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The coordination of food intake, energy storage, and expenditure involves complex interactions between hypothalamic neurons and peripheral tissues including pancreatic islets, adipocytes, muscle, and liver. Previous research shows that deficiency of the transcription factor Alx3 alters pancreatic islet-dependent glucose homeostasis. In this study we carried out a comprehensive assessment of metabolic alterations in Alx3 deficiency. We report that Alx3-deficient mice exhibit decreased food intake without changes in body weight, along with reduced energy expenditure and altered respiratory exchange ratio. Magnetic resonance imaging reveals increased adiposity and decreased muscle mass, which was associated with markers of motor and sympathetic denervation. By contrast, Alx3-deficient mice on a high-fat diet show attenuated weight gain and improved insulin sensitivity, compared to control mice. Gene expression analysis demonstrates altered lipogenic and lipolytic gene profiles. In wild type mice Alx3 is expressed in hypothalamic arcuate nucleus neurons, but not in major peripheral metabolic organs. Functional diffusion-weighted magnetic resonance imaging reveals selective hypothalamic responses to fasting in the arcuate nucleus of Alx3-deficient mice. Additionally, altered expression of proopiomelanocortin and melanocortin-3 receptor mRNA in the hypothalamus suggests impaired regulation of feeding behavior. This study highlights the crucial role for Alx3 in governing food intake, energy homeostasis, and metabolic nutrient partitioning, thereby influencing body mass composition.
Assuntos
Composição Corporal , Ingestão de Alimentos , Metabolismo Energético , Proteínas de Homeodomínio , Homeostase , Hipotálamo , Camundongos Knockout , Animais , Metabolismo Energético/genética , Hipotálamo/metabolismo , Camundongos , Ingestão de Alimentos/genética , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Dieta Hiperlipídica , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Masculino , Camundongos Endogâmicos C57BL , Neurônios/metabolismo , Pró-Opiomelanocortina/metabolismo , Pró-Opiomelanocortina/genética , Resistência à Insulina/genética , Núcleo Arqueado do Hipotálamo/metabolismoRESUMO
BACKGROUND: The current study sought to explore whether cancer pain (CP) already exists in patients at colorectal cancer (CRC) diagnosis before treatment compared with patients with colorectal cancer (CRC) after treatment and a healthy matched control group. The study also sought to examine whether factors related to physical health status could enhance pain processes. METHODS: An observational cross-sectional study was conducted following the STROBE checklist. Twenty-nine newly diagnosed and forty post-treatment patients with CRC and 40 healthy age/sex-matched controls were included for comparison. Pain, local muscle function, and body composition outcomes were assessed by a physiotherapist with > 3 years of experience. ANCOVA and Kruskal-Wallis tests were performed, with Bonferroni and Dunn-Bonferroni post hoc analyses and Cohen's d and Hedge's effect size, as appropriate. RESULTS: The analysis detected lower values of pressure pain threshold (PPT) points, the PPT index, and abdominal strength and higher values of self-reported abdominal pain in newly diagnosed patients, with even more marked results observed in the post-treatment patients, where lower lean mass and skeletal muscle index values were also found than those in the healthy matched controls (p < 0.05). In the post-treatment and healthy matched control groups, positive associations were observed between the PPT lumbar dominant side points and abdominal isometric strength and lean mass, and negative associations were observed between the lumbar dominant side points and body fat (p < 0.05). CONCLUSION: Upon diagnosis, patients with CRC already show signs of hyperalgesia and central sensitization and deteriorated physical conditions and body composition, and this state could be aggravated by subsequent treatments.
