequent detrimental health problems.
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Microbioma Gastrointestinal , Obesidad Infantil , Niño , Humanos , Animales , Ratas , Masculino , Femenino , Disbiosis/complicaciones , ARN Ribosómico 16S/genética , Obesidad Infantil/complicaciones , Retardo del Crecimiento Fetal/etiología , Dieta , Dieta Alta en Grasa/efectos adversosRESUMEN
Associations between exponential childhood growth superimposed on low birth weight and adult onset cardiovascular disease with glucose intolerance/type 2 diabetes mellitus exist in epidemiological investigations. To determine the metabolic adaptations that guard against myocardial failure on subsequent exposure to hypoxia, we compared with controls (CON), the effect of intrauterine (IUGR), postnatal (PNGR), and intrauterine and postnatal (IPGR) calorie and growth restriction (n = 6/group) on myocardial macronutrient transporter (fatty acid and glucose) -mediated uptake in pregestational young female adult rat offspring. A higher myocardial FAT/CD36 protein expression in IUGR, PNGR, and IPGR, with higher FATP1 in IUGR, FATP6 in PNGR, FABP-c in PNGR and IPGR, and no change in GLUT4 of all groups was observed. These adaptive macronutrient transporter protein changes were associated with no change in myocardial [(3)H]bromopalmitate accumulation but a diminution in 2-deoxy-[(14)C]glucose uptake. Examination of the sarcolemmal subfraction revealed higher basal concentrations of FAT/CD36 in PNGR and FATP1 and GLUT4 in IUGR, PNGR, and IPGR vs. CON. Exogenous insulin uniformly further enhanced sarcolemmal association of these macronutrient transporter proteins above that of basal, with the exception of insulin resistance of FATP1 and GLUT4 in IUGR and FAT/CD36 in PNGR. The basal sarcolemmal macronutrient transporter adaptations proved protective against subsequent chronic hypoxic exposure (7 days) only in IUGR and PNGR, with notable deterioration in IPGR and CON of the echocardiographic ejection fraction. We conclude that the IUGR and PNGR pregestational adult female offspring displayed a resistance to insulin-induced translocation of FATP1, GLUT4, or FAT/CD36 to the myocardial sarcolemma due to preexistent higher basal concentrations. This basal adaptation of myocardial macronutrient transporters ensured adequate fatty acid uptake, thereby proving protective against chronic hypoxia-induced myocardial compromise.
Asunto(s)
Proteínas Portadoras/metabolismo , Retardo del Crecimiento Fetal/metabolismo , Trastornos del Crecimiento/metabolismo , Miocardio/metabolismo , Adaptación Fisiológica , Análisis de Varianza , Animales , Animales Recién Nacidos , Western Blotting , Peso Corporal/fisiología , Restricción Calórica/efectos adversos , Cateterismo Cardíaco , Membrana Celular/metabolismo , Desoxiglucosa/metabolismo , Ecocardiografía , Femenino , Hormonas/metabolismo , Hipoxia/metabolismo , Músculo Esquelético/citología , Músculo Esquelético/metabolismo , Miocardio/citología , Tamaño de los Órganos/fisiología , Palmitatos/farmacocinética , Embarazo , Radiofármacos/farmacocinética , Ratas , Ratas Sprague-DawleyRESUMEN
Energy balance is regulated by circulating leptin concentrations and hypothalamic leptin receptor (ObRb) signaling via STAT3 but is inhibited by SOCS3 and PTP1B. Leptin signaling enhances anorexigenic neuropeptides and receptor (POMC, MC3-R, MC4-R) activation while suppressing orexigenic neuropeptides (NPY, AgRP). We investigated in a sex-specific manner the early (PN2) and late (PN21) postnatal hypothalamic mechanisms in response to intrauterine (IUGR), postnatal (PNGR), and combined (IPGR) calorie and growth restriction. At PN2, both male and female IUGR were hypoleptinemic, but hypothalamic leptin signaling in females was activated as seen by enhanced STAT3. In addition, increased SOCS3 and PTP1B supported early initiation of leptin resistance in females that led to elevated AgRP but diminished MC3-R and MC4-R. In contrast, males demonstrated leptin sensitivity seen as a reduction in PTP1B and MC3-R and MC4-R with no effect on neuropeptide expression. At PN21, with adequate postnatal caloric intake, a sex-specific dichotomy in leptin concentrations was seen in IUGR, with euleptinemia in males indicative of persisting leptin sensitivity and hyperleptinemia in females consistent with leptin resistance, both with normal hypothalamic ObRb signaling, neuropeptides, and energy balance. In contrast, superimposition of PNGR upon IUGR (IPGR) led to diminished leptin concentrations with enhanced PTP1B and an imbalance in arcuate nuclear NPY/AgRP and POMC expression that favored exponential hyperphagia and diminished energy expenditure postweaning. We conclude that IUGR results in sex-specific leptin resistance observed mainly in females, whereas PNGR and IPGR abolish this sex-specificity, setting the stage for acquiring obesity after weaning.
