RESUMEN
During CNS development, interneuron precursors have to migrate extensively before they integrate in specific microcircuits. Known regulators of neuronal motility include classical neurotransmitters, yet the mechanisms that assure interneuron dispersal and interneuron/projection neuron matching during histogenesis remain largely elusive. We combined time-lapse video microscopy and electrophysiological analysis of the nascent cerebellum of transgenic Pax2-EGFP mice to address this issue. We found that cerebellar interneuronal precursors regularly show spontaneous postsynaptic currents, indicative of synaptic innervation, well before settling in the molecular layer. In keeping with the sensitivity of these cells to neurotransmitters, ablation of synaptic communication by blocking vesicular release in acute slices of developing cerebella slows migration. Significantly, abrogation of exocytosis primarily impedes the directional persistence of migratory interneuronal precursors. These results establish an unprecedented function of the early synaptic innervation of migrating neuronal precursors and demonstrate a role for synapses in the regulation of migration and pathfinding.
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Movimiento Celular , Interneuronas/citología , Células-Madre Neurales/citología , Sinapsis/metabolismo , Animales , Forma de la Célula , Cerebelo/citología , Cerebelo/ultraestructura , Fenómenos Electrofisiológicos , Femenino , Glutamatos/metabolismo , Interneuronas/metabolismo , Masculino , Ratones Endogámicos C57BL , Modelos Biológicos , Células-Madre Neurales/metabolismo , Factor de Transcripción PAX2/metabolismo , Ácido gamma-Aminobutírico/metabolismoRESUMEN
The present study describes for the first time the neural expression and distribution of UGS148, a protein encoded by the RIKEN cDNA63330403K07 gene that has been shown to be prominently and characteristically expressed in neural stem cells (NSCs). Based on its molecular structure, UGS148 is an intracellular protein expected to be involved in intracellular sorting, trafficking, exocytosis and membrane insertion of proteins. We demonstrate that UGS148 is highly expressed in embryonic NSCs as well as, albeit at low level, in the adult neurogenic niches, the subventricular zone and the hippocampal dentate gyrus. Interestingly, the highest expression level of UGS148 in the adult mouse brain was observed specifically in the neurogenic cells lining the third ventricle, the tanycytes. Our in vitro studies show the involvement of UGS148 in the regulation of the proliferation of NSCs.
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Giro Dentado/metabolismo , Células Ependimogliales/metabolismo , Proteínas del Tejido Nervioso/genética , Células-Madre Neurales/metabolismo , Secuencia de Aminoácidos , Animales , Células Cultivadas , Giro Dentado/citología , Giro Dentado/crecimiento & desarrollo , Ratones , Ratones Endogámicos C57BL , Datos de Secuencia Molecular , Proteínas del Tejido Nervioso/metabolismo , Transporte de ProteínasRESUMEN
BACKGROUND: Cigarette smoking is the major risk factor for COPD, leading to chronic airway inflammation. We hypothesized that cigarette smoke induces structural and functional changes of airway epithelial mitochondria, with important implications for lung inflammation and COPD pathogenesis. METHODS: We studied changes in mitochondrial morphology and in expression of markers for mitochondrial capacity, damage/biogenesis and fission/fusion in the human bronchial epithelial cell line BEAS-2B upon 6-months from ex-smoking COPD GOLD stage IV patients to age-matched smoking and never-smoking controls. RESULTS: We observed that long-term CSE exposure induces robust changes in mitochondrial structure, including fragmentation, branching and quantity of cristae. The majority of these changes were persistent upon CSE depletion. Furthermore, long-term CSE exposure significantly increased the expression of specific fission/fusion markers (Fis1, Mfn1, Mfn2, Drp1 and Opa1), oxidative phosphorylation (OXPHOS) proteins (Complex II, III and V), and oxidative stress (Mn-SOD) markers. These changes were accompanied by increased levels of the pro-inflammatory mediators IL-6, IL-8, and IL-1ß. Importantly, COPD primary bronchial epithelial cells (PBECs) displayed similar changes in mitochondrial morphology as observed in long-term CSE-exposure BEAS-2B cells. Moreover, expression of specific OXPHOS proteins was higher in PBECs from COPD patients than control smokers, as was the expression of mitochondrial stress marker PINK1. CONCLUSION: The observed mitochondrial changes in COPD epithelium are potentially the consequence of long-term exposure to cigarette smoke, leading to impaired mitochondrial function and may play a role in the pathogenesis of COPD.