Assuntos
Dor do Câncer , Neoplasias Colorretais , Composição Corporal , Dor do Câncer/diagnóstico , Dor do Câncer/etiologia , Neoplasias Colorretais/complicações , Neoplasias Colorretais/diagnóstico , Estudos Transversais , Humanos , Limiar da Dor/fisiologiaRESUMO
Postnatal overfeeding increases the risk of chronic diseases later in life, including obesity, insulin resistance, hepatic steatosis, and type 2 diabetes. Epigenetic mechanisms might underlie the long-lasting effects associated with early nutrition. Here we aimed to explore the molecular pathways involved in early development of insulin resistance and hepatic steatosis, and we examined the potential contribution of DNA methylation and histone modifications to long-term programming of metabolic disease. We used a well-characterized mouse model of neonatal overfeeding and early adiposity by litter size reduction. Neonatal overfeeding led to hepatic insulin resistance very early in life that persisted throughout adulthood despite normalizing food intake. Up-regulation of monoacylglycerol O-acyltransferase ( Mogat) 1 conceivably mediates hepatic steatosis and insulin resistance through increasing intracellular diacylglycerol content. Early and sustained deregulation of Mogat1 was associated with a combination of histone modifications that might favor Mogat1 expression. In sum, postnatal overfeeding causes extremely rapid derangements of hepatic insulin sensitivity that remain relatively stable until adulthood. Epigenetic mechanisms, particularly histone modifications, could contribute to such long-lasting effects. Our data suggest that targeting hepatic monoacylglycerol acyltransferase activity during early life might provide a novel strategy to improve hepatic insulin sensitivity and prevent late-onset insulin resistance and fatty liver disease.-Ramon-Krauel, M., Pentinat, T., Bloks, V. W., Cebrià, J., Ribo, S., Pérez-Wienese, R., Vilà, M., Palacios-Marin, I., Fernández-Pérez, A., Vallejo, M., Téllez, N., Rodríguez, M. À., Yanes, O., Lerin, C., Díaz, R., Plosch, T., Tietge, U. J. F., Jimenez-Chillaron, J. C. Epigenetic programming at the Mogat1 locus may link neonatal overnutrition with long-term hepatic steatosis and insulin resistance.
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Efficient intracellular cargo delivery is a key hurdle for the translation of many emerging stem cell and cellular reprogramming therapies. Recently, a microfluidic-based device constructed from silicon was shown to transduce macromolecules into cells via shear-induced formation of plasma membrane pores. However, the scalability and widespread application of the current platform is limited since physical deformation-mediated delivery must be optimized for each therapeutic application. Therefore, we sought to create a low-cost, versatile device that could facilitate rapid prototyping and application-specific optimization in most academic research labs. Here we describe the design and implementation of a microfluidic device constructed from Polydimethylsiloxane (PDMS) that we call Cyto-PDMS (Cytoplasmic PDMS-based Delivery and Modification System). Using a systematic Cyto-PDMS workflow, we demonstrate intracellular cargo delivery with minimal effects on cellular viability. We identify specific flow rates at which a wide range of cargo sizes (1-70 kDa) can be delivered to the cell interior. As a proof-of-principle for the biological utility of Cyto-PDMS, we show (i) F-actin labeling in live human fibroblasts and (ii) intracellular delivery of recombinant Cre protein with appropriate genomic recombination in recipient fibroblasts. Taken together, our results demonstrate that Cyto-PDMS can deliver small-molecules to the cytoplasm and biologically active cargo to the nucleus without major effects on viability. We anticipate that the cost and versatility of PDMS can be leveraged to optimize delivery to a broad array of possible cell types and thus expand the potential impact of cellular therapies.
Assuntos
Fibroblastos/metabolismo , Dispositivos Lab-On-A-Chip , Actinas/metabolismo , Animais , Linhagem Celular , Sobrevivência Celular , Dimetilpolisiloxanos , Portadores de Fármacos/química , Desenho de Equipamento , Fibroblastos/citologia , Humanos , Camundongos , Peso MolecularRESUMO
Various combinations of cardiogenic transcription factors, including Gata4 (G), Hand2 (H), Mef2c (M) and Tbx5 (T), can reprogram fibroblasts into induced cardiac-like myocytes (iCLMs) in vitro and in vivo. Given that optimal cardiac function relies on distinct yet functionally interconnected atrial, ventricular and pacemaker (PM) cardiomyocytes (CMs), it remains to be seen which subtypes are generated by direct reprogramming and whether this process can be harnessed to produce a specific CM of interest. Here, we employ a PM-specific Hcn4-GFP reporter mouse and a spectrum of CM subtype-specific markers to investigate the range of cellular phenotypes generated by reprogramming of primary fibroblasts. Unexpectedly, we find that a combination of four transcription factors (4F) optimized for Hcn4-GFP expression does not generate beating PM cells due to inadequate sarcomeric protein expression and organization. However, applying strict single-cell criteria to GHMT-reprogrammed cells, we observe induction of diverse cellular phenotypes, including those resembling immature forms of all three major cardiac subtypes (i.e. atrial, ventricular and pacemaker). In addition, we demonstrate that cells induced by GHMT are directly reprogrammed and do not arise from an Nxk2.5(+) progenitor cell intermediate. Taken together, our results suggest a remarkable degree of plasticity inherent to GHMT reprogramming and provide a starting point for optimization of CM subtype-specific reprogramming protocols.
