RESUMO
Interactions between the gut microbiota, diet, and the host potentially contribute to the development of metabolic diseases. Here, we identify imidazole propionate as a microbially produced histidine-derived metabolite that is present at higher concentrations in subjects with versus without type 2 diabetes. We show that imidazole propionate is produced from histidine in a gut simulator at higher concentrations when using fecal microbiota from subjects with versus without type 2 diabetes and that it impairs glucose tolerance when administered to mice. We further show that imidazole propionate impairs insulin signaling at the level of insulin receptor substrate through the activation of p38γ MAPK, which promotes p62 phosphorylation and, subsequently, activation of mechanistic target of rapamycin complex 1 (mTORC1). We also demonstrate increased activation of p62 and mTORC1 in liver from subjects with type 2 diabetes. Our findings indicate that the microbial metabolite imidazole propionate may contribute to the pathogenesis of type 2 diabetes.
Assuntos
Diabetes Mellitus Tipo 2/metabolismo , Microbioma Gastrointestinal , Imidazóis/metabolismo , Insulina/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Transdução de Sinais , Animais , Células Cultivadas , Diabetes Mellitus Tipo 2/microbiologia , Células HEK293 , Histidina/metabolismo , Humanos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína Sequestossoma-1/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
While the commensal microbiota is considered an important regulator of immunity and metabolism, the mechanisms controlling the interplay between diet, cytokine signaling, and the microbiota in atherosclerosis remains unknown. In this issue of Immunity, Fatkhullina et al. (2018) demonstrate that interlukin-23-22 axis regulates diet-induced atherosclerosis by repressing pro-atherogenic microbiota.
Assuntos
Aterosclerose , Microbioma Gastrointestinal , Microbiota , Dieta , Homeostase , Humanos , Interleucina-23 , Interleucinas , Interleucina 22RESUMO
CRISPR-Cas genome-editing nucleases hold substantial promise for developing human therapeutic applications1-6 but identifying unwanted off-target mutations is important for clinical translation7. A well-validated method that can reliably identify off-targets in vivo has not been described to date, which means it is currently unclear whether and how frequently these mutations occur. Here we describe 'verification of in vivo off-targets' (VIVO), a highly sensitive strategy that can robustly identify the genome-wide off-target effects of CRISPR-Cas nucleases in vivo. We use VIVO and a guide RNA deliberately designed to be promiscuous to show that CRISPR-Cas nucleases can induce substantial off-target mutations in mouse livers in vivo. More importantly, we also use VIVO to show that appropriately designed guide RNAs can direct efficient in vivo editing in mouse livers with no detectable off-target mutations. VIVO provides a general strategy for defining and quantifying the off-target effects of gene-editing nucleases in whole organisms, thereby providing a blueprint to foster the development of therapeutic strategies that use in vivo gene editing.
Assuntos
Proteínas Associadas a CRISPR/metabolismo , Sistemas CRISPR-Cas/genética , Edição de Genes/métodos , Edição de Genes/normas , Genoma/genética , Mutação , Especificidade por Substrato/genética , Animais , Proteínas Associadas a CRISPR/genética , Feminino , Humanos , Mutação INDEL , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pró-Proteína Convertase 9/genética , Transgenes/genéticaRESUMO
BACKGROUND: Plasma concentration of low-density lipoprotein (LDL) cholesterol is a well-established risk factor for cardiovascular disease. Inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9), which regulates cholesterol homeostasis, has recently emerged as an approach to reduce cholesterol levels. The development of humanized animal models is an important step to validate and study human drug targets, and use of genome and base editing has been proposed as a mean to target disease alleles. RESULTS: To address the lack of validated models to test the safety and efficacy of techniques to target human PCSK9, we generated a liver-specific human PCSK9 knock-in mouse model (hPCSK9-KI). We showed that plasma concentrations of total cholesterol were higher in hPCSK9-KI than in wildtype mice and increased with age. Treatment with evolocumab, a monoclonal antibody that targets human PCSK9, reduced cholesterol levels in hPCSK9-KI but not in wildtype mice, showing that the hypercholesterolemic phenotype was driven by overexpression of human PCSK9. CRISPR-Cas9-mediated genome editing of human PCSK9 reduced plasma levels of human and not mouse PCSK9, and in parallel reduced plasma concentrations of total cholesterol; genome editing of mouse Pcsk9 did not reduce cholesterol levels. Base editing using a guide RNA that targeted human and mouse PCSK9 reduced plasma levels of human and mouse PCSK9 and total cholesterol. In our mouse model, base editing was more precise than genome editing, and no off-target editing nor chromosomal translocations were identified. CONCLUSIONS: Here, we describe a humanized mouse model with liver-specific expression of human PCSK9 and a human-like hypercholesterolemia phenotype, and demonstrate that this mouse can be used to evaluate antibody and gene editing-based (genome and base editing) therapies to modulate the expression of human PCSK9 and reduce cholesterol levels. We predict that this mouse model will be used in the future to understand the efficacy and safety of novel therapeutic approaches for hypercholesterolemia.
