RESUMEN
Neuronal phenotypes are controlled by terminal selector transcription factors in invertebrates, but only a few examples of such regulators have been provided in vertebrates. We hypothesised that TCF7L2 regulates different stages of postmitotic differentiation in the thalamus, and functions as a thalamic terminal selector. To investigate this hypothesis, we used complete and conditional knockouts of Tcf7l2 in mice. The connectivity and clustering of neurons were disrupted in the thalamo-habenular region in Tcf7l2-/- embryos. The expression of subregional thalamic and habenular transcription factors was lost and region-specific cell migration and axon guidance genes were downregulated. In mice with a postnatal Tcf7l2 knockout, the induction of genes that confer thalamic terminal electrophysiological features was impaired. Many of these genes proved to be direct targets of TCF7L2. The role of TCF7L2 in terminal selection was functionally confirmed by impaired firing modes in thalamic neurons in the mutant mice. These data corroborate the existence of master regulators in the vertebrate brain that control stage-specific genetic programmes and regional subroutines, maintain regional transcriptional network during embryonic development, and induce terminal selection postnatally.
Asunto(s)
Diferenciación Celular , Regulación del Desarrollo de la Expresión Génica , Mitosis , Transmisión Sináptica , Tálamo/embriología , Factor de Transcripción 4/metabolismo , Animales , Ratones , Ratones Noqueados , Tálamo/citología , Factor de Transcripción 4/genéticaRESUMEN
Ischemic stroke is a disturbance in cerebral blood flow caused by brain tissue ischemia and hypoxia. We optimized a multifactorial in vitro model of acute ischemic stroke using rat primary neural cultures. This model was exploited to investigate the pro-viable activity of cell-penetrating peptides: arginine-rich Tat(49-57)-NH2 (R49KKRRQRRR57-amide) and its less basic analogue, PTD4 (Y47ARAAARQARA57-amide). Our model included glucose deprivation, oxidative stress, lactic acidosis, and excitotoxicity. Neurotoxicity of these peptides was excluded below a concentration of 50 µm, and PTD4-induced pro-survival was more pronounced. Circular dichroism spectroscopy and molecular dynamics (MD) calculations proved potential contribution of the peptide conformational properties to neuroprotection: in MD, Tat(49-57)-NH2 adopted a random coil and polyproline type II helical structure, whereas PTD4 adopted a helical structure. In an aqueous environment, the peptides mostly adopted a random coil conformation (PTD4) or a polyproline type II helical (Tat(49-57)-NH2) structure. In 30% TFE, PTD4 showed a tendency to adopt a helical structure. Overall, the pro-viable activity of PTD4 was not correlated with the arginine content but rather with the peptide's ability to adopt a helical structure in the membrane-mimicking environment, which enhances its cell membrane permeability. PTD4 may act as a leader sequence in novel drugs for the treatment of acute ischemic stroke.