Assuntos
Diferenciação Celular/fisiologia , Indução Embrionária/fisiologia , Fibroblastos/citologia , Coração/embriologia , Miócitos Cardíacos/fisiologia , Fatores de Transcrição/metabolismo , Potenciais de Ação/fisiologia , Análise de Variância , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Primers do DNA/genética , Fibroblastos/metabolismo , Fibroblastos/fisiologia , Fator de Transcrição GATA4/genética , Fator de Transcrição GATA4/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Canais Disparados por Nucleotídeos Cíclicos Ativados por Hiperpolarização/metabolismo , Imuno-Histoquímica , Fatores de Transcrição MEF2/genética , Fatores de Transcrição MEF2/metabolismo , Camundongos , Miócitos Cardíacos/citologia , Reação em Cadeia da Polimerase em Tempo Real , Proteínas com Domínio T/genética , Proteínas com Domínio T/metabolismo , Fatores de Transcrição/genéticaRESUMO
OBJECTIVE: To investigate the efficacy of an eight-week lumbopelvic stabilization program (CO-CUIDATE) for colon cancer survivors. DESIGN: A secondary analysis of a randomized controlled clinical trial. SETTINGS: A blinded, trained researcher performed the end point assessments for pain (Pressure Pain Threshold and Brief Pain Inventory) and muscle architecture (ultrasound imaging measurements). SUBJECTS: Forty-six colon cancer survivors who were assigned to the CO-CUIDATE group or usual care group. METHODS: The CO-CUIDATE program was conducted for eight weeks. A trained researcher who was blinded to patient group performed the assessments. The tests were carried out with multiple observations. Intention-to-treat analyses were performed. RESULTS: The program had an adherence rate of 88.36% and two dropouts (10.5%). The participants reported some minor side effects during the first exercise sessions. The analysis revealed significant differences in the group x time interactions for the lumbar side (dominant: F = 3.1, P < 0.001; nondominant: F = 3.0, P = 0.01) and the infra-umbilical dominant side (F = 1.2, P = 0.04) after the program and at the six-month follow-up and for the internal oblique thickness (F = 5.1, P = 0.030) after the program. The experimental group experienced a greater improvement in all values after the program compared with the control group. There were no significant changes in the other pressure pain threshold points, pain severity, interference of pain, or the remaining ultrasound imaging measurements. CONCLUSION: The CO-CUIDATE program is effective for improving the musculoskeletal conditions related to the lumbopelvic area in colon cancer survivors, specifically in relation to pain and the internal oblique thickness.
Assuntos
Músculos Abdominais/diagnóstico por imagem , Sobreviventes de Câncer , Neoplasias do Colo/diagnóstico por imagem , Neoplasias do Colo/terapia , Terapia por Exercício/tendências , Medição da Dor/tendências , Músculos Abdominais/fisiologia , Idoso , Exercício Físico/fisiologia , Terapia por Exercício/métodos , Feminino , Seguimentos , Humanos , Região Lombossacral/diagnóstico por imagem , Masculino , Pessoa de Meia-Idade , Força Muscular/fisiologia , Pelve/diagnóstico por imagem , Exercício Pliométrico/métodos , Exercício Pliométrico/tendênciasRESUMO
Although all authors report beneficial health changes following training based on the Pilates method, no explicit analysis has been performed of its cardiorespiratory effects. The objective of this study was to evaluate possible changes in cardiorespiratory parameters with the Pilates method. A total of 45 university students aged 18-35 years (77.8% female and 22.2% male), who did not routinely practice physical exercise or sports, volunteered for the study and signed informed consent. The Pilates training was conducted over 10 weeks, with three 1-hour sessions per week. Physiological cardiorespiratory responses were assessed using a MasterScreen CPX apparatus. After the 10-week training, statistically significant improvements were observed in mean heart rate (135.4-124.2 beats/min), respiratory exchange ratio (1.1-0.9) and oxygen equivalent (30.7-27.6) values, among other spirometric parameters, in submaximal aerobic testing. These findings indicate that practice of the Pilates method has a positive influence on cardiorespiratory parameters in healthy adults who do not routinely practice physical exercise activities.