Assuntos
Colesterol/sangue , Hipercolesterolemia/genética , Fígado/metabolismo , Pró-Proteína Convertase 9/genética , Animais , Modelos Animais de Doenças , Edição de Genes , Genoma , Humanos , Hipercolesterolemia/metabolismo , Camundongos , Camundongos TransgênicosRESUMO
Activation of brown adipose tissue (BAT) and browning of white adipose tissue (WAT) present potential new therapies for obesity and type 2 diabetes. Here, we examined the effects of ß3-adrenergic stimulation on tissue-specific uptake and storage of free fatty acids (FFA) and its implications for whole body FFA metabolism in diet-induced obese rats using a multi-radiotracer technique. Male Wistar rats were high fat-fed for 12 wk and administered ß3-agonist CL316,243 (CL, 1 mg·kg-1·day-1) or saline via osmotic minipumps during the last 3 wk. The rats were then fasted and acutely infused with a tracer mixture ([14C]palmitate and the partially metabolized R-[3H]bromopalmitate) under anesthesia. CL infusion decreased body weight gain and fasting plasma glucose levels. While core body temperature was unaffected, infrared thermography showed an increase in tail heat dissipation following CL infusion. Interestingly, CL markedly increased both FFA storage and utilization in interscapular and perirenal BAT, whereas the flux of FFA to skeletal muscle was decreased. In this rat model of obesity, only sporadic populations of beige adipocytes were detected in the epididymal WAT depot of CL-infused rats, and there was no change in FFA uptake or utilization in WAT following CL infusion. In summary, ß3-agonism robustly increased FFA flux to BAT coupled with enhanced utilization. Increased BAT activation most likely drove the increased tail heat dissipation to maintain thermostasis. Our results emphasize the quantitative role of brown fat as the functional target of ß3-agonism in obesity.
Assuntos
Tecido Adiposo Marrom/efeitos dos fármacos , Tecido Adiposo Branco/efeitos dos fármacos , Agonistas de Receptores Adrenérgicos beta 3/farmacologia , Dieta Hiperlipídica , Dioxóis/farmacologia , Ácidos Graxos não Esterificados/metabolismo , Obesidade/metabolismo , Tecido Adiposo Marrom/metabolismo , Tecido Adiposo Marrom/patologia , Tecido Adiposo Branco/metabolismo , Tecido Adiposo Branco/patologia , Animais , Western Blotting , Radioisótopos de Carbono , Imuno-Histoquímica , Masculino , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/metabolismo , Palmitatos/metabolismo , Ratos , Ratos Wistar , Receptores Adrenérgicos beta 3 , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Termografia , Trítio , Proteína Desacopladora 1/efeitos dos fármacos , Proteína Desacopladora 1/metabolismoRESUMO
Sleep disorders are highly prevalent during late pregnancy and can impose adverse effects, such as preeclampsia and diabetes. However, the consequences of sleep fragmentation (SF) on offspring metabolism and epigenomic signatures are unclear. We report that physical activity during early life, but not later, reversed the increased body weight, altered glucose and lipid homeostasis, and increased visceral adipose tissue in offspring of mice subjected to gestational SF (SFo). The reversibility of this phenotype may reflect epigenetic mechanisms induced by SF during gestation. Accordingly, we found that the metabolic master switch Foxo1 was epigenetically misregulated in SFo livers in a temporally regulated fashion. Temporal Foxo1 analysis and its gluconeogenetic targets revealed that the epigenetic abnormalities of Foxo1 precede the metabolic syndrome phenotype. Importantly, regular physical activity early, but not later in life, reversed Foxo1 epigenetic misregulation and altered the metabolic phenotype in gestationally SF-exposed offspring. Thus, we have identified a restricted postnatal period during which lifestyle interventions may reverse the Foxo1 epigenetically mediated risk for metabolic dysfunction later in the life, as induced by gestational sleep disorders.