Asunto(s)
Isquemia Encefálica/prevención & control , Péptidos de Penetración Celular/farmacología , Modelos Animales de Enfermedad , Accidente Cerebrovascular Isquémico/prevención & control , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Animales , Isquemia Encefálica/etiología , Isquemia Encefálica/patología , Permeabilidad de la Membrana Celular , Femenino , Accidente Cerebrovascular Isquémico/etiología , Accidente Cerebrovascular Isquémico/patología , Ratas , Ratas WistarRESUMEN
AIMS/HYPOTHESIS: We aimed to identify microRNAs (miRNAs) under transcriptional control of the HNF1ß transcription factor, and investigate whether its effect manifests in serum. METHODS: The Polish cohort (N = 60) consisted of 11 patients with HNF1B-MODY, 17 with HNF1A-MODY, 13 with GCK-MODY, an HbA1c-matched type 1 diabetic group (n = 9) and ten healthy controls. Replication was performed in 61 clinically-matched British patients mirroring the groups in the Polish cohort. The Polish cohort underwent miRNA serum level profiling with quantitative real-time PCR (qPCR) arrays to identify differentially expressed miRNAs. Validation was performed using qPCR. To determine whether serum content reflects alterations at a cellular level, we quantified miRNA levels in a human hepatocyte cell line (HepG2) with small interfering RNA knockdowns of HNF1α or HNF1ß. RESULTS: Significant differences (adjusted p < 0.05) were noted for 11 miRNAs. Five of them differed between HNF1A-MODY and HNF1B-MODY, and, amongst those, four (miR-24, miR-27b, miR-223 and miR-199a) showed HNF1B-MODY-specific expression levels in the replication group. In all four cases the miRNA expression level was lower in HNF1B-MODY than in all other tested groups. Areas under the receiver operating characteristic curves ranged from 0.79 to 0.86, with sensitivity and specificity reaching 91.7% (miR-24) and 82.1% (miR-199a), respectively. The cellular expression pattern of miRNA was consistent with serum levels, as all were significantly higher in HNF1α- than in HNF1ß-deficient HepG2 cells. CONCLUSIONS/INTERPRETATION: We have shown that expression of specific miRNAs depends on HNF1ß function. The impact of HNF1ß deficiency was evidenced at serum level, making HNF1ß-dependent miRNAs potentially applicable in the diagnosis of HNF1B-MODY.
Asunto(s)
Factor Nuclear 1-alfa del Hepatocito/metabolismo , Factor Nuclear 1-beta del Hepatocito/metabolismo , MicroARNs/sangre , MicroARNs/genética , Regulación de la Expresión Génica/genética , Regulación de la Expresión Génica/fisiología , Hemoglobina Glucada/genética , Hemoglobina Glucada/metabolismo , Células Hep G2 , Factor Nuclear 1-alfa del Hepatocito/genética , Factor Nuclear 1-beta del Hepatocito/genética , Humanos , ARN Interferente Pequeño/genética , Curva ROC , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Autophagy is indispensable for the proper architecture and flawless functioning of pancreatic ß-cells. A growing body of evidence indicates reciprocal communication between autophagic pathways, apoptosis, and intracellular lipids. The way in which elevated levels of free saturated or unsaturated FAs contribute to progressive ß-cell failure remains incompletely understood. Stearoyl-CoA desaturase (SCD)1, a key regulatory enzyme in biosynthesis of MUFAs, was shown to play an important role in regulation of ß-cell function. Here, we investigated whether SCD1 activity is engaged in palmitate-induced pancreatic ß-cell autophagy. We found augmented apoptosis and diminished autophagy upon cotreatment of INS-1E cells with palmitate and an SCD1 inhibitor. Furthermore, we found that additional treatment of the cells with monensin, an inhibitor of autophagy at the step of fusion, exacerbates palmitate-induced apoptosis. Accordingly, diminished SCD1 activity affected the accumulation, composition, and saturation status of cellular membrane phospholipids and neutral lipids. Such an effect was accompanied by aberrant endoplasmic reticulum stress, mitochondrial injury, and decreases in insulin secretion and cell proliferation. Our data reveal a novel mechanism by which the inhibition of SCD1 activity affects autophagosome-lysosome fusion because of perturbations in cellular membrane integrity, thus leading to an aberrant stress response and ß-cell failure.