Asunto(s)
Restricción Calórica , Metabolismo Energético/fisiología , Hipotálamo/metabolismo , Neuropéptidos/metabolismo , Efectos Tardíos de la Exposición Prenatal/patología , Factores de Edad , Proteína Relacionada con Agouti/genética , Análisis de Varianza , Animales , Animales Recién Nacidos , Peso Corporal/efectos de los fármacos , Peso Corporal/fisiología , Ingestión de Líquidos , Ingestión de Alimentos , Ingestión de Energía , Ensayo de Inmunoadsorción Enzimática , Femenino , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica/fisiología , Hipotálamo/efectos de los fármacos , Inyecciones Intraventriculares , Leptina/administración & dosificación , Leptina/sangre , Masculino , Leche/metabolismo , Neuropéptido Y/genética , Neuropéptido Y/metabolismo , Neuropéptidos/genética , Embarazo , Efectos Tardíos de la Exposición Prenatal/metabolismo , Efectos Tardíos de la Exposición Prenatal/fisiopatología , Proopiomelanocortina/genética , Proopiomelanocortina/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas Sprague-Dawley , Receptores de Leptina/genética , Receptores de Leptina/metabolismo , Fenómenos Fisiológicos Respiratorios , Factor de Transcripción STAT3/genética , Factor de Transcripción STAT3/metabolismo , Factores Sexuales , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiologíaRESUMEN
Paraoxonase 2 deficiency (PON2-def) alters mitochondrial function and exacerbates the development of atherosclerosis in mice. PON2 overexpression protects against ER stress in cell culture. In this paper, we examined the role of PON2 in the unexplored link between ER stress and mitochondrial dysfunction and tested whether restoration of PON2 in macrophages is sufficient to reduce aggravated atherosclerosis in PON2-def/apoE(-/-) mice on a Western diet. ER stress response genes, intracellular calcium levels, and apoptotic nuclei were significantly elevated in PON2-def/apoE(-/-) macrophages compared to apoE(-/-) macrophages in response to ER stressors, but not at the basal level. In contrast, PON2-def/apoE(-/-) macrophages exhibited greater mitochondrial stress at the basal level, which was further worsened in response to ER stressors. There was no difference in ER stress response genes and apoptotic nuclei between apoE(-/-) and PON2-def/apoE(-/-) macrophages when pretreated with xestospongin (which blocks the release of calcium from ER) suggesting that PON2 modulates cell survival and ER stress by maintaining calcium homeostasis. Treatment with a mitochondrial calcium uptake inhibitor, RU360, attenuated ER stressor mediated mitochondrial dysfunction in PON2-def/apoE(-/-) macrophages. CHOP expression (ER stress marker) and apoptotic nuclei were significantly higher in aortic lesions of PON2-def/apoE(-/-) mice compared to apoE(-/-) mice fed a Western diet. Restoration of PON2 in macrophages reduced ER stress, mitochondrial dysfunction and apoptosis in response to ER stressors. Furthermore, restoration of PON2 in macrophages reduced lesional apoptosis and atherosclerosis in PON2-def/apoE(-/-) mice on a Western diet. Our data suggest that macrophage PON2 modulates mechanisms that link ER stress, mitochondrial dysfunction and the development of atherosclerosis.