Assuntos
Metabolismo Energético/genética , Epigênese Genética , Fatores de Transcrição Forkhead/genética , Fígado/metabolismo , Atividade Motora , Efeitos Tardios da Exposição Pré-Natal , Transtornos do Sono-Vigília/genética , Sono , Adiposidade , Fatores Etários , Animais , Biomarcadores/sangue , Glicemia/metabolismo , Feminino , Proteína Forkhead Box O1 , Fatores de Transcrição Forkhead/metabolismo , Regulação da Expressão Gênica , Idade Gestacional , Gordura Intra-Abdominal/metabolismo , Gordura Intra-Abdominal/fisiopatologia , Estilo de Vida , Lipídeos/sangue , Fígado/fisiopatologia , Camundongos Endogâmicos C57BL , Gravidez , Fatores de Risco , Comportamento de Redução do Risco , Transtornos do Sono-Vigília/sangue , Transtornos do Sono-Vigília/fisiopatologia , Aumento de PesoRESUMO
RATIONALE: An increased cancer aggressiveness and mortality have been recently reported among patients with obstructive sleep apnea (OSA). Intermittent hypoxia (IH), a hallmark of OSA, enhances melanoma growth and metastasis in mice. OBJECTIVES: To assess whether OSA-related adverse cancer outcomes occur via IH-induced changes in host immune responses, namely tumor-associated macrophages (TAMs). MEASUREMENTS AND MAIN RESULTS: Lung epithelial TC1 cell tumors were 84% greater in mice subjected to IH for 28 days compared with room air (RA). In addition, TAMs in IH-exposed tumors exhibited reductions in M1 polarity with a shift toward M2 protumoral phenotype. Although TAMs from tumors harvested from RA-exposed mice increased TC1 migration and extravasation, TAMs from IH-exposed mice markedly enhanced such effects and also promoted proliferative rates and invasiveness of TC1 cells. Proliferative rates of melanoma (B16F10) and TC1 cells exposed to IH either in single culture or in coculture with macrophages (RAW 264.7) increased only when RAW 264.7 macrophages were concurrently present. CONCLUSIONS: Our findings support the notion that IH-induced alterations in TAMs participate in the adverse cancer outcomes reported in OSA.
Assuntos
Hipóxia/imunologia , Neoplasias Pulmonares/patologia , Macrófagos/patologia , Melanoma Experimental/patologia , Apneia Obstrutiva do Sono/fisiopatologia , Animais , Linhagem Celular Tumoral , Proliferação de Células , Modelos Animais de Doenças , Citometria de Fluxo , Hipóxia/etiologia , Neoplasias Pulmonares/imunologia , Masculino , Melanoma Experimental/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Invasividade Neoplásica , Fenótipo , Apneia Obstrutiva do Sono/imunologia , Linfócitos T Citotóxicos/patologiaRESUMO
GFRAL-expressing neurons actuate aversion and nausea, are targets for obesity treatment, and may mediate metformin effects by long-term GDF15-GFRAL agonism. Whether GFRAL+ neurons acutely regulate glucose and energy homeostasis is, however, underexplored. Here, we report that cell-specific activation of GFRAL+ neurons using a variety of techniques causes a torpor-like state, including hypothermia, the release of stress hormones, a shift from glucose to lipid oxidation, and impaired insulin sensitivity, glucose tolerance, and skeletal muscle glucose uptake but augmented glucose uptake in visceral fat. Metabolomic analysis of blood and transcriptomics of muscle and fat indicate alterations in ketogenesis, insulin signaling, adipose tissue differentiation and mitogenesis, and energy fluxes. Our findings indicate that acute GFRAL+ neuron activation induces endocrine and gluco- and thermoregulatory responses associated with nausea and torpor. While chronic activation of GFRAL signaling promotes weight loss in obesity, these results show that acute activation of GFRAL+ neurons causes hypothermia and hyperglycemia.