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Bronquios/patología , Células Epiteliales/patología , Mitocondrias/fisiología , Mitocondrias/ultraestructura , Dinámicas Mitocondriales/fisiología , Recambio Mitocondrial/fisiología , Fumar/efectos adversos , Adulto , Anciano , Bronquios/metabolismo , Estudios de Casos y Controles , Línea Celular , Células Cultivadas , Citocinas/metabolismo , Dinaminas , Células Epiteliales/metabolismo , Femenino , GTP Fosfohidrolasas/metabolismo , Humanos , Técnicas In Vitro , Masculino , Proteínas de la Membrana/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Persona de Mediana Edad , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Proteínas Mitocondriales/metabolismo , Proteínas Quinasas/metabolismo , Enfermedad Pulmonar Obstructiva Crónica/metabolismo , Enfermedad Pulmonar Obstructiva Crónica/patología , Enfermedad Pulmonar Obstructiva Crónica/fisiopatología , Factores de Riesgo , Superóxido Dismutasa/metabolismo , Factores de TiempoRESUMEN
Pantothenate kinase-associated neurodegeneration (PKAN), a progressive neurodegenerative disorder, is associated with impairment of pantothenate kinase function. Pantothenate kinase is the first enzyme required for de novo synthesis of CoA, an essential metabolic cofactor. The pathophysiology of PKAN is not understood, and there is no cure to halt or reverse the symptoms of this devastating disease. Recently, we and others presented a PKAN Drosophila model, and we demonstrated that impaired function of pantothenate kinase induces a neurodegenerative phenotype and a reduced lifespan. We have explored this Drosophila model further and have demonstrated that impairment of pantothenate kinase is associated with decreased levels of CoA, mitochondrial dysfunction, and increased protein oxidation. Furthermore, we searched for compounds that can rescue pertinent phenotypes of the Drosophila PKAN model and identified pantethine. Pantethine feeding restores CoA levels, improves mitochondrial function, rescues brain degeneration, enhances locomotor abilities, and increases lifespan. We show evidence for the presence of a de novo CoA biosynthesis pathway in which pantethine is used as a precursor compound. Importantly, this pathway is effective in the presence of disrupted pantothenate kinase function. Our data suggest that pantethine may serve as a starting point to develop a possible treatment for PKAN.
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Drosophila/metabolismo , Regulación de la Expresión Génica , Panteteína/análogos & derivados , Neurodegeneración Asociada a Pantotenato Quinasa/tratamiento farmacológico , Animales , Encéfalo/patología , Coenzima A/química , Drosophila/genética , Humanos , Mitocondrias/metabolismo , Modelos Biológicos , Mutación , Estrés Oxidativo , Oxígeno/química , Panteteína/farmacología , Fenotipo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismoRESUMEN
OBJECTIVE: Indoleamine-2,3-dioxygenase (IDO) suppresses the function of T-lymphocytes and is an important immune escape mechanism for cancer. Therefore, it is to be expected that IDO influences prognosis of cancer patients. This study aimed to investigate the prognostic role of IDO expression in a large cohort of endometrial carcinoma (EC) patients. METHODS: A tissue microarray containing primary EC tissue of 355 patients treated in a single institution was used to evaluate IDO expression. Expression of IDO was associated with clinicopathological characteristics, survival and previously determined numbers of CD8(+) and Foxp3(+) T-lymphocytes. RESULTS: IDO(high) expression was associated with lower numbers of intratumoral CD8(+) T-lymphocytes (p=0.031). Next to well-known prognostic parameters, IDO(high) expression was independently associated with poor disease specific survival in the general cohort of EC patients (HR 2.62, 95% C.I. 1.48-4.66, p=0.001) and among patients with early stage EC (HR 3.06, 95% C.I. 1.10-8.54, p=0.032). CONCLUSION: Our results show that IDO expression is associated with poor survival. This provides evidence that further research into the use of IDO blocking agents in cancer treatment is valid where it might be a promising new therapeutic strategy.