Asunto(s)
Apolipoproteínas E/deficiencia , Arildialquilfosfatasa/deficiencia , Aterosclerosis/metabolismo , Calcio/metabolismo , Estrés del Retículo Endoplásmico , Macrófagos/metabolismo , Animales , Aorta/efectos de los fármacos , Aorta/metabolismo , Aorta/patología , Apolipoproteínas E/genética , Apoptosis , Arildialquilfosfatasa/genética , Aterosclerosis/genética , Aterosclerosis/patología , Señalización del Calcio , Núcleo Celular/efectos de los fármacos , Supervivencia Celular , Dieta , Retículo Endoplásmico/metabolismo , Expresión Génica , Homeostasis , Compuestos Macrocíclicos/farmacología , Ratones , Ratones Noqueados , Mitocondrias/metabolismo , Oxazoles/farmacología , Estrés Oxidativo , Compuestos de Rutenio/farmacologíaRESUMEN
Since GLUT3 is vital for fueling neurotransmission, we examined in-vivo the adult phenotype carrying the conditional homozygous glut3 gene mutation (KO) in glutamate-excitatory neurons. These KO mice demonstrated sex-specific differences in brain and body weights (p = 0.0001 and p = 0.01 each) with reduced GLUT3 protein in cerebral cortices and brain stem (p = 0.005). In patch clamp studies the glut3 KO mice displayed a shorter latency to and enhanced paroxysmal activity (p = 0.01 and p = 0.015 each) in pyramidal neurons upon application of a GABAA antagonist, supporting hyperexcitability. Further, associated changes in neurobehavior consisted of reduced latency to fall in the rotorod motor test related to incoordination, increased distance traveled in total and periphery versus center in open field testing suggesting hyperactivity with anxiety (p = 0.0013 in male, p = 0.045 in female), reduced time freezing reminiscent of disrupted contextual fear conditioning (p = 0.0033), decreased time in target quadrant seen with spatial cognitive memory water maze testing (p = 0.034), and enhanced sociability particularly for novelty reflecting a lack of inhibition/impulsivity (p = 0.038). Some of these features were equally pronounced in males and females (cognitive) while others were seen in females (anxiety and impulsivity). We conclude that GLUT3 in adult glutamate-excitatory neurons is essential for maintaining neurotransmitory equipoise regulating excitation with maintenance of motor coordination and activity, cognition, spatial memory and normal fear for both contextual events and novelty with tempered sociability. While sex-specificity was forthcoming for some of these behaviors, our findings collectively suggest that loss-of-function glut3 gene mutations or polymorphisms may underlie an endophenotype of attention deficit-hyperactivity disorder.
Asunto(s)
Conducta Animal/fisiología , Encéfalo/fisiopatología , Transportador de Glucosa de Tipo 3/genética , Trastornos del Neurodesarrollo/genética , Animales , Encéfalo/metabolismo , Modelos Animales de Enfermedad , Femenino , Transportador de Glucosa de Tipo 3/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Trastornos del Neurodesarrollo/metabolismo , Trastornos del Neurodesarrollo/fisiopatología , Células Piramidales/metabolismoRESUMEN
We examined the effects of 45-min hypoxia (FiO(2) 0.08; Hx) vs. normoxia (FiO(2) 0.21; Nx) on the ipsilateral (Ipsi) and contralateral (Ctrl) sides of the brain in neuronal glucose transporter isoform 3 (Glut3) heterozygous null mice (glut3(+/-)) and their wild-type littermates (WT), undergoing unilateral carotid artery ligation. Glut3(+/-) mice, under Nx, demonstrated a compensatory increase in blood-brain barrier/glial Glut1 protein concentration and a concomitant increase in neuronal nitric oxide synthase (nNOS) enzyme activity and Bax protein, with a decrease in procaspase 3 protein (P < 0.05 each). After Hx, reoxygenation in FiO(2) of 0.21 led to no comparable adaptive up-regulation of the ipsilateral brain Glut3 or Glut1 protein at 4 hr and Glut1 at 24 hr in glut3(+/-) vs. WT. These brain Glut changes in glut3(+/-) but not WT mice were associated with an increase in proapoptotic Bax protein and caspase-3 enzyme activity (P < 0.01 each) and a decline in the antiapoptotic Bcl-2 and procaspase-3 proteins (P < 0.05 each). Glut3(+/-) mice after Hx demonstrated TUNEL-positive neurons with nuclear pyknosis in most ipsilateral (hypoxic-ischemia) brain regions. A subset (â¼55%) of glut3(+/-) mice developed spontaneous seizures after hypoxic-ischemia, confirmed by electroencephalography, but the WT mice remained seizure-free. Pentylenetetrazole testing demonstrated an increased occurrence of longer lasting clinical seizures at a lower threshold in glut3(+/-) vs. WT mice, with no detectable differences in monamine neurotransmitters. We conclude that hypoxic-ischemic brain injury in glut3(+/-) mice exacerbates cellular apoptosis and necrosis and precipitates spontaneous seizures.
