RESUMEN
Behavioral state is known to influence interactions between thalamus and cortex, which are important for sensation, action, and cognition. The thalamic reticular nucleus (TRN) is hypothesized to regulate thalamo-cortical interactions, but the underlying functional architecture of this process and its state dependence are unknown. By combining the first TRN ensemble recording with psychophysics and connectivity-based optogenetic tagging, we found reticular circuits to be composed of distinct subnetworks. While activity of limbic-projecting TRN neurons positively correlates with arousal, sensory-projecting neurons participate in spindles and show elevated synchrony by slow waves during sleep. Sensory-projecting neurons are suppressed by attentional states, demonstrating that their gating of thalamo-cortical interactions is matched to behavioral state. Bidirectional manipulation of attentional performance was achieved through subnetwork-specific optogenetic stimulation. Together, our findings provide evidence for differential inhibition of thalamic nuclei across brain states, where the TRN separately controls external sensory and internal limbic processing facilitating normal cognitive function. PAPERFLICK:
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Cognición , Núcleos Talámicos/fisiología , Animales , Atención , Conducta Animal , Sistema Límbico/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Percepción VisualRESUMEN
Interactions between the mediodorsal thalamus and the prefrontal cortex are critical for cognition. Studies in humans indicate that these interactions may resolve uncertainty in decision-making1, but the precise mechanisms are unknown. Here we identify two distinct mediodorsal projections to the prefrontal cortex that have complementary mechanistic roles in decision-making under uncertainty. Specifically, we found that a dopamine receptor (D2)-expressing projection amplifies prefrontal signals when task inputs are sparse and a kainate receptor (GRIK4) expressing-projection suppresses prefrontal noise when task inputs are dense but conflicting. Collectively, our data suggest that there are distinct brain mechanisms for handling uncertainty due to low signals versus uncertainty due to high noise, and provide a mechanistic entry point for correcting decision-making abnormalities in disorders that have a prominent prefrontal component2-6.
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Vías Nerviosas , Corteza Prefrontal/citología , Corteza Prefrontal/fisiología , Tálamo/citología , Tálamo/fisiología , Animales , Toma de Decisiones , Femenino , Humanos , Interneuronas/fisiología , Masculino , Núcleo Talámico Mediodorsal/citología , Núcleo Talámico Mediodorsal/fisiología , Ratones , Receptores Dopaminérgicos/metabolismo , Receptores de Ácido Kaínico/metabolismo , IncertidumbreRESUMEN
The thalamus has long been suspected to have an important role in cognition, yet recent theories have favored a more corticocentric view. According to this view, the thalamus is an excitatory feedforward relay to or between cortical regions, and cognitively relevant computations are exclusively cortical. Here, we review anatomical, physiological, and behavioral studies along evolutionary and theoretical dimensions, arguing for essential and unique thalamic computations in cognition. Considering their architectural features as well as their ability to initiate, sustain, and switch cortical activity, thalamic circuits appear uniquely suited for computing contextual signals that rapidly reconfigure task-relevant cortical representations. We introduce a framework that formalizes this notion, show its consistency with several findings, and discuss its prediction of thalamic roles in perceptual inference and behavioral flexibility. Overall, our framework emphasizes an expanded view of the thalamus in cognitive computations and provides a roadmap to test several of its theoretical and experimental predictions.
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Corteza Cerebral/fisiología , Cognición/fisiología , Modelos Neurológicos , Vías Nerviosas/fisiología , Tálamo/fisiología , Animales , Corteza Cerebral/anatomía & histología , Simulación por Computador , Humanos , Vías Nerviosas/anatomía & histología , Tálamo/anatomía & histologíaRESUMEN
A subset of children with autism spectrum disorder appear to show an improvement in their behavioural symptoms during the course of a fever, a sign of systemic inflammation1,2. Here we elucidate the molecular and neural mechanisms that underlie the beneficial effects of inflammation on social behaviour deficits in mice. We compared an environmental model of neurodevelopmental disorders in which mice were exposed to maternal immune activation (MIA) during embryogenesis3,4 with mouse models that are genetically deficient for contactin-associated protein-like 2 (Cntnap2)5, fragile X mental retardation-1 (Fmr1)6 or Sh3 and multiple ankyrin repeat domains 3 (Shank3)7. We establish that the social behaviour deficits in offspring exposed to MIA can be temporarily rescued by the inflammatory response elicited by the administration of lipopolysaccharide (LPS). This behavioural rescue was accompanied by a reduction in neuronal activity in the primary somatosensory cortex dysgranular zone (S1DZ), the hyperactivity of which was previously implicated in the manifestation of behavioural phenotypes associated with offspring exposed to MIA8. By contrast, we did not observe an LPS-induced rescue of social deficits in the monogenic models. We demonstrate that the differences in responsiveness to the LPS treatment between the MIA and the monogenic models emerge from differences in the levels of cytokine production. LPS treatment in monogenic mutant mice did not induce amounts of interleukin-17a (IL-17a) comparable to those induced in MIA offspring; bypassing this difference by directly delivering IL-17a into S1DZ was sufficient to promote sociability in monogenic mutant mice as well as in MIA offspring. Conversely, abrogating the expression of IL-17 receptor subunit a (IL-17Ra) in the neurons of the S1DZ eliminated the ability of LPS to reverse the sociability phenotypes in MIA offspring. Our data support a neuroimmune mechanism that underlies neurodevelopmental disorders in which the production of IL-17a during inflammation can ameliorate the expression of social behaviour deficits by directly affecting neuronal activity in the central nervous system.