Assuntos
Adaptação Fisiológica , Técnicas de Exercício e de Movimento , Condicionamento Físico Humano/fisiologia , Adolescente , Adulto , Teste de Esforço , Feminino , Frequência Cardíaca , Humanos , Masculino , Consumo de Oxigênio , Troca Gasosa Pulmonar , Espirometria , Adulto JovemRESUMO
Alterations in transcription factors expressed in insulin-producing islet ß-cells generate pancreatic dysfunction leading to diabetes. The homeodomain transcription factor Alx3 (aristaless-like homeobox 3) expressed in pancreatic islets participates in the regulated expression of several islet genes, and its deficiency in mice leads to islet cell apoptosis and glucose intolerance. In the present study, we investigated the mechanisms that regulate expression of Alx3 in pancreatic islets at the transcriptional level. We found that the Alx3 promoter contains at least eight putative regulatory elements with an E-box consensus sequence, three of which were determined to be functional and required for Alx3 promoter activity by mutational analysis in transfected MIN6 ß-cells. We determined that these E-box elements are recognized by the basic helix-loop-helix transcription factors USF1 (upstream stimulatory factor 1) and USF2. We also identified a highly conserved A-box in the Alx3 promoter that is recognized by the islet-specific transcription factor Pdx1 (pancreatic and duodenal homeobox 1). Pdx1-mediated transactivation of the Alx3 promoter requires the integrity of the three functional E-boxes and the co-operation with USF transcription factors bound to them. The results from the present study indicate that Pdx1 contributes to the transcriptional transactivation of Alx3 in pancreatic ß-cells by acting in co-ordination with USF1 and USF2.
Assuntos
Regulação da Expressão Gênica , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Ilhotas Pancreáticas/metabolismo , Transativadores/metabolismo , Fatores Estimuladores Upstream/metabolismo , Animais , Linhagem Celular , Camundongos , Transativadores/genética , Fatores Estimuladores Upstream/genéticaRESUMO
During embryonic development, the aristaless-type homeodomain protein Alx3 is expressed in the forehead mesenchyme and contributes to the regulation of craniofacial development. In the adult, Alx3 is expressed in pancreatic islets where it participates in the control of glucose homoeostasis. In the present study, we investigated the transcriptional regulation of Alx3 gene expression in these two cell types. We found that the Alx3 promoter contains two E-box regulatory elements, named EB1 and EB2, that provide binding sites for the basic helix-loop-helix transcription factors Twist1, E47, USF (upstream stimulatory factor) 1 and USF2. In primary mouse embryonic mesenchymal cells isolated from the forehead, EB2 is bound by Twist1, whereas EB1 is bound by USF1 and USF2. Integrity of both EB1 and EB2 is required for Twist1-mediated transactivation of the Alx3 promoter, even though Twist1 does not bind to EB1, indicating that binding of USF1 and USF2 to this element is required for Twist1-dependent Alx3 promoter activity. In contrast, in pancreatic islet insulin-producing cells, the integrity of EB2 is not required for proximal promoter activity. The results of the present study indicate that USF1 and USF2 are important regulatory factors for Alx3 gene expression in different cell types, whereas Twist1 contributes to transcriptional transactivation in mesenchymal, but not in pancreatic, cells.
Assuntos
Proteínas de Homeodomínio/genética , Proteínas Nucleares/genética , Regiões Promotoras Genéticas , Proteína 1 Relacionada a Twist/genética , Fatores Estimuladores Upstream/genética , Animais , Sítios de Ligação , Células COS , Linhagem Celular , Chlorocebus aethiops , Embrião de Mamíferos/metabolismo , Feminino , Células Secretoras de Glucagon/metabolismo , Células HeLa , Proteínas de Homeodomínio/metabolismo , Humanos , Mesoderma/metabolismo , Camundongos , Células NIH 3T3 , Proteínas Nucleares/metabolismo , Gravidez , Proteína 1 Relacionada a Twist/metabolismo , Fatores Estimuladores Upstream/metabolismoRESUMO
Metabolic homeostasis is under circadian regulation to adapt energy requirements to light-dark cycles. Feeding cycles are regulated by photic stimuli reaching the suprachiasmatic nucleus via retinohypothalamic axons and by nutritional information involving dopaminergic neurotransmission. Previously, we reported that Pitx3-mutant Aphakia mice with altered development of the retinohypothalamic tract and the dopaminergic neurons projecting to the striatum, are resistant to locomotor and metabolic entrainment by time-restricted feeding. In their Matters Arising article, Scarpa et al. (2022) challenge this conclusion using mice from the same strain but following a different experimental paradigm involving calorie restriction. Here, we address their concerns by extending the analyses of our previous data, by identifying important differences in the experimental design between both studies and by presenting additional results on the dopaminergic deficit in the brain of Aphakia mice. This Matters Arising Response article addresses the Matters Arising article by Scarpa et al. (2022), published concurrently in Cell Reports.