Asunto(s)
Autofagia/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Células Secretoras de Insulina/efectos de los fármacos , Palmitatos/farmacología , Estearoil-CoA Desaturasa/antagonistas & inhibidores , Animales , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Insulina/farmacología , Células Secretoras de Insulina/citología , Células Secretoras de Insulina/enzimología , Células Secretoras de Insulina/metabolismo , Insulinoma , Lisosomas/metabolismo , Fusión de Membrana/efectos de los fármacos , Ácido Palmítico/farmacología , Fosfolípidos/metabolismo , Ratas , Estearoil-CoA Desaturasa/genética , Estearoil-CoA Desaturasa/metabolismoRESUMEN
Obesity-related type 2 diabetes develops in individuals with the onset of ß-cell dysfunction. Pancreatic islet lipotoxicity is now recognized as a primary reason for the onset and progression of the disease. Such dysfunction is reflected by the aberrant secretory capacity and detrimental loss of ß-cell mass and survival. Elevated circulating serum fatty acid levels and disordered lipid metabolism management are particularly interesting in the search for biologically relevant triggers of ß-cell demise. Herein, we review various types of toxic lipid metabolites that may play a significant role in pancreatic islet failure. The lipotoxic effect on ß-cells depends on the type of lipid mediator (e.g., long-chain fatty acids, diacylglycerols, ceramides, phospholipids), cellular location of its action (e.g., endoplasmic reticulum, mitochondria), and associated-organelle conditions (e.g., membranes, vesicles). We also discuss various aspects of lipid action in ß-cells, including effects on metabolic pathways, stress responses (e.g., oxidative stress, endoplasmic reticulum stress, and autophagy), and gene expression.
Asunto(s)
Diabetes Mellitus Tipo 2/fisiopatología , Islotes Pancreáticos/fisiopatología , Autofagia , Ceramidas/metabolismo , Retículo Endoplásmico/metabolismo , Retículo Endoplásmico/fisiología , Humanos , Mitocondrias/fisiologíaRESUMEN
The Wnt signaling pathway plays an important role in morphogenesis, differentiation, cell survival and proliferation. Wnt activators are secreted proteins that work in an auto-, para- and endocrine manner and their synthesis, secretion and transport are tightly regulated. Frizzled/LRP is the main receptor complex in the canonical Wnt pathway. Its activation triggers ß-catenin translocation to the nucleus and increases activity of TCF transcription factor. Disruption in Wnt signaling has been found in many pathophysiological states such as different types of cancer, neurodegenerative diseases and metabolic disorders. Recent studies revealed the important role of Wnt signaling in maintaining carbohydrate and lipid homeostasis. Activation of the Frizzled/LRP receptor complex leads to increase in the activity of transcription factors and nuclear receptors that regulate expression of genes involved in lipid utilization (PPARδ, RAR, LXR) and inhibits adipogenesis. The Wnt signaling pathway is also involved in the regulation of gluconeogenesis and glycolysis. This review summarizes the current state of knowledge about mechanisms that regulate canonical Wnt signaling and its role in cell metabolism regulation.
Asunto(s)
Fenómenos Fisiológicos Celulares/fisiología , Proteínas Wnt/metabolismo , Vía de Señalización Wnt/fisiología , Adipogénesis/fisiología , Metabolismo de los Hidratos de Carbono/fisiología , Diferenciación Celular/fisiología , Proliferación Celular , Expresión Génica/fisiología , Gluconeogénesis/fisiología , Glucólisis/fisiología , Homeostasis/fisiología , Humanos , Metabolismo de los Lípidos/fisiología , Morfogénesis/fisiología , Receptores Citoplasmáticos y Nucleares/metabolismo , Transducción de Señal/fisiología , Factores de Transcripción TCF/genética , Factores de Transcripción TCF/metabolismo , Factores de Transcripción/metabolismo , beta Catenina/metabolismoRESUMEN
Nuclear magnetic resonance is a "workhorse technique" used in metabolomics, complementary to mass spectrometry. Unfortunately, only the most basic NMR methods are sensitive enough to allow fast medical screening. The most common of them, a simple 1H NMR, suffers from low dispersion of resonance frequencies, which often hampers the identification of metabolites. In this article we show that 1H NMR spectra contain previously overlooked parameters potentially helpful in metabolite identification, namely the rates of temperature-induced changes of chemical shifts. We prove that they are reproducible between various metabolite mixtures and can be determined quickly when Radon transform is used to process the data.