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Interleucina-17/inmunología , Trastornos del Neurodesarrollo/inmunología , Animales , Conducta Animal , Modelos Animales de Enfermedad , Femenino , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil , Lipopolisacáridos/farmacología , Ratones , Ratones Endogámicos C57BL , Embarazo , Efectos Tardíos de la Exposición Prenatal , Conducta SocialRESUMEN
The thalamic reticular nucleus (TRN), the major source of thalamic inhibition, regulates thalamocortical interactions that are critical for sensory processing, attention and cognition1-5. TRN dysfunction has been linked to sensory abnormality, attention deficit and sleep disturbance across multiple neurodevelopmental disorders6-9. However, little is known about the organizational principles that underlie its divergent functions. Here we performed an integrative study linking single-cell molecular and electrophysiological features of the mouse TRN to connectivity and systems-level function. We found that cellular heterogeneity in the TRN is characterized by a transcriptomic gradient of two negatively correlated gene-expression profiles, each containing hundreds of genes. Neurons in the extremes of this transcriptomic gradient express mutually exclusive markers, exhibit core or shell-like anatomical structure and have distinct electrophysiological properties. The two TRN subpopulations make differential connections with the functionally distinct first-order and higher-order thalamic nuclei to form molecularly defined TRN-thalamus subnetworks. Selective perturbation of the two subnetworks in vivo revealed their differential role in regulating sleep. In sum, our study provides a comprehensive atlas of TRN neurons at single-cell resolution and links molecularly defined subnetworks to the functional organization of thalamocortical circuits.
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Redes Reguladoras de Genes , Núcleos Talámicos/citología , Núcleos Talámicos/metabolismo , Animales , Análisis por Conglomerados , Femenino , Perfilación de la Expresión Génica , Hibridación Fluorescente in Situ , Metaloendopeptidasas/metabolismo , Ratones , Vías Nerviosas , Neuronas/metabolismo , Osteopontina/metabolismo , Técnicas de Placa-Clamp , RNA-Seq , Análisis de la Célula Individual , Sueño/genética , Sueño/fisiología , Núcleos Talámicos/fisiología , TranscriptomaRESUMEN
BACKGROUND & AIMS: Excess copper causes hepatocyte death in hereditary Wilson's disease (WD). Current WD treatments by copper-binding chelators may gradually reduce copper overload; they fail, however, to bring hepatic copper close to normal physiological levels. Consequently, lifelong daily dose regimens are required to hinder disease progression. This may result in severe issues due to nonadherence or unwanted adverse drug reactions and also due to drug switching and ultimate treatment failures. This study comparatively tested bacteria-derived copper binding agents-methanobactins (MBs)-for efficient liver copper depletion in WD rats as well as their safety and effect duration. METHODS: Copper chelators were tested in vitro and in vivo in WD rats. Metabolic cage housing allowed the accurate assessment of animal copper balances and long-term experiments related to the determination of minimal treatment phases. RESULTS: We found that copper-binding ARBM101 (previously known as MB-SB2) depletes WD rat liver copper dose dependently via fecal excretion down to normal physiological levels within 8 days, superseding the need for continuous treatment. Consequently, we developed a new treatment consisting of repetitive cycles, each of â¼1 week of ARBM101 applications, followed by months of in-between treatment pauses to ensure a healthy long-term survival in WD rats. CONCLUSIONS: ARBM101 safely and efficiently depletes excess liver copper from WD rats, thus allowing for short treatment periods as well as prolonged in-between rest periods.