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Afacia , Núcleo Supraquiasmático , Animais , Dopamina , Metabolismo Energético , Camundongos , FotoperíodoRESUMO
The cardiac conduction system (CCS) ensures regular contractile function, and injury to any of its components can cause cardiac dysrhythmia. Although all cardiomyocytes (CMs) originate from common progenitors, the CCS is composed of biologically distinct cell types with unique functional and developmental characteristics. In contrast to ventricular cardiomyocytes, which continue to proliferate after birth, most CCS cells terminally exit the cell cycle during fetal development. Although the CCS should thus provide a poor substrate for postnatal injury repair, its regenerative capacity remains untested. Here, we describe a genetic system for ablating CMs that reside within the atrioventricular conduction system (AVCS). Adult mouse AVCS ablation resulted in regenerative failure characterized by persistent atrioventricular conduction defects and contractile dysfunction. In contrast, AVCS injury in neonatal mice led to recovery in a subset of these mice, thus providing evidence for CCS plasticity. Furthermore, CM proliferation did not appear to completely account for the observed functional recovery, suggesting that mechanisms regulating recovery from dysrhythmia are likely to be distinct from cardiac regeneration associated with ventricular injury. Taken together, we anticipate that our results will motivate further mechanistic studies of CCS plasticity and enable the exploration of rhythm restoration as an alternative therapeutic strategy.
Assuntos
Nó Atrioventricular/lesões , Miócitos Cardíacos/fisiologia , Regeneração/fisiologia , Animais , Nó Atrioventricular/fisiologia , Plasticidade Celular/fisiologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
OBJECTIVE: Pancreatic ß-cell dysfunction is a central feature in the pathogenesis of type 2 diabetes (T2D). Accumulating evidence indicates that ß-site APP-cleaving enzyme 2 (BACE2) inhibition exerts a beneficial effect on ß-cells in different models of T2D. Thus, targeting BACE2 may represent a potential therapeutic strategy for the treatment of this disease. Here, we aimed to investigate the effects of BACE2 suppression on glucose homeostasis in a model of diet-induced obesity. METHODS: BACE2 knock-out (BKO) and wild-type (WT) mice were fed with a high-fat diet (HFD) for 2 or 16 weeks. Body weight, food intake, respiratory exchange ratio, locomotor activity, and energy expenditure were determined. Glucose homeostasis was evaluated by glucose and insulin tolerance tests. ß-cell proliferation was assessed by Ki67-positive nuclei, and ß-cell function was determined by measuring glucose-stimulated insulin secretion. Leptin sensitivity was evaluated by quantifying food intake and body weight after an intraperitoneal leptin injection. Neuropeptide gene expression and insulin signaling in the mediobasal hypothalamus were determined by qPCR and Akt phosphorylation, respectively. RESULTS: After 16 weeks of HFD feeding, BKO mice exhibited an exacerbated body weight gain and hyperphagia, in comparison to WT littermates. Glucose tolerance was similar in both groups, whereas HFD-induced hyperinsulinemia, insulin resistance, and ß-cell expansion were more pronounced in BKO mice. In turn, leptin-induced food intake inhibition and hypothalamic insulin signaling were impaired in BKO mice, regardless of the diet, in accordance with deregulation of the expression of hypothalamic neuropeptide genes. Importantly, BKO mice already showed increased ß-cell proliferation and glucose-stimulated insulin secretion with respect to WT littermates after two weeks of HFD feeding, before the onset of obesity. CONCLUSIONS: Collectively, these results reveal that BACE2 suppression in an obesogenic setting leads to exacerbated body weight gain, hyperinsulinemia, and insulin resistance. Thus, we conclude that inhibition of BACE2 may aggravate the adverse metabolic effects associated with obesity.