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Carcinoma/enzimología , Carcinoma/inmunología , Neoplasias Endometriales/enzimología , Neoplasias Endometriales/inmunología , Indolamina-Pirrol 2,3,-Dioxigenasa/metabolismo , Linfocitos T , Anciano , Linfocitos T CD8-positivos , Carcinoma/patología , Línea Celular Tumoral , Intervalos de Confianza , Supervivencia sin Enfermedad , Neoplasias Endometriales/patología , Femenino , Factores de Transcripción Forkhead , Humanos , Recuento de Linfocitos , Persona de Mediana Edad , Análisis Multivariante , Invasividad Neoplásica , Oportunidad Relativa , Modelos de Riesgos ProporcionalesRESUMEN
The most common clinical tachycardia, Atrial Fibrillation (AF), is a progressive disease, caused by cardiomyocyte remodeling, which finally results in contractile dysfunction and AF persistence. Recently, we identified a protective role of heat shock proteins (HSPs), especially the small HSPB1 member, against tachycardia remodeling in experimental AF models. Our understanding of tachycardia remodeling and anti-remodeling drugs is currently hampered by the lack of suitable (genetic) manipulatable in vivo models for rapid screening of key targets in remodeling. We hypothesized that Drosophila melanogaster can be exploited to study tachycardia remodeling and protective effects of HSPs by drug treatments or by utilizing genetically manipulated small HSP-overexpressing strains. Tachypacing of Drosophila pupae resulted in gradual and significant cardiomyocyte remodeling, demonstrated by reduced contraction rate, increase in arrhythmic episodes and reduction in heart wall shortening, compared to normal paced pupae. Heat shock, or pre-treatment with HSP-inducers GGA and BGP-15, resulted in endogenous HSP overexpression and protection against tachycardia remodeling. DmHSP23 overexpressing Drosophilas were protected against tachycardia remodeling, in contrast to overexpression of other small HSPs (DmHSP27, DmHSP67Bc, DmCG4461, DmCG7409, and DmCG14207). (Ultra)structural evaluation of the tachypaced heart wall revealed loss of sarcomeres and mitochondrial damage which were absent in tachypaced DmHSP23 overexpressing Drosophila. In addition, tachypacing induced a significant increase in calpain activity, which was prevented in tachypaced Drosophila overexpressing DmHSP23. Tachypacing of Drosophila resulted in cardiomyocyte remodeling, which was prevented by general HSP-inducing treatments and overexpression of a single small HSP, DmHSP23. Thus, tachypaced D. melanogaster can be used as an in vivo model system for rapid identification of novel targets to combat AF associated cardiomyocyte remodeling.