Assuntos
Glucose , Hipotermia , Náusea , Neurônios , Torpor , Animais , Neurônios/metabolismo , Náusea/metabolismo , Hipotermia/metabolismo , Torpor/fisiologia , Glucose/metabolismo , Camundongos , Masculino , Músculo Esquelético/metabolismo , Camundongos Endogâmicos C57BL , Insulina/metabolismo , Resistência à Insulina , Transdução de SinaisRESUMO
BACKGROUND: Sleepiness and cognitive dysfunction are recognized as prominent consequences of sleep deprivation. Experimentally induced short-term sleep fragmentation, even in the absence of any reductions in total sleep duration, will lead to the emergence of excessive daytime sleepiness and cognitive impairments in humans. Tumor necrosis factor (TNF)-α has important regulatory effects on sleep, and seems to play a role in the occurrence of excessive daytime sleepiness in children who have disrupted sleep as a result of obstructive sleep apnea, a condition associated with prominent sleep fragmentation. The aim of this study was to examine role of the TNF-α pathway after long-term sleep fragmentation in mice. METHODS: The effect of chronic sleep fragmentation during the sleep-predominant period on sleep architecture, sleep latency, cognitive function, behavior, and inflammatory markers was assessed in C57BL/6 J and in mice lacking the TNF-α receptor (double knockout mice). In addition, we also assessed the above parameters in C57BL/6 J mice after injection of a TNF-α neutralizing antibody. RESULTS: Mice subjected to chronic sleep fragmentation had preserved sleep duration, sleep state distribution, and cumulative delta frequency power, but also exhibited excessive sleepiness, altered cognitive abilities and mood correlates, reduced cyclic AMP response element-binding protein phosphorylation and transcriptional activity, and increased phosphodiesterase-4 expression, in the absence of AMP kinase-α phosphorylation and ATP changes. Selective increases in cortical expression of TNF-α primarily circumscribed to neurons emerged. Consequently, sleepiness and cognitive dysfunction were absent in TNF-α double receptor knockout mice subjected to sleep fragmentation, and similarly, treatment with a TNF-α neutralizing antibody abrogated sleep fragmentation-induced learning deficits and increases in sleep propensity. CONCLUSIONS: Taken together, our findings show that recurrent arousals during sleep, as happens during sleep apnea, induce excessive sleepiness via activation of inflammatory mechanisms, and more specifically TNF-α-dependent pathways, despite preserved sleep duration.
Assuntos
Transtornos Cognitivos/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral/deficiência , Transdução de Sinais/fisiologia , Privação do Sono/metabolismo , Fases do Sono/fisiologia , Fator de Necrose Tumoral alfa/metabolismo , Animais , Anticorpos Neutralizantes/farmacologia , Nível de Alerta/genética , Nível de Alerta/fisiologia , Encéfalo/metabolismo , Encéfalo/fisiologia , Transtornos Cognitivos/genética , Transtornos Cognitivos/psicologia , Masculino , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Vias Neurais/fisiologia , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Recidiva , Transdução de Sinais/genética , Privação do Sono/genética , Privação do Sono/psicologia , Fases do Sono/genética , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/genéticaRESUMO
Intermittent hypoxia (IH) is a frequent occurrence in sleep and respiratory disorders. Both human and murine studies show that IH may be implicated in metabolic dysfunction. Although the effects of nocturnal low-frequency intermittent hypoxia (IH(L)) have not been extensively examined, it would appear that IH(L) and high-frequency intermittent hypoxia (IH(H)) may elicit distinct metabolic adaptations. To this effect, C57BL/6J mice were randomly assigned to IH(H) (cycles of 90 s 6.4% O(2) and 90 s 21% O(2) during daylight), IH(L) (8% O(2) during daylight hours), or control (CTL) for 5 wk. At the end of exposures, some of the mice were subjected to a glucose tolerance test (GTT; after intraperitoneal injection of 2 mg glucose/g body wt), and others were subjected to an insulin tolerance test (ITT; 0.25 units Humulin/kg body wt), with plasma leptin and insulin levels being measured in fasting conditions. Skeletal muscles were harvested for GLUT4 and proliferator-activated receptor gamma coactivator 1-α (PGC1-α) expression. Both IH(H) and IH(L) displayed reduced body weight increases compared with CTL. CTL mice had higher basal glycemic levels, but GTT kinetics revealed marked differences between IH(L) and IH(H), with IH(L) manifesting the lowest insulin sensitivity compared with either IH(H) or CTL, and such findings were further confirmed by ITT. No differences emerged in PGC1-α expression across the three experimental groups. However, while cytosolic GLUT4 protein expression remained similar in IH(L), IH(H), and CTL, significant decreases in GLUT4 membrane fraction occurred in hypoxia and were most pronounced in IH(L)-exposed mice. Thus IH(H) and IH(L) elicit differential glucose homeostatic responses despite similar cumulative hypoxic profiles.