RESUMEN
In the setting of metabolic overload, chronic elevations of free fatty acids in blood and tissues are associated with pancreatic ß-cell lipotoxicity and failure. Ultimately, obesity combined with insulin resistance increases the dysfunctional demand of ß-cells and contributes to the development of type 2 diabetes. Forkhead box O1 (FoxO1) is a potent transcriptional regulator of pancreatic ß-cell function and tolerance to lipid stress. The present study examined the effects of stearoyl-CoA desaturase 1 (SCD1)-metabolized precursors and products, notably oleic acid, on the compensatory capacity of ß-cells and their relationship with regulation of the FoxO1 and Wnt pathways. The trioleate-induced compromise of insulin sensitivity blunted the compensatory response of pancreatic ß-cells in primary rat islets. These events were associated with increases in the nuclear accumulation and transcriptional activity of FoxO1. Such effects were also observed in INS-1E cells that were subjected to oleate treatment. The overexpression of human SCD1 that was accompanied by endogenously generated oleic acid also led to an increase in the nuclear abundance of FoxO1. The mechanism of the oleate-mediated subcellular localization of FoxO1 was independent of the fatty acid receptor GPR40. Instead, the mechanism involved diversion of the active ß-catenin pool from an interaction with transcription factor 7-like 2 toward FoxO1-mediated transcription in ß-cells. Our findings identify a unique role for oleic acid in the compensatory response of pancreatic ß-cells and emphasize the importance of FoxO1 in ß-cell failure in obesity-induced insulin resistance.
Asunto(s)
Proteína Forkhead Box O1/metabolismo , Regulación de la Expresión Génica , Células Secretoras de Insulina/metabolismo , Ácido Oléico/metabolismo , Transporte de Proteínas/fisiología , beta Catenina/metabolismo , Animales , Núcleo Celular , Diabetes Mellitus Tipo 2/metabolismo , Proteínas de Homeodominio , Masculino , Proteínas del Tejido Nervioso/metabolismo , Ratas , Ratas Wistar , Receptores Acoplados a Proteínas G , Estearoil-CoA Desaturasa/metabolismo , Transactivadores , Proteína 2 Similar al Factor de Transcripción 7/metabolismoRESUMEN
Canonical Wnt signaling, which is transduced by ß-catenin and lymphoid enhancer factor 1/T cell-specific transcription factors (LEF1/TCFs), regulates many aspects of metazoan development and tissue renewal. Although much evidence has associated canonical Wnt/ß-catenin signaling with mood disorders, the mechanistic links are still unknown. Many components of the canonical Wnt pathway are involved in cellular processes that are unrelated to classical canonical Wnt signaling, thus further blurring the picture. The present review critically evaluates the involvement of classical Wnt/ß-catenin signaling in developmental processes that putatively underlie the pathology of mental illnesses. Particular attention is given to the roles of LEF1/TCFs, which have been discussed surprisingly rarely in this context. Highlighting recent discoveries, we propose that alterations in the activity of LEF1/TCFs, and particularly of transcription factor 7-like 2 (TCF7L2), result in defects previously associated with neuropsychiatric disorders, including imbalances in neurogenesis and oligodendrogenesis, the functional disruption of thalamocortical circuitry and dysfunction of the habenula.
Asunto(s)
Encéfalo/citología , Encéfalo/crecimiento & desarrollo , Trastornos Mentales/metabolismo , Trastornos Mentales/patología , Proteína 2 Similar al Factor de Transcripción 7/metabolismo , Vía de Señalización Wnt , Animales , Encéfalo/metabolismo , Encéfalo/patología , Humanos , NeurogénesisRESUMEN
The association between mental disorders and diabetes has a long history. Recent large-scale, well-controlled epidemiological studies confirmed a link between diabetes and psychiatric illnesses. The scope of this review is to summarize our current understanding of this relationship from a molecular perspective. We first discuss the potential contribution of diabetes-associated metabolic impairments to the etiology of mental conditions. Then, we focus on possible shared molecular risk factors and mechanisms. Simple comorbidity, shared susceptibility loci, and common pathophysiological processes in diabetes and mental illnesses have changed our traditional way of thinking about mental illness. We conclude that schizophrenia and affective disorders are not limited to an imbalance in dopaminergic and serotoninergic neurotransmission in the brain. They are also systemic disorders that can be considered, to some extent, as metabolic disorders.