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Degeneración Hepatolenticular , Ratas , Animales , Degeneración Hepatolenticular/tratamiento farmacológico , Degeneración Hepatolenticular/metabolismo , Cobre , Eliminación Hepatobiliar , Hígado/metabolismo , Quelantes/farmacología , Quelantes/uso terapéuticoRESUMEN
BACKGROUND AND AIMS: In patients with acute liver failure (ALF) who suffer from massive hepatocyte loss, liver progenitor cells (LPCs) take over key hepatocyte functions, which ultimately determines survival. This study investigated how the expression of hepatocyte nuclear factor 4α (HNF4α), its regulators, and targets in LPCs determines clinical outcome of patients with ALF. APPROACH AND RESULTS: Clinicopathological associations were scrutinized in 19 patients with ALF (9 recovered and 10 receiving liver transplantation). Regulatory mechanisms between follistatin, activin, HNF4α, and coagulation factor expression in LPC were investigated in vitro and in metronidazole-treated zebrafish. A prospective clinical study followed up 186 patients with cirrhosis for 80 months to observe the relevance of follistatin levels in prevalence and mortality of acute-on-chronic liver failure. Recovered patients with ALF robustly express HNF4α in either LPCs or remaining hepatocytes. As in hepatocytes, HNF4α controls the expression of coagulation factors by binding to their promoters in LPC. HNF4α expression in LPCs requires the forkhead box protein H1-Sma and Mad homolog 2/3/4 transcription factor complex, which is promoted by the TGF-ß superfamily member activin. Activin signaling in LPCs is negatively regulated by follistatin, a hepatocyte-derived hormone controlled by insulin and glucagon. In contrast to patients requiring liver transplantation, recovered patients demonstrate a normal activin/follistatin ratio, robust abundance of the activin effectors phosphorylated Sma and Mad homolog 2 and HNF4α in LPCs, leading to significantly improved coagulation function. A follow-up study indicated that serum follistatin levels could predict the incidence and mortality of acute-on-chronic liver failure. CONCLUSIONS: These results highlight a crucial role of the follistatin-controlled activin-HNF4α-coagulation axis in determining the clinical outcome of massive hepatocyte loss-induced ALF. The effects of insulin and glucagon on follistatin suggest a key role of the systemic metabolic state in ALF.
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Activinas/genética , Folistatina/metabolismo , Factor Nuclear 4 del Hepatocito/metabolismo , Fallo Hepático Agudo/metabolismo , Activinas/metabolismo , Insuficiencia Hepática Crónica Agudizada/sangre , Adulto , Anciano , Animales , Coagulación Sanguínea , Línea Celular , Factor V/genética , Femenino , Folistatina/sangre , Estudios de Seguimiento , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/metabolismo , Expresión Génica , Factor Nuclear 4 del Hepatocito/genética , Hepatocitos/metabolismo , Humanos , Fallo Hepático Agudo/inducido químicamente , Fallo Hepático Agudo/patología , Fallo Hepático Agudo/cirugía , Regeneración Hepática , Trasplante de Hígado , Masculino , Metronidazol , Ratones , Persona de Mediana Edad , Pronóstico , Regiones Promotoras Genéticas , Estudios Prospectivos , Protrombina/genética , Transducción de Señal , Proteína Smad2/genética , Proteína Smad2/metabolismo , Proteína smad3/genética , Proteína smad3/metabolismo , Proteína Smad4/genética , Células Madre/metabolismo , Factor de Crecimiento Transformador beta1/genética , Pez CebraRESUMEN
Although interactions between the thalamus and cortex are critical for cognitive function, the exact contribution of the thalamus to these interactions remains unclear. Recent studies have shown diverse connectivity patterns across the thalamus, but whether this diversity translates to thalamic functions beyond relaying information to or between cortical regions is unknown. Here we show, by investigating the representation of two rules used to guide attention in the mouse prefrontal cortex (PFC), that the mediodorsal thalamus sustains these representations without relaying categorical information. Specifically, mediodorsal input amplifies local PFC connectivity, enabling rule-specific neural sequences to emerge and thereby maintain rule representations. Consistent with this notion, broadly enhancing PFC excitability diminishes rule specificity and behavioural performance, whereas enhancing mediodorsal excitability improves both. Overall, our results define a previously unknown principle in neuroscience; thalamic control of functional cortical connectivity. This function, which is dissociable from categorical information relay, indicates that the thalamus has a much broader role in cognition than previously thought.
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Atención/fisiología , Corteza Prefrontal/fisiología , Tálamo/fisiología , Animales , Cognición/fisiología , Masculino , Ratones , Vías Nerviosas , Optogenética , Corteza Prefrontal/citología , Tálamo/citologíaRESUMEN
Developmental disabilities, including attention-deficit hyperactivity disorder (ADHD), intellectual disability (ID), and autism spectrum disorders (ASD), affect one in six children in the USA. Recently, gene mutations in patched domain containing 1 (PTCHD1) have been found in ~1% of patients with ID and ASD. Individuals with PTCHD1 deletion show symptoms of ADHD, sleep disruption, hypotonia, aggression, ASD, and ID. Although PTCHD1 is probably critical for normal development, the connection between its deletion and the ensuing behavioural defects is poorly understood. Here we report that during early post-natal development, mouse Ptchd1 is selectively expressed in the thalamic reticular nucleus (TRN), a group of GABAergic neurons that regulate thalamocortical transmission, sleep rhythms, and attention. Ptchd1 deletion attenuates TRN activity through mechanisms involving small conductance calcium-dependent potassium currents (SK). TRN-restricted deletion of Ptchd1 leads to attention deficits and hyperactivity, both of which are rescued by pharmacological augmentation of SK channel activity. Global Ptchd1 deletion recapitulates learning impairment, hyper-aggression, and motor defects, all of which are insensitive to SK pharmacological targeting and not found in the TRN-restricted deletion mouse. This study maps clinically relevant behavioural phenotypes onto TRN dysfunction in a human disease model, while also identifying molecular and circuit targets for intervention.