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Secretases da Proteína Precursora do Amiloide/metabolismo , Ácido Aspártico Endopeptidases/metabolismo , Obesidade/metabolismo , Animais , Dieta/efeitos adversos , Masculino , Camundongos , Camundongos TransgênicosRESUMO
Childhood obesity is a strong risk factor for adult obesity, type 2 diabetes, and cardiovascular disease. The mechanisms that link early adiposity with late-onset chronic diseases are poorly characterised. We developed a mouse model of early adiposity through litter size reduction. Mice reared in small litters (SLs) developed obesity, insulin resistance, and hepatic steatosis during adulthood. The liver played a major role in the development of the disease. OBJECTIVE: To gain insight into the molecular mechanisms that link early development and childhood obesity with adult hepatic steatosis and insulin resistance. METHODS: We analysed the hepatic transcriptome (Affymetrix) of control and SL mice to uncover potential pathways involved in the long-term programming of disease in our model. RESULTS: The circadian rhythm was the most significantly deregulated Gene Ontology term in the liver of adult SL mice. Several core clock genes, such as period 1-3 and cryptochrome 1-2, were altered in two-week-old SL mice and remained altered throughout their life course until they reached 4-6 months of age. Defective circadian rhythm was restricted to the periphery since the expression of clock genes in the hypothalamus, the central pacemaker, was normal. The period-cryptochrome genes were primarily entrained by dietary signals. Hence, restricting food availability during the light cycle only uncoupled the central rhythm from the peripheral and completely normalised hepatic triglyceride content in adult SL mice. This effect was accompanied by better re-alignment of the hepatic period genes, suggesting that they might have played a causal role in mediating hepatic steatosis in the adult SL mice. Functional downregulation of Per2 in hepatocytes in vitro confirmed that the period genes regulated lipid-related genes in part through peroxisome proliferator-activated receptor alpha (Ppara). CONCLUSIONS: The hepatic circadian rhythm matures during early development, from birth to postnatal day 30. Hence, nutritional challenges during early life may misalign the hepatic circadian rhythm and secondarily lead to metabolic derangements. Specific time-restricted feeding interventions improve metabolic health in the context of childhood obesity by partially re-aligning the peripheral circadian rhythm.
Assuntos
Ritmo Circadiano/fisiologia , Lactação , Fígado/metabolismo , Hepatopatia Gordurosa não Alcoólica/metabolismo , Adiposidade , Adulto , Animais , Ritmo Circadiano/genética , Diabetes Mellitus Tipo 2/metabolismo , Jejum , Feminino , Humanos , Hipotálamo/metabolismo , Recém-Nascido , Resistência à Insulina/fisiologia , Doenças Metabólicas/metabolismo , Camundongos , Camundongos Endogâmicos ICR , Hepatopatia Gordurosa não Alcoólica/genética , Obesidade/metabolismo , Obesidade InfantilRESUMO
Accelerated postnatal growth is a potentially modifiable risk factor for future obesity. To study how specific breast milk components contribute to early growth and obesity risk, we quantified one-carbon metabolism-related metabolites in human breast milk and found an inverse association between milk betaine content and infant growth. This association was replicated in an independent and geographically distinct cohort. To determine the potential role of milk betaine in modulating offspring obesity risk, we performed maternal betaine supplementation experiments in mice. Higher betaine intake during lactation increased milk betaine content in dams and led to lower adiposity and improved glucose homeostasis throughout adulthood in mouse offspring. These effects were accompanied by a transient increase in Akkermansia spp. abundance in the gut during early life and a long-lasting increase in intestinal goblet cell number. The link between breast milk betaine and Akkermansia abundance in the gut was also observed in humans, as infants exposed to higher milk betaine content during breastfeeding showed higher fecal Akkermansia muciniphila abundance. Furthermore, administration of A. muciniphila to mouse pups during the lactation period partially replicated the effects of maternal breast milk betaine, including increased intestinal goblet cell number, lower adiposity, and improved glucose homeostasis during adulthood. These data demonstrate a link between breast milk betaine content and long-term metabolic health of offspring.