Asunto(s)
Encéfalo/metabolismo , Trastornos Mentales/metabolismo , Redes y Vías Metabólicas , Animales , Comorbilidad , Diabetes Mellitus Tipo 2/epidemiología , Diabetes Mellitus Tipo 2/metabolismo , Glucosa/metabolismo , Humanos , Trastornos Mentales/epidemiologíaRESUMEN
Wnt signaling molecules are associated with obesity, hyperlipidemia, and type 2 diabetes (T2D). Here, we show that two Wnt proteins, WNT3a and WNT4, are specifically secreted by skeletal muscle and adipose tissue during the development of insulin resistance and play an important role in cross-talk between insulin-resistant tissues and pancreatic beta cells. The activation of Frizzled receptor and Wnt signaling in pancreatic islets via circulating WNT3a in blood resulted in higher insulin secretion and an increase in beta cell proliferation, thus leading to islet adaptation in a pre-diabetic state. Interestingly, in fully developed T2D, the expression profiles of Wnt3a and Wnt4 in adipose tissue and muscle cells and blood plasma levels of these proteins were opposite to the pre-diabetic state, thus favoring the downregulation of Wnt signaling in beta cells and resulting in dysfunctional pancreatic islets. These results demonstrate that alterations in the secretion profile of a canonical Wnt activator (WNT3a) and inhibitor (WNT4) from insulin-resistant tissues during the development of T2D are responsible for triggering progression from a pre-diabetic to a diabetic state. We also show here that WNT3a and WNT4 are potent myokines, and their expression and secretion are regulated in response to nutritional and metabolic changes.
Asunto(s)
Tejido Adiposo/metabolismo , Islotes Pancreáticos/fisiopatología , Músculo Esquelético/metabolismo , Proteína Wnt3A/metabolismo , Proteína Wnt4/metabolismo , Células 3T3-L1 , Animales , Medios de Cultivo Condicionados , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patología , Dieta Alta en Grasa , Progresión de la Enfermedad , Resistencia a la Insulina , Islotes Pancreáticos/metabolismo , Islotes Pancreáticos/patología , Masculino , Ratones , Ratas , Ratas WistarRESUMEN
Adipocytes are one of the primary sources of inflammatory cytokines that drive the low-grade inflammation associated with obesity and obesity-related diseases. Stearoyl-CoA desaturase, a key adipogenic enzyme in rodents and humans, plays significant role in the regulation of adipocyte inflammation via a mechanism that involves the regulation of inflammatory gene expression. In the present study, we tested the hypothesis that the stearoyl-CoA desaturase 1-related regulation of gene expression might be driven by changes in DNA methylation. We showed that stearoyl-CoA desaturase 1 overexpression causes the global hypomethylation of DNA, even as early as 12h after the induction of differentiation, with the greatest difference seen in mature adipocytes. In contrast, both the silencing of stearoyl-CoA desaturase 1 gene expression by siRNA and inhibition of stearoyl-CoA desaturase 1 activity resulted in DNA hypermethylation in 3T3-L1 adipocytes. The analysis of the promoter methylation of 22 genes that are related to the inflammatory response showed that the level of methylation of CpG sites in interleukin-10 receptor a, interleukin-4 receptor a, interleukin-6 signal transducer, and transforming growth factor ß 1 promoters was strongly related to stearoyl-CoA desaturase 1 expression or activity. The changes in methylation at CpG promoter sites correlated with differential expression of the aforementioned genes. The results show that stearoyl-CoA desaturase 1 regulates the level of DNA methylation in adipocytes and suggest that the mechanism by which stearoyl-CoA desaturase 1 affects adipocyte inflammation may involve changes in the methylation of inflammatory genes.