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Trastorno por Déficit de Atención con Hiperactividad/fisiopatología , Trastorno por Déficit de Atención con Hiperactividad/psicología , Eliminación de Gen , Proteínas de la Membrana/deficiencia , Proteínas de la Membrana/genética , Núcleos Talámicos/fisiopatología , Agresión , Animales , Animales Recién Nacidos , Atención , Trastorno por Déficit de Atención con Hiperactividad/genética , Conducta Animal , Modelos Animales de Enfermedad , Conductividad Eléctrica , Femenino , Neuronas GABAérgicas/metabolismo , Neuronas GABAérgicas/patología , Humanos , Discapacidades para el Aprendizaje/genética , Discapacidades para el Aprendizaje/fisiopatología , Masculino , Proteínas de la Membrana/metabolismo , Ratones , Ratones Noqueados , Trastornos Motores/genética , Trastornos Motores/fisiopatología , Inhibición Neural , Canales de Potasio Calcio-Activados/metabolismo , Sueño , Privación de Sueño/genética , Privación de Sueño/fisiopatología , Núcleos Talámicos/patologíaRESUMEN
How the brain selects appropriate sensory inputs and suppresses distractors is unknown. Given the well-established role of the prefrontal cortex (PFC) in executive function, its interactions with sensory cortical areas during attention have been hypothesized to control sensory selection. To test this idea and, more generally, dissect the circuits underlying sensory selection, we developed a cross-modal divided-attention task in mice that allowed genetic access to this cognitive process. By optogenetically perturbing PFC function in a temporally precise window, the ability of mice to select appropriately between conflicting visual and auditory stimuli was diminished. Equivalent sensory thalamocortical manipulations showed that behaviour was causally dependent on PFC interactions with the sensory thalamus, not sensory cortex. Consistent with this notion, we found neurons of the visual thalamic reticular nucleus (visTRN) to exhibit PFC-dependent changes in firing rate predictive of the modality selected. visTRN activity was causal to performance as confirmed by bidirectional optogenetic manipulations of this subnetwork. Using a combination of electrophysiology and intracellular chloride photometry, we demonstrated that visTRN dynamically controls visual thalamic gain through feedforward inhibition. Our experiments introduce a new subcortical model of sensory selection, in which the PFC biases thalamic reticular subnetworks to control thalamic sensory gain, selecting appropriate inputs for further processing.
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Atención/fisiología , Células Receptoras Sensoriales/fisiología , Tálamo/fisiología , Estimulación Acústica , Animales , Giro del Cíngulo/fisiología , Masculino , Ratones , Ratones Endogámicos C57BL , Modelos Neurológicos , Inhibición Neural/fisiología , Vías Nerviosas/fisiología , Optogenética , Estimulación Luminosa , Corteza Prefrontal/fisiología , Núcleos Talámicos/citología , Núcleos Talámicos/fisiología , Tálamo/citologíaRESUMEN
Non-alcoholic fatty liver disease (NAFLD) is rising in prevalence, and a better pathophysiologic understanding of the transition to its inflammatory phenotype (NASH) is key to the development of effective therapies. To evaluate the contribution of the NLRP3 inflammasome and its downstream effectors IL-1 and IL-18 in this process, we applied the true-to-life "American lifestyle-induced obesity syndrome" (ALiOS) diet mouse model. Development of obesity, fatty liver and liver damage was investigated in mice fed for 24 weeks according to the ALiOS protocol. Lipidomic changes in mouse livers were compared to human NAFLD samples. Receptor knockout mice for IL-1 and IL-18 were used to dissect the impact of downstream signals of inflammasome activity on the development of NAFLD. The ALiOS diet induced obesity and liver steatosis. The lipidomic changes closely mimicked changes in human NAFLD. A pro-inflammatory gene expression pattern in liver tissue and increased serum liver transaminases indicated early liver damage in the absence of histological evidence of NASH. Mechanistically, Il-18r-/-- but not Il-1r-/- mice were protected from early liver damage, possibly due to silencing of the pro-inflammatory gene expression pattern. Our study identified NLRP3 activation and IL-18R-dependent signaling as potential modulators of early liver damage in NAFLD, preceding development of histologic NASH.