Assuntos
Adaptação Fisiológica/fisiologia , Ritmo Circadiano/fisiologia , Hipóxia/metabolismo , Descanso/fisiologia , Animais , Peso Corporal/fisiologia , Transportador de Glucose Tipo 4/metabolismo , Insulina/sangue , Leptina/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais , Músculo Esquelético/metabolismo , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo , Fatores de Tempo , Transativadores/metabolismo , Fatores de TranscriçãoRESUMO
BACKGROUND: The aim was to test the hypothesis that the blood serum of rats subjected to recurrent airway obstructions mimicking obstructive sleep apnea (OSA) induces early activation of bone marrow-derived mesenchymal stem cells (MSC) and enhancement of endothelial wound healing. METHODS: We studied 30 control rats and 30 rats subjected to recurrent obstructive apneas (60 per hour, lasting 15 s each, for 5 h). The migration induced in MSC by apneic serum was measured by transwell assays. MSC-endothelial adhesion induced by apneic serum was assessed by incubating fluorescent-labelled MSC on monolayers of cultured endothelial cells from rat aorta. A wound healing assay was used to investigate the effect of apneic serum on endothelial repair. RESULTS: Apneic serum showed significant increase in chemotaxis in MSC when compared with control serum: the normalized chemotaxis indices were 2.20 +/- 0.58 (m +/- SE) and 1.00 +/- 0.26, respectively (p < 0.05). MSC adhesion to endothelial cells was greater (1.75 +/- 0.14 -fold; p < 0.01) in apneic serum than in control serum. When compared with control serum, apneic serum significantly increased endothelial wound healing (2.01 +/- 0.24 -fold; p < 0.05). CONCLUSIONS: The early increases induced by recurrent obstructive apneas in MSC migration, adhesion and endothelial repair suggest that these mechanisms play a role in the physiological response to the challenges associated to OSA.
Assuntos
Células Endoteliais/metabolismo , Células-Tronco Mesenquimais/metabolismo , Apneia Obstrutiva do Sono/sangue , Cicatrização , Animais , Adesão Celular , Células Cultivadas , Quimiotaxia , Técnicas de Cocultura , Modelos Animais de Doenças , Masculino , Ratos , Ratos Endogâmicos Lew , Ratos Sprague-Dawley , Fatores de TempoRESUMO
OBJECTIVE: Gut-derived inflammatory factors can impair glucose homeostasis, but the underlying mechanisms are not fully understood. In this study, we investigated how hepatic gene expression is regulated by gut colonization status through myeloid differentiation primary response 88 (MYD88) and how one of the regulated genes, lipopolysaccharide-binding protein (Lbp), affects insulin signaling and systemic glucose homeostasis. METHODS: Liver transcriptomics analysis was conducted on four groups of mice fed a chow diet: conventionally raised (CONV-R) wild-type, germ-free (GF) wild-type, CONV-R Myd88 KO, and GF Myd88 KO. Primary hepatocytes were exposed to combinations of lipopolysaccharide (LPS), LBP, and the LBP-blocking peptide LBPK95A, and the effect on insulin signaling was determined. To assess how LBP affects glucose metabolism in vivo, two mouse models were applied: treatment with LBPK95A and hepatic knockdown of Lbp using CRISPR-CAS9. RESULTS: We showed that the colonization status regulates gene expression in the liver and that a subset of these genes, including Lbp, is regulated through MYD88. Furthermore, we demonstrated that LBP impairs insulin signaling in hepatocytes in the presence of low levels of LPS and that the effect of LBP is abolished by LBPK95A. We showed that both systemic pharmacological blocking of LBP by LBPK95A and CRISPR-CAS9-mediated downregulation of hepatic Lbp improve glucose homeostasis. CONCLUSIONS: Our results demonstrate that the gut microbiota regulates hepatic expression of Lbp through MYD88-dependent signaling. LBP potentiates LPS inhibition of insulin signaling in vitro and impairs systemic glucose homeostasis in vivo.
Assuntos
Proteínas de Fase Aguda/metabolismo , Proteínas de Transporte/metabolismo , Glucose/metabolismo , Glicoproteínas de Membrana/metabolismo , Fator 88 de Diferenciação Mieloide/metabolismo , Proteínas de Fase Aguda/genética , Animais , Metabolismo dos Carboidratos/fisiologia , Proteínas de Transporte/genética , Microbioma Gastrointestinal/genética , Microbioma Gastrointestinal/fisiologia , Expressão Gênica , Teste de Tolerância a Glucose , Hepatócitos/metabolismo , Inflamação/metabolismo , Lipopolissacarídeos/metabolismo , Fígado/metabolismo , Fígado/patologia , Masculino , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Fator 88 de Diferenciação Mieloide/farmacologia , Fator 88 de Diferenciação Mieloide/fisiologia , Obesidade/metabolismo , Obesidade/fisiopatologia , Transdução de SinaisRESUMO
STUDY OBJECTIVES: To investigate whether noninvasive application of recurrent airway obstructions induces early release of mesenchymal stem cells into the circulating blood in a rat model of obstructive sleep apnea. DESIGN: Prospective controlled animal study. SETTING: University laboratory. PATIENTS OR PARTICIPANTS: Twenty male Sprague-Dawley rats (250-300 g). INTERVENTIONS: A specially designed nasal mask was applied to the anesthetized rats. Ten rats were subjected to a pattern of recurrent obstructive apneas (60 per hour, lasting 15 seconds each) for 5 hours. Ten anesthetized rats were used as controls. MEASUREMENTS AND RESULTS: Mesenchymal stem cells from the blood and bone marrow samples were isolated and cultured to count the total number of colony-forming unit fibroblasts (CFU-F) of adherent cells after 9 days in culture. The number of CFU-F from circulating blood was significantly (P = 0.02) higher in the rats subjected to recurrent obstructive apneas (5.00 +/- 1.16; mean +/- SEM) than in controls (1.70 +/- 0.72). No significant (P = 0.54) differences were observed in CFU-F from bone marrow. CONCLUSIONS: Application of a pattern of airway obstructions similar to those experienced by patients with sleep apnea induced an early mobilization of mesenchymal stem cells into circulating blood.