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Betaína , Leite Humano , Akkermansia , Animais , Dieta Hiperlipídica , Feminino , Lactação , CamundongosRESUMO
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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In the developing mouse brain, once the generation of neurons is mostly completed during the prenatal period, precisely coordinated signals act on competent neural precursors to direct their differentiation into astrocytes, which occurs mostly after birth. Among these signals, those provided by neurotrophic cytokines and bone morphogenetic proteins appear to have a key role in triggering the neurogenic to gliogenic switch and in regulating astrocyte numbers. In addition, we have reported previously that the neurotrophic peptide pituitary adenylate cyclase-activating polypeptide (PACAP) is able to promote astrocyte differentiation of cortical precursors via activation of a cAMP-dependent pathway. Signals acting on progenitor cells of the developing cortex to generate astrocytes activate glial fibrillary acidic protein (GFAP) gene expression, but the transcriptional mechanisms that regulate this activation are unclear. Here, we identify the previously known transcriptional repressor downstream regulatory element antagonist modulator (DREAM) as an activator of GFAP gene expression. We found that DREAM occupies specific sites on the GFAP promoter before and after differentiation is initiated by exposure of cortical progenitor cells to PACAP. PACAP raises intracellular calcium concentration via a mechanism that requires cAMP, and DREAM-mediated transactivation of the GFAP gene requires the integrity of calcium-binding domains. Cortical progenitor cells from dream(-/-) mice fail to express GFAP in response to PACAP. Moreover, the neonatal cortex of dream(-/-) mice exhibits a reduced number of astrocytes and increased number of neurons. These results identify the PACAP-cAMP-Ca(2+)-DREAM cascade as a new pathway to activate GFAP gene expression during astrocyte differentiation.
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Astrócitos/metabolismo , Sinalização do Cálcio/genética , Córtex Cerebral/embriologia , AMP Cíclico/metabolismo , Proteína Glial Fibrilar Ácida/genética , Proteínas Interatuantes com Canais de Kv/genética , Proteínas Repressoras/genética , Animais , Astrócitos/citologia , Astrócitos/efeitos dos fármacos , Cálcio/metabolismo , Cálcio/farmacologia , Sinalização do Cálcio/efeitos dos fármacos , Contagem de Células , Diferenciação Celular/genética , Células Cultivadas , Córtex Cerebral/citologia , Córtex Cerebral/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios/metabolismo , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase/genética , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase/metabolismo , Regiões Promotoras Genéticas/genética , Ratos , Ratos Wistar , Transdução de Sinais/genéticaRESUMO
Pituitary adenylate cyclase-activating polypeptide (PACAP) acts on cortical precursor cells to trigger glial fibrillary acidic protein (GFAP) gene expression and astrocyte differentiation by stimulation of intracellular cAMP production. Here, we show that as expected, PACAP activates cAMP-dependent protein kinase A. However, inhibition of protein kinase A does not prevent PACAP-induced GFAP gene expression or astrocytogenesis. PACAP also activates the small GTPases Rap1 and Ras, but either activation of Rap1 alone by selective stimulation of the guanine nucleotide exchange factor Epac, or expression of a constitutively active form of Ras, do not induce GFAP gene expression. Ras is activated by PACAP in a cAMP-dependent manner, and inhibition of Ras and/or Rap1 decreases PACAP-induced GFAP promoter stimulation. Thus, cAMP-dependent PACAP-induced GFAP expression during astrocytogenesis involves the coordinated activation of both Ras and Rap1, but activation of either one of them in isolation is not sufficient to trigger this response.
Assuntos
Córtex Cerebral/citologia , Regulação da Expressão Gênica , Proteína Glial Fibrilar Ácida/metabolismo , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase/metabolismo , Células-Tronco/fisiologia , Proteínas rap1 de Ligação ao GTP/metabolismo , Proteínas ras/metabolismo , Animais , Biomarcadores/metabolismo , Diferenciação Celular/fisiologia , Células Cultivadas , Córtex Cerebral/metabolismo , Ativação Enzimática , Proteína Glial Fibrilar Ácida/genética , Fatores de Troca do Nucleotídeo Guanina/genética , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Polipeptídeo Hipofisário Ativador de Adenilato Ciclase/genética , Regiões Promotoras Genéticas , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Wistar , Células-Tronco/citologia , Proteínas rap1 de Ligação ao GTP/genética , Proteínas ras/genéticaRESUMO
OBJECTIVE: The purpose of this study was to investigate if the application of a cervical myofascial induction technique targeted to the ligamentum nuchae resulted in changes in cervical range of motion and pressure pain thresholds (PPT) in asymptomatic subjects. METHODS: Thirty-five subjects, 8 men and 27 women (mean age, 21 +/- 4 years), without a current history of neck, shoulder, or arm pain participated. Participants were randomly divided into 2 groups: the experimental group, which received a real cervical myofascial induction technique, and the control group, which received a sham-manual procedure. Bilateral PPT levels over C5-C6 zygapophyseal joints and tibialis anterior muscles and neck mobility were assessed preintervention and 5 minutes postintervention by an assessor blinded to the treatment allocation of the subject. Separate mixed-model analyses of variance were used to examined the effects of the treatment on neck mobility and PPT levels as the dependent variable, with group (experimental or control) as the between-subjects variable and time (pre-post test) or side (dominant, nondominant) as the within-subjects variable. The hypothesis of interest was the group x time interaction at an a priori alpha level equal to .05. RESULTS: The group x time interaction was statistically significant for cervical flexion (F = 5.4; P = .03), extension (F = 3.3; P = .045), and left lateral-flexion (F = 4.6; P = .04), but not for right lateral-flexion (F = 2.5; P = .1), right rotation (F = 0.5; P = .5), and left rotation (F = 0.09; P = .2). Subjects receiving the real cervical myofascial induction technique experienced greater improvement in cervical mobility when compared with the control group. The group x time interaction did not reveal any significance for PPT in the C5-C6 zygapophyseal joints (F = 0.5; P = .5) and in the tibialis anterior muscle (F = 0.2; P = .8). CONCLUSIONS: The application of a cervical myofascial induction technique resulted in an increase in cervical flexion, extension, and left lateral-flexion, but not rotation motion in a cohort of healthy subjects. No changes in PPT in either C5-C6 zygapophyseal joint (local point) or tibialis anterior muscle (distant point) were found.