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Interleucina-18/metabolismo , Interleucina-1/metabolismo , Hígado/lesiones , Hígado/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Transducción de Señal , Animales , Interleucina-1/genética , Interleucina-18/genética , Hígado/patología , Masculino , Ratones , Ratones Noqueados , Enfermedad del Hígado Graso no Alcohólico/genética , Enfermedad del Hígado Graso no Alcohólico/patología , Receptores de Interleucina-1/genética , Receptores de Interleucina-1/metabolismo , Receptores de Interleucina-18/genética , Receptores de Interleucina-18/metabolismoRESUMEN
Liver cirrhosis is a life-threatening consequence of liver fibrosis. The aim of this study was to investigate the antifibrotic potential of clinically available vitamin D analogs compared to that of calcitriol in vitro and in vivo. Murine hepatic stellate cells, Kupffer cells, and human LX-2 cells were treated with vitamin D analogs, and the profibrotic behavior of these cells was studied. In vivo liver fibrosis was induced using CCl4 until measurable fibrosis was established. Animals were then treated with calcitriol and paricalcitol. Vitamin D and its analogs showed antifibrotic effects in vitro. Treatment with active vitamin D (calcitriol, CAL) and its analogs reduced the protein expression of α-smooth muscle actin (α-SMA) in mHSC. In human LX-2 cells alfacalcidol reduced transforming growth factor-ß (TGF-ß) induced platelet-derived growth factor receptor-ß protein expression and contractility while paricalcitol (PCT), in its equipotent dose to CAL, reduced TGF-ß induced α-SMA protein expression, and ACTA2 and TGF-ß mRNA expression. No effects of a treatment with vitamin D and its analogs were observed in Kupffer cells. In vivo, PCT-treated mice had significantly lower calcium levels than CAL-treated mice. CAL and PCT reduced the hepatic infiltration of CD11b-positive cells and alanine transaminase levels, while PCT but not CAL significantly inhibited fibrosis progression, with a favorable side effect profile in the CCl4 model. We conclude that hypocalcemic vitamin D analogs should be considered in future studies investigating vitamin D for the treatment of liver fibrosis.
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Ergocalciferoles/uso terapéutico , Células Estrelladas Hepáticas/efectos de los fármacos , Cirrosis Hepática/tratamiento farmacológico , Animales , Calcitriol/farmacología , Calcitriol/uso terapéutico , Calcio/sangre , Tetracloruro de Carbono , Línea Celular , Evaluación Preclínica de Medicamentos , Ergocalciferoles/farmacología , Humanos , Macrófagos del Hígado/efectos de los fármacos , Cirrosis Hepática/inducido químicamente , Masculino , Ratones Endogámicos C57BL , Cultivo Primario de Células , Factor de Crecimiento Transformador beta , Vitamina D/análogos & derivadosRESUMEN
The thalamus has traditionally been considered as only a relay source of cortical inputs, with hierarchically organized cortical circuits serially transforming thalamic signals to cognitively relevant representations. Given the absence of local excitatory connections within the thalamus, the notion of thalamic relay seemed like a reasonable description over the past several decades. Recent advances in experimental approaches and theory provide a broader perspective on the role of the thalamus in cognitively relevant cortical computations and suggest that only a subset of thalamic circuit motifs fits the relay description. Here, we discuss this perspective and highlight the potential role for the thalamus, and specifically the mediodorsal (MD) nucleus, in the dynamic selection of cortical representations through a combination of intrinsic thalamic computations and output signals that change cortical network functional parameters. We suggest that through the contextual modulation of cortical computation, the thalamus and cortex jointly optimize the information and cost trade-off in an emergent fashion. We emphasize that coordinated experimental and theoretical efforts will provide a path to understanding the role of the thalamus in cognition, along with an understanding to augment cognitive capacity in health and disease.
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Inteligencia Artificial , Cognición/fisiología , Vías Nerviosas/fisiología , Tálamo/fisiología , Corteza Cerebral/fisiología , HumanosRESUMEN
The prevalence of obesity-related nonalcoholic fatty liver disease (NAFLD) is rising. NAFLD may result in nonalcoholic steatohepatitis (NASH), progressing to liver cirrhosis. Weight loss is recommended to treat obesity-related NASH. Lifestyle intervention may improve NASH; however, pertinent trials have so far focused on overweight patients, whereas patients with obesity are at highest risk of developing NAFLD. Furthermore, reports of effects on liver fibrosis are scarce. We evaluated the effect of lifestyle intervention on NAFLD in a real-life cohort of morbidly obese patients. In our observational study, 152 patients underwent lifestyle intervention, with a follow-up of 52 weeks. Noninvasive measures of obesity, metabolic syndrome, liver steatosis, liver damage, and liver fibrosis were analyzed. Treatment response in terms of weight loss was achieved in 85.1% of patients. Dysglycemia and dyslipidemia improved. The proportion of patients with fatty liver dropped from 98.1 to 54.3% ( P < 0.001). Weight loss >10% was associated with better treatment response ( P = 0.0009). Prevalence of abnormal serum transaminases fell from 81.0 to 50.5% ( P < 0.001). The proportion fibrotic patients, as determined by the NAFLD fibrosis score, dropped from 11.8 to 0% ( P < 0.05). Low serum levels of adiponectin correlated with degree of liver damage, i.e., serum liver transaminases ( r = -0,32, P < 0.05). Serum levels of adiponectin improved with intervention. In conclusion, lifestyle intervention effectively targeted obesity and the metabolic syndrome. Liver steatosis, damage and fibrosis were ameliorated in this real-life cohort of morbidly obese patients, mediated in part by changes in the adipokine profile. Patients with weight loss of >10% seemed to benefit most. NEW & NOTEWORTHY We demonstrate new evidence that lifestyle intervention is effective in treating NAFLD in the important group of patients with (morbid) obesity. Although current guidelines on the therapy of NASH recommend weight loss of 5-7%, weight reduction >10% may be favorable in morbid obesity. Serum levels of adipokines correlate with liver damage, which is indicative of their pathogenetic importance in human NASH. Our study adds to the limited body of evidence that NAFLD-associated liver fibrosis may resolve with lifestyle intervention.