Assuntos
Contagem de Células Sanguíneas , Células da Medula Óssea/citologia , Células-Tronco Mesenquimais/citologia , Apneia Obstrutiva do Sono/sangue , Adipócitos/citologia , Animais , Adesão Celular/fisiologia , Contagem de Células , Diferenciação Celular/fisiologia , Divisão Celular/fisiologia , Modelos Animais de Doenças , Fibroblastos/citologia , Masculino , Osteócitos/citologia , Ratos , Ratos Sprague-Dawley , Células-Tronco/citologiaRESUMO
Liver X receptors limit cellular lipid uptake by stimulating the transcription of Inducible Degrader of the LDL Receptor (IDOL), an E3 ubiquitin ligase that targets lipoprotein receptors for degradation. The function of IDOL in systemic metabolism is incompletely understood. Here we show that loss of IDOL in mice protects against the development of diet-induced obesity and metabolic dysfunction by altering food intake and thermogenesis. Unexpectedly, analysis of tissue-specific knockout mice revealed that IDOL affects energy balance, not through its actions in peripheral metabolic tissues (liver, adipose, endothelium, intestine, skeletal muscle), but by controlling lipoprotein receptor abundance in neurons. Single-cell RNA sequencing of the hypothalamus demonstrated that IDOL deletion altered gene expression linked to control of metabolism. Finally, we identify VLDLR rather than LDLR as the primary mediator of IDOL effects on energy balance. These studies identify a role for the neuronal IDOL-VLDLR pathway in metabolic homeostasis and diet-induced obesity.
Assuntos
Metabolismo Energético/fisiologia , Neurônios/metabolismo , Receptores de LDL/metabolismo , Ubiquitina-Proteína Ligases/fisiologia , Animais , Glicemia/metabolismo , Dieta , Metabolismo Energético/genética , Hipotálamo/metabolismo , Resistência à Insulina , Camundongos , Camundongos Knockout , Obesidade/metabolismo , Obesidade/prevenção & controle , Ubiquitina-Proteína Ligases/genéticaRESUMO
Chronic sleep fragmentation (SF) commonly occurs in human populations, and although it does not involve circadian shifts or sleep deprivation, it markedly alters feeding behaviors ultimately promoting obesity and insulin resistance. These symptoms are known to be related to the host gut microbiota. Mice were exposed to SF for 4 weeks and then allowed to recover for 2 weeks. Taxonomic profiles of fecal microbiota were obtained prospectively, and conventionalization experiments were performed in germ-free mice. Adipose tissue insulin sensitivity and inflammation, as well as circulating measures of inflammation, were assayed. Effect of fecal water on colonic epithelial permeability was also examined. Chronic SF-induced increased food intake and reversible gut microbiota changes characterized by the preferential growth of highly fermentative members of Lachnospiraceae and Ruminococcaceae and a decrease of Lactobacillaceae families. These lead to systemic and visceral white adipose tissue inflammation in addition to altered insulin sensitivity in mice, most likely via enhanced colonic epithelium barrier disruption. Conventionalization of germ-free mice with SF-derived microbiota confirmed these findings. Thus, SF-induced metabolic alterations may be mediated, in part, by concurrent changes in gut microbiota, thereby opening the way for gut microbiome-targeted therapeutics aimed at reducing the major end-organ morbidities of chronic SF.