Assuntos
Braço/fisiopatologia , Vértebras Cervicais/fisiopatologia , Quiroprática/métodos , Nível de Saúde , Manejo da Dor , Limiar da Dor , Dor/etiologia , Pressão/efeitos adversos , Amplitude de Movimento Articular , Ombro/fisiopatologia , Fáscia , Feminino , Humanos , Masculino , Manipulação da Coluna/métodos , Dor/diagnóstico , Medição da Dor , Índice de Gravidade de Doença , Método Simples-Cego , Adulto JovemRESUMO
The hypothalamus is the principal regulator of global energy balance, enclosing additionally essential neuronal centers for glucose-sensing and osmoregulation. Disturbances in these tightly regulated neuronal networks are thought to underlie the development of severe pandemic syndromes, including obesity and diabetes. In this work, we investigate in vivo the response of individual hypothalamic nuclei to the i.p. administration of glucose or vehicle solutions, using two groups of adult male C57BL6/J fasted mice and a combination of non-invasive T2 ∗-weighted and diffusion-weighted functional magnetic resonance imaging (fMRI) approaches. MRI parameters were assessed in both groups of animals before, during and in a post-stimulus phase, following the administration of glucose or vehicle solutions. Hypothalamic nuclei depicted different patterns of activation characterized by: (i) generalized glucose-induced increases of neuronal activation and perfusion-markers in the lateral hypothalamus, arcuate and dorsomedial nuclei, (ii) cellular shrinking events and decreases in microvascular blood flow in the dorsomedial, ventromedial and lateral hypothalamus, following the administration of vehicle solutions and (iii) increased neuronal activity markers and decreased microperfusion parameters in the ARC nuclei of vehicle-administered animals. Immunohistochemical studies performed after the post-stimulus phase confirmed the presence of c-Fos immunoreactive neurons in the arcuate nucleus (ARC) from both animal groups, with significantly higher numbers in the glucose-treated animals. Together, our results reveal that fMRI methods are able to detect in vivo diverse patterns of glucose or vehicle-induced effects in the different hypothalamic nuclei.
RESUMO
The atrioventricular node (AVN) coordinates the timing of atrial and ventricular contraction to optimize cardiac performance. To study this critical function using mouse genetics, however, new reagents are needed that allow AVN-specific manipulation. Here we describe a novel Gjd3-CreEGFP mouse line that successfully recombines floxed alleles within the AVN beginning at E12.5. These mice have been engineered to express CreEGFP under the control of endogenous Gjd3 regulatory elements without perturbing native protein expression. Detailed histological analysis of Gjd3-CreEGFP mice reveals specific labeling of AVN cardiomyocytes and a subset of cardiac endothelial cells. Importantly, we show that Gjd3-CreEGFP mice have preserved cardiac mechanical and electrical function. In one application of our newly described mouse line, we provide a three-dimensional (3D) view of the AVN using tissue clearing combined with confocal microscopy. With this 3D model as a reference, we identify specific AVN sub-structures based on marker staining characteristics. In addition, we use our Gjd3-CreEGFP mice to guide microdissection of the AVN and construction of a single-cell atlas. Thus, our results establish a new transgenic tool for AVN-specific recombination, provide an updated model of AVN morphology, and describe a roadmap for exploring AVN cellular heterogeneity.