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Adiponectina/sangre , Dietoterapia/métodos , Cirrosis Hepática , Enfermedad del Hígado Graso no Alcohólico , Obesidad Mórbida , Pérdida de Peso/fisiología , Adipoquinas/sangre , Adulto , Índice de Masa Corporal , Femenino , Alemania/epidemiología , Conductas Relacionadas con la Salud/fisiología , Estilo de Vida Saludable/fisiología , Humanos , Estilo de Vida , Hígado/metabolismo , Hígado/patología , Cirrosis Hepática/metabolismo , Cirrosis Hepática/patología , Cirrosis Hepática/prevención & control , Pruebas de Función Hepática/métodos , Masculino , Persona de Mediana Edad , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/patología , Enfermedad del Hígado Graso no Alcohólico/prevención & control , Enfermedad del Hígado Graso no Alcohólico/psicología , Obesidad Mórbida/fisiopatología , Obesidad Mórbida/psicología , Obesidad Mórbida/terapiaRESUMEN
BACKGROUND/PURPOSE OF THE STUDY: Vitamin D3-deficiency is common in patients with chronic liver-disease and may promote disease progression. Vitamin D3-administration has thus been proposed as a therapeutic approach. Vitamin D3 has immunomodulatory effects and may modulate autoimmune liver-disease such as primary sclerosing cholangitis. Although various mechanisms of action have been proposed, experimental evidence is limited. Here we test the hypothesis that active 1,25-(OH)2-vitamin D3 inhibits activation of hepatic stellate cells (HSC) in vitro and modulates liver-injury in vivo. METHODS: Proliferation and activation of primary murine HSC were assessed by BrdU- and PicoGreen(®)-assays, immunoblotting, immunofluorescence-microscopy, quantitative-PCR, and zymography following calcitriol-treatment. Wild-type and ATP-binding cassette transporter b4(-/-) (Abcb4(-/-))-mice received calcitriol for 4 weeks. Liver-damage, inflammation, and fibrosis were assessed by serum liver-tests, Sirius-red staining, quantitative-PCR, immunoblotting, immunohistochemistry and hydroxyproline quantification. RESULTS: In vitro, calcitriol inhibited activation and proliferation of murine HSC as shown by reduced α-smooth muscle actin and platelet-derived growth factor-receptor-ß-protein-levels, BrdU and PicoGreen®-assays. Furthermore, mRNA-levels and activity of matrix metalloproteinase 13 were profoundly increased. In vivo, calcitriol ameliorated inflammatory liver-injury reflected by reduced levels of alanine aminotransferase in Abcb4(-/-)-mice. In accordance, their livers had lower mRNA-levels of F4/80, tumor necrosis factor-receptor 1 and a lower count of portal CD11b positive cells. In contrast, no effect on overall fibrosis was observed. CONCLUSION: Calcitriol inhibits activation and proliferation of HSCs in vitro. In Abcb4(-/-)-mice, administration of calcitriol ameliorates inflammatory liver-damage but has no effect on biliary fibrosis after 4 weeks of treatment.
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Subfamilia B de Transportador de Casetes de Unión a ATP/deficiencia , Calcitriol/farmacología , Células Estrelladas Hepáticas/efectos de los fármacos , Hepatitis Animal/tratamiento farmacológico , Cirrosis Hepática/tratamiento farmacológico , Subfamilia B de Transportador de Casetes de Unión a ATP/genética , Animales , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Modelos Animales de Enfermedad , Células Estrelladas Hepáticas/inmunología , Células Estrelladas Hepáticas/patología , Hepatitis Animal/inmunología , Hepatitis Animal/patología , Factores Inmunológicos/farmacología , Cirrosis Hepática/inmunología , Cirrosis Hepática/patología , Metaloproteinasa 13 de la Matriz/genética , Metaloproteinasa 13 de la Matriz/metabolismo , Ratones , Ratones Noqueados , ARN Mensajero/genética , ARN Mensajero/metabolismo , Miembro 4 de la Subfamilia B de Casete de Unión a ATPRESUMEN
The cell-toxic bile salt glycochenodeoxycholic acid (GCDCA) and taurochenodeoxycholic acid (TCDCA) are responsible for hepatocyte demise in cholestatic liver diseases, while tauroursodeoxycholic acid (TUDCA) is regarded hepatoprotective. We demonstrate the direct mitochondrio-toxicity of bile salts which deplete the mitochondrial membrane potential and induce the mitochondrial permeability transition (MPT). The bile salt mediated mechanistic mode of destruction significantly differs from that of calcium, the prototype MPT inducer. Cell-toxic bile salts initially bind to the mitochondrial outer membrane. Subsequently, the structure of the inner boundary membrane disintegrates. And it is only thereafter that the MPT is induced. This progressive destruction occurs in a dose- and time-dependent way. We demonstrate that GCDCA and TCDCA, but not TUDCA, preferentially permeabilize liposomes containing the mitochondrial membrane protein ANT, a process resembling the MPT induction in whole mitochondria. This suggests that ANT is one decisive target for toxic bile salts. To our knowledge this is the first report unraveling the consecutive steps leading to mitochondrial destruction by cell-toxic bile salts.