Assuntos
Tecido Adiposo/metabolismo , Microbioma Gastrointestinal , Resistência à Insulina , Privação do Sono/microbiologia , Animais , Insulina/sangue , Interleucinas/sangue , Lactobacillaceae/isolamento & purificação , Leptina/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Privação do Sono/sangue , Privação do Sono/metabolismoRESUMO
BACKGROUND: Excessive sleep fragmentation (SF) is common in pregnant women. Adult-onset metabolic disorders may begin during early development and exhibit substantial sex dimorphism. We hypothesized that metabolic dysfunction induced by gestational SF in male mice would not be apparent in female littermates. METHODS: Body weight and food consumption were measured weekly in male and female offspring after late gestational SF or control sleep (SC). At 20 weeks, plasma leptin, adiponectin, lipid profiles, and insulin and glucose tolerance tests were assessed. Leptin and adiponectin, M1, and M2 macrophage messenger RNA expression and polarity were examined. Adiponectin gene promoter methylation levels in several tissues were assessed. RESULTS: Food intake, body weight, visceral fat mass, and insulin resistance were higher, and adiponectin levels lower in male but not female offspring exposed to gestational SF. However, dyslipidemia was apparent in both male and female offspring exposed to SF, albeit of lesser magnitude. In visceral fat, leptin messenger RNA expression was selectively increased and adiponectin expression was decreased in male offspring exposed to gestational SF, but adiponectin was increased in exposed female offspring. Differences in adipokine expression also emerged in liver, subcutaneous fat, and muscle. Increased M1 macrophage markers were present in male offspring exposed to SF (SFOM) while increased M2 markers emerged in SF in female offspring (SFOF). Similarly, significant differences emerged in the methylation patterns of adiponectin promoter in SFOM and SFOF. CONCLUSION: Gestational sleep fragmentation increases the susceptibility to obesity and metabolic syndrome in male but not in female offspring, most likely via epigenetic changes. Thus, sleep perturbations impose long-term detrimental effects to the fetus manifesting as sex dimorphic metabolic dysfunction in adulthood.
Assuntos
Suscetibilidade a Doenças , Complicações na Gravidez/fisiopatologia , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Caracteres Sexuais , Privação do Sono/fisiopatologia , Adiponectina/sangue , Adiponectina/genética , Animais , Peso Corporal , Metilação de DNA , Ingestão de Alimentos , Epigênese Genética , Feminino , Teste de Tolerância a Glucose , Insulina/sangue , Resistência à Insulina , Gordura Intra-Abdominal/metabolismo , Leptina/sangue , Lipídeos/sangue , Fígado/metabolismo , Macrófagos/metabolismo , Masculino , Síndrome Metabólica/metabolismo , Camundongos , Músculos/metabolismo , Obesidade/metabolismo , Gravidez , Regiões Promotoras Genéticas/genética , Sono/genética , Gordura Subcutânea/metabolismoRESUMO
The molecular mechanisms underlying how sleep fragmentation (SF) influences cancer growth and progression remain largely elusive. Here, we present evidence that SF reduced ROS production by downregulating gp91phox expression and activity in TC1 cell tumor associated macrophages (TAMs), while genetic ablation of phagocytic Nox2 activity increased tumor cell proliferation, motility, invasion, and extravasation in vitro. Importantly, the in vivo studies using immunocompetent syngeneic murine tumor models suggested that Nox2 deficiency mimics SF-induced TAMs infiltration and subsequent tumor growth and invasion. Taken together, these studies reveal that perturbed sleep could adversely affect innate immunity within the tumor by altering Nox2 expression and activity, and indicate that selective potentiation of Nox2 activity may present a novel therapeutic strategy in the treatment of cancer.
RESUMO
Chronic intermittent hypoxia during sleep (IH), as occurs in sleep apnea, promotes systemic insulin resistance. Resveratrol (Resv) has been reported to ameliorate high-fat diet-induced obesity, inflammation, and insulin resistance. To examine the effect of Resv on IH-induced metabolic dysfunction, male mice were subjected to IH or room air conditions for 8 weeks and treated with either Resv or vehicle (Veh). Fasting plasma levels of glucose, insulin, and leptin were obtained, homeostatic model assessment of insulin resistance index levels were calculated, and insulin sensitivity tests (phosphorylated AKT [also known as protein kinase B]/total AKT) were performed in 2 visceral white adipose tissue (VWAT) depots (epididymal [Epi] and mesenteric [Mes]) along with flow cytometry assessments for VWAT macrophages and phenotypes (M1 and M2). IH-Veh and IH-Resv mice showed initial reductions in food intake with later recovery, with resultant lower body weights after 8 weeks but with IH-Resv showing better increases in body weight vs IH-Veh. IH-Veh and IH-Resv mice exhibited lower fasting glucose levels, but only IH-Veh had increased homeostatic model assessment of insulin resistance index vs all 3 other groups. Leptin levels were preserved in IH-Veh but were significantly lower in IH-Resv. Reduced VWAT phosphorylated-AKT/AKT responses to insulin emerged in both Mes and Epi in IH-Veh but normalized in IH-Resv. Increases total macrophage counts and in M1 to M2 ratios occurred in IH-Veh Mes and Epi compared all other 3 groups. Thus, Resv ameliorates food intake and weight gain during IH exposures and markedly attenuates VWAT inflammation and insulin resistance, thereby providing a potentially useful adjunctive therapy for metabolic morbidity in the context of sleep apnea.