Asunto(s)
Apoptosis/fisiología , Transportador de Glucosa de Tipo 3/metabolismo , Hipoxia-Isquemia Encefálica/fisiopatología , Neuronas/patología , Convulsiones/fisiopatología , Animales , Western Blotting , Cromatografía Líquida de Alta Presión , Femenino , Técnica del Anticuerpo Fluorescente , Transportador de Glucosa de Tipo 1/metabolismo , Transportador de Glucosa de Tipo 3/genética , Heterocigoto , Hipoxia-Isquemia Encefálica/genética , Hipoxia-Isquemia Encefálica/metabolismo , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Mutantes , Neuronas/metabolismo , Fenotipo , Convulsiones/genética , Convulsiones/metabolismoRESUMEN
We examined the role of hepatocyte micro-RNA-122 and hypothalamic neuropeptides, in weanling (21d) female rats exposed to calorie restriction induced growth restriction either prenatally (IUGR), postnatally (PNGR) or both (IPGR) vs. ad lib fed controls (CON). IUGR were hyperinsulinemic, hyperleptinemic and dyslipidemic with high circulating miR-122. In contrast, PNGR and IPGR displayed insufficient glucose, insulin and leptin amidst high ketones with a dichotomy in circulating miR-122 of PNGRAsunto(s)
Restricción Calórica/efectos adversos
, Metabolismo de los Lípidos/genética
, Metabolismo de los Lípidos/fisiología
, MicroARNs/fisiología
, Animales
, Peso Corporal
, Ritmo Circadiano
, Metabolismo Energético
, Ácidos Grasos/genética
, Ácidos Grasos/metabolismo
, Femenino
, Retardo del Crecimiento Fetal/etiología
, Expresión Génica
, Trastornos del Crecimiento/etiología
, Hipotálamo/química
, Hígado/química
, Hígado/metabolismo
, MicroARNs/genética
, Tamaño de los Órganos
, Embarazo
, ARN Mensajero/análisis
, Ratas
, Ratas Sprague-Dawley
, Destete
RESUMEN
Diet during pregnancy has long lasting consequences on the offspring, warranting a study on the impact of early exposure to a high fat diet on the adult offspring. We hypothesized that a prenatal n-6 enriched diet will have adverse metabolic outcomes on the adult offspring that may be reversed with a postnatal n-3 enriched diet. To test this hypothesis, we examined the adult offspring from three groups: (1) n-6 group: during gestation and lactation, dams consumed an n-6 polyunsaturated fatty acid enriched diet, (2) n-3 group: gestational n-6 diet was followed by an n-3 enriched diet during lactation, and (3) a control (CD) group that received standard diet throughout gestation and lactation. Offspring from all groups weaned to a control diet ad libitum. Beginning at postnatal day 2 (Pâ¯<â¯.03) and persisting at 360 days in males (Pâ¯<â¯.04), an increase in hypothalamic AgRP expression occurred in the n-6 and n-3 groups, with an increase in food intake (Pâ¯=â¯.01), and the n-3 group displaying lower body (Pâ¯<â¯.03) and brain (Pâ¯<â¯.05) weights. At 360 days, the n-6 and n-3 groups remained glucose tolerant and insulin sensitive, with increased phosphorylated-AMP-activated protein kinase (Pâ¯<â¯.05). n-6 group developed hepatic steatosis with reduced hepatic reflected as higher plasma microRNA-122 (Pâ¯<â¯.04) that targets pAMPK. We conclude that early life exposure to n-6 and n-3 led to hypothalamic AgRP-related higher food intake, with n-6 culminating in a fatty liver partially mitigated by postnatal n-3. While both diets preserved glucose tolerance and insulin sensitivity, postnatal n-3 displayed detrimental effects on the brain.