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Ácido Glicoquenodesoxicólico/toxicidad , Mitocondrias Hepáticas/efectos de los fármacos , Translocasas Mitocondriales de ADP y ATP/agonistas , Ácido Tauroquenodesoxicólico/farmacología , Animales , Permeabilidad de la Membrana Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Liposomas/química , Hígado/química , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias Cardíacas/química , Mitocondrias Hepáticas/metabolismo , Mitocondrias Hepáticas/patología , Translocasas Mitocondriales de ADP y ATP/aislamiento & purificación , Proteínas de Transporte de Membrana Mitocondrial/agonistas , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Membranas Mitocondriales/química , Membranas Mitocondriales/efectos de los fármacos , Poro de Transición de la Permeabilidad Mitocondrial , Miocardio/química , Ratas , Ácido Tauroquenodesoxicólico/toxicidad , Canales Aniónicos Dependientes del Voltaje/química , Canales Aniónicos Dependientes del Voltaje/aislamiento & purificaciónRESUMEN
BACKGROUND & AIMS: Hydrophobic bile salts such as glycochenodeoxycholate (GCDC) accumulate in cholestatic liver disease and induce hepatocellular apoptosis, promoting profibrotic signalling. The tissue microenvironment is an integral player in cellular pathophysiology, but it is not routinely incorporated into laboratory studies. Tissue oxygen partial pressure (pO2) may be an underestimated component of the microenvironment: in the liver, a pO2 of 30-45 mmHg (approximately 6% O2) is physiological, because of predominant portal blood supply. It was the aim of this project to investigate the impact of physiological hypoxia (i.e. 6% O2) on hepatocellular function, namely, bile salt-induced apoptosis. METHODS: Human hepatoma cells (HepG2-Ntcp) and primary rat hepatocytes were cultured at standard laboratory (hyperoxic) conditions (21% O2) and at physiological hypoxia (6% O2) in parallel for 1-8 days to study hepatocellular apoptosis and activation of signalling pathways. Standard laboratory analyses were applied for bile salt uptake, caspase-3/-7 activity, western blotting and gene-array analysis. RESULTS: Culturing at physiological hypoxia protected both human and rat hepatocytes against GCDC-induced apoptosis: caspase-3/-7 activation was diminished by 3.1 ± 0.5-fold in human HepG2-Ntcp and completely abolished in primary rat hepatocytes. Bile salt uptake was unaffected. Induction of hypoxia-inducible factor-1α indicated adaption to physiological hypoxia. The MEK/ERK cascade was activated and anti-apoptotic mediators were induced: N-Myc down-regulated gene, gelsolin and carbonic anhydrase IX were upregulated 12.4-, 6.5- and 5.2-fold respectively. CONCLUSIONS: We conclude from these data that (i) physiological hypoxia protects hepatocytes from bile salt-induced apoptosis, (ii) tissue pO2 is a crucial, underestimated component of the microenvironment and should (iii) be considered when studying hepatocellular physiology in vitro.
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Apoptosis/fisiología , Puntos de Control del Ciclo Celular/fisiología , Hipoxia de la Célula/fisiología , Hepatocitos/fisiología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Animales , Apoptosis/efectos de los fármacos , Ácidos y Sales Biliares/efectos adversos , Western Blotting , Puntos de Control del Ciclo Celular/efectos de los fármacos , Citometría de Flujo , Células Hep G2 , Humanos , Análisis por Micromatrices , Ratas , Transducción de Señal/fisiologíaRESUMEN
AIM: Hepatic apoptosis is involved in the pathogenesis of immune-mediated liver diseases such as autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC). The aim of our study was to quantify distinct markers of apoptosis in sera of patients with AIH, PBC and PSC, and to evaluate correlation with markers of disease activity and prognosis. METHODS: Sera of patients with AIH, PBC and PSC, and of healthy controls were collected and distinct cell death markers were quantified using a bead-based multiplex enzyme linked immunosorbent assay (soluble intracellular adhesion molecule [sICAM], macrophage migration inhibitory factor [MIF], soluble Fas [sFas], plasminogen activator inhibitor 1 [PAI-1]) or single enzyme-linked immunosorbent assays (DNAse, M30, M65). RESULTS: In comparison with healthy controls, the apoptotic markers sFas, sICAM (only in PSC patients), M30 and the cell death marker M65 were substantially elevated in sera of patients with immune-mediated liver diseases, whereas DNAse activity was reduced. Interestingly, patients with advanced PSC presented with higher levels of sICAM, M30 and M65 than patients with mild PSC. Regression analysis revealed correlations between serum levels of sICAM, M30 and M65 with the Mayo Risk Score for PSC, and of M65 with the Mayo Risk Score for PBC. CONCLUSION: Concentrations of the serum markers of apoptosis sFas and M30 and of the marker of total cell death M65 are elevated in patients with immune-mediated liver diseases, whereas activity of DNAse is reduced. In patients with PSC, sICAM, M30 and M65 may serve as indicators for disease activity and prognosis.