Assuntos
Fármacos Antiobesidade/farmacologia , Hipóxia/imunologia , Resistência à Insulina , Gordura Intra-Abdominal/imunologia , Macrófagos/efeitos dos fármacos , Estilbenos/farmacologia , Animais , Avaliação Pré-Clínica de Medicamentos , Ingestão de Alimentos , Insulina/sangue , Leptina/sangue , Masculino , Camundongos Endogâmicos C57BL , Distribuição Aleatória , Resveratrol , Aumento de PesoRESUMO
BACKGROUND: Sleep fragmentation (SF) is highly prevalent and may constitute an important contributing factor to excessive weight gain and the metabolic syndrome. Increased endoplasmic reticulum (ER) stress and activation of the unfolded protein response (UPR) leading to the attenuation of leptin receptor signaling in the hypothalamus leads to obesity and metabolic dysfunction. METHODS: Mice were exposed to SF and sleep control (SC) for varying periods of time during which ingestive behaviors were monitored. UPR pathways and leptin receptor signaling were assessed in hypothalami. To further examine the mechanistic role of ER stress, changes in leptin receptor (ObR) signaling were also examined in wild-type mice treated with the ER chaperone tauroursodeoxycholic acid (TUDCA), as well as in CHOP-/+ transgenic mice. RESULTS: Fragmented sleep in male mice induced increased food intake starting day 3 and thereafter, which was preceded by increases in ER stress and activation of all three UPR pathways in the hypothalamus. Although ObR expression was unchanged, signal transducer and activator of transcription 3 (STAT3) phosphorylation was decreased, suggesting reduced ObR signaling. Unchanged suppressor of cytokine signaling-3 (SOCS3) expression and increases in protein-tyrosine phosphatase 1B (PTP1B) expression and activity emerged with SF, along with reduced p-STAT3 responses to exogenous leptin. SF-induced effects were reversed following TUDCA treatment and were absent in CHOP -/+ mice. CONCLUSIONS: SF induces hyperphagic behaviors and reduced leptin signaling in hypothalamus that are mediated by activation of ER stress, and ultimately lead to increased PTP1B activity. ER stress pathways are therefore potentially implicated in SF-induced weight gain and metabolic dysfunction, and may represent a viable therapeutic target.
Assuntos
Estresse do Retículo Endoplasmático , Hipotálamo/metabolismo , Leptina/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 1/metabolismo , Privação do Sono/fisiopatologia , Animais , Ingestão de Alimentos , Heterozigoto , Hiperfagia/etiologia , Hiperfagia/fisiopatologia , Hipotálamo/efeitos dos fármacos , Hipotálamo/patologia , Leptina/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Obesidade/metabolismo , Receptores para Leptina/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/efeitos dos fármacos , Sono/efeitos dos fármacos , Privação do Sono/complicações , Privação do Sono/metabolismo , Ácido Tauroquenodesoxicólico/farmacologia , Fator de Transcrição CHOP/deficiência , Fator de Transcrição CHOP/genética , Resposta a Proteínas não DobradasRESUMO
Intermittent hypoxia (IH)-induces alterations in tumor-associated macrophages (TAMs) that are associated with adverse cancer outcomes, as reported in patients suffering from sleep apnea. Adipose tissues (AT) and bone-marrow (BM)-derived cells are the inferred sources of macrophages infiltrating malignant tumors. Here, the sources of TAMs and the phenotypic changes induced by IH in the ipsilateral and contralateral AT were investigated by using a syngeneic murine solid tumor model (TC1). C57/B6 male mice were exposed to either IH or room air (RA) for 6 weeks, with TC1 cells being inoculated in the 2nd week. Macrophage content, phenotype and tissue origin were assessed in tumors, and ipsilateral and contralateral AT. IH induced a ~2.2-fold increase in TAM tumor infiltration. However, differential responses in the tumor ipsilateral and contralateral AT emerged: IH increased infiltration of preferentially M1 macrophages in contralateral AT, while reductions in macrophages emerged in ipsilateral AT and primarily consisted of the M2 phenotype. These changes were accompanied by reciprocal increases in resident and BM-derived TAMs in the tumor. IH-induced phenotypic alterations in AT macrophages surrounding the tumor and their increased infiltration within the tumor may contribute to the accelerated tumor progression associated with IH.