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Low-threshold (T-type) Ca(2+) channels encoded by the Ca(V)3 genes endow neurons with oscillatory properties that underlie slow waves characteristic of the non-rapid eye movement (NREM) sleep EEG. Three Ca(V)3 channel subtypes are expressed in the thalamocortical (TC) system, but their respective roles for the sleep EEG are unclear. Ca(V)3.3 protein is expressed abundantly in the nucleus reticularis thalami (nRt), an essential oscillatory burst generator. We report the characterization of a transgenic Ca(V)3.3(-/-) mouse line and demonstrate that Ca(V)3.3 channels are indispensable for nRt function and for sleep spindles, a hallmark of natural sleep. The absence of Ca(V)3.3 channels prevented oscillatory bursting in the low-frequency (4-10 Hz) range in nRt cells but spared tonic discharge. In contrast, adjacent TC neurons expressing Ca(V)3.1 channels retained low-threshold bursts. Nevertheless, the generation of synchronized thalamic network oscillations underlying sleep-spindle waves was weakened markedly because of the reduced inhibition of TC neurons via nRt cells. T currents in Ca(V)3.3(-/-) mice were <30% compared with those in WT mice, and the remaining current, carried by Ca(V)3.2 channels, generated dendritic [Ca(2+)](i) signals insufficient to provoke oscillatory bursting that arises from interplay with Ca(2+)-dependent small conductance-type 2 K(+) channels. Finally, naturally sleeping Ca(V)3.3(-/-) mice showed a selective reduction in the power density of the σ frequency band (10-12 Hz) at transitions from NREM to REM sleep, with other EEG waves remaining unaltered. Together, these data identify a central role for Ca(V)3.3 channels in the rhythmogenic properties of the sleep-spindle generator and provide a molecular target to elucidate the roles of sleep spindles for brain function and development.
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Canales de Calcio Tipo T/fisiología , Sueño/fisiología , Tálamo/fisiología , Animales , Ondas Encefálicas , Señalización del Calcio , Electroencefalografía , Ratones , Ratones Noqueados , Neuronas/fisiología , Sueño REMRESUMEN
Primary sclerosing cholangitis (PSC) represents a chronic liver disease characterized by poor prognosis and lacking causal treatment options. Yes-associated protein (YAP) functions as a critical mediator of fibrogenesis; however, its therapeutic potential in chronic biliary diseases such as PSC remains unestablished. The objective of this study is to elucidate the possible significance of YAP inhibition in biliary fibrosis by examining the pathophysiology of hepatic stellate cells (HSC) and biliary epithelial cells (BEC). Human liver tissue samples from PSC patients were analyzed to assess the expression of YAP/connective tissue growth factor (CTGF) relative to non-fibrotic control samples. The pathophysiological relevance of YAP/CTGF in HSC and BEC was investigated in primary human HSC (phHSC), LX-2, H69, and TFK-1 cell lines through siRNA or pharmacological inhibition utilizing verteporfin (VP) and metformin (MF). The Abcb4-/- mouse model was employed to evaluate the protective effects of pharmacological YAP inhibition. Hanging droplet and 3D matrigel culture techniques were utilized to investigate YAP expression and activation status of phHSC under various physical conditions. YAP/CTGF upregulation was observed in PSC patients. Silencing YAP/CTGF led to inhibition of phHSC activation and reduced contractility of LX-2 cells, as well as suppression of epithelial-mesenchymal transition (EMT) in H69 cells and proliferation of TFK-1 cells. Pharmacological inhibition of YAP mitigated chronic liver fibrosis in vivo and diminished ductular reaction and EMT. YAP expression in phHSC was effectively modulated by altering extracellular stiffness, highlighting YAP's role as a mechanotransducer. In conclusion, YAP regulates the activation of HSC and EMT in BEC, thereby functioning as a checkpoint of fibrogenesis in chronic cholestasis. Both VP and MF demonstrate effectiveness as YAP inhibitors, capable of inhibiting biliary fibrosis. These findings suggest that VP and MF warrant further investigation as potential therapeutic options for the treatment of PSC.