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Fibrilación Atrial/metabolismo , Fibrilación Atrial/fisiopatología , Drosophila melanogaster/metabolismo , Proteínas de Choque Térmico Pequeñas/metabolismo , Contracción Miocárdica , Animales , Fibrilación Atrial/patología , Calpaína/metabolismo , Modelos Animales de Enfermedad , Diterpenos/farmacología , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/genética , Expresión Génica/genética , Regulación de la Expresión Génica/efectos de los fármacos , Corazón/fisiopatología , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Proteínas de Choque Térmico Pequeñas/genética , Contracción Miocárdica/genética , Oximas/farmacología , Piperidinas/farmacología , Taquicardia/patología , Taquicardia/fisiopatología , Taquicardia/prevención & controlRESUMEN
The geometric and subcellular organization of axon arbors distributes and regulates electrical signaling in neurons and networks, but the underlying mechanisms have remained elusive. In rodent cerebellar cortex, stellate interneurons elaborate characteristic axon arbors that selectively innervate Purkinje cell dendrites and likely regulate dendritic integration. We used GFP BAC transgenic reporter mice to examine the cellular processes and molecular mechanisms underlying the development of stellate cell axons and their innervation pattern. We show that stellate axons are organized and guided towards Purkinje cell dendrites by an intermediate scaffold of Bergmann glial (BG) fibers. The L1 family immunoglobulin protein Close Homologue of L1 (CHL1) is localized to apical BG fibers and stellate cells during the development of stellate axon arbors. In the absence of CHL1, stellate axons deviate from BG fibers and show aberrant branching and orientation. Furthermore, synapse formation between aberrant stellate axons and Purkinje dendrites is reduced and cannot be maintained, leading to progressive atrophy of axon terminals. These results establish BG fibers as a guiding scaffold and CHL1 a molecular signal in the organization of stellate axon arbors and in directing their dendritic innervation.
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Axones/metabolismo , Moléculas de Adhesión Celular/metabolismo , Dendritas/metabolismo , Neuroglía/metabolismo , Células de Purkinje/ultraestructura , Animales , Axones/ultraestructura , Moléculas de Adhesión Celular/inmunología , Corteza Cerebelosa/metabolismo , Dendritas/ultraestructura , Ratones , Ratones Transgénicos , Microscopía Electrónica , Neuroglía/ultraestructura , Células de Purkinje/metabolismo , Sinapsis/fisiología , Ácido gamma-Aminobutírico/metabolismoRESUMEN
AIMS: We previously showed that enhanced myogenic constriction (MC) of peripheral resistance arteries involves active AT(1) receptors in chronic heart failure (CHF). Recent data suggest both transactivation of EGF receptors and caveolae-like microdomains to be implicated in the activity of AT(1) receptors. Thus, we assessed their roles in increased MC in mesenteric arteries of CHF rats. METHODS AND RESULTS: Male Wistar rats underwent myocardial infarction to induce CHF and were sacrificed after 12 weeks. The number of caveolae in smooth muscle cells (SMC) of mesenteric arteries of CHF rats was decreased by 43.6 +/- 4.0%, this was accompanied by increased MC, which was fully normalized to the level of sham by antagonists of the AT(1)-receptor (losartan) or EGF-receptor (AG1478). Acute disruption of caveolae in sham rats affected caveolae numbers and MC to a similar extent as CHF, however MC was only reversed by the antagonist of the EGF-receptor, but not by the AT(1)-receptor antagonist. Further, in sham rats, MC was increased by a sub-threshold concentration of angiotensin II and reversed by both AT(1)- as well as EGF-receptor inhibition. In contrast, increased MC by a sub-threshold concentration of EGF was only reversed by EGF receptor inhibition. CONCLUSION: These findings provide the first evidence that decreased SMC caveolae numbers are involved in enhanced MC in small mesenteric arteries, by affecting AT(1)- and EGF-receptor function. This suggests a novel mechanism involved in increased peripheral resistance in CHF.
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Caveolas/ultraestructura , Receptores ErbB/metabolismo , Insuficiencia Cardíaca/fisiopatología , Arterias Mesentéricas/fisiopatología , Músculo Liso Vascular/fisiopatología , Receptor de Angiotensina Tipo 1/metabolismo , Vasoconstricción/fisiología , Animales , Recuento de Células , Modelos Animales de Enfermedad , Técnica del Anticuerpo Fluorescente , Insuficiencia Cardíaca/metabolismo , Insuficiencia Cardíaca/patología , Inmunohistoquímica , Masculino , Arterias Mesentéricas/metabolismo , Arterias Mesentéricas/ultraestructura , Microscopía Electrónica , Músculo Liso Vascular/metabolismo , Músculo Liso Vascular/ultraestructura , Contracción Miocárdica/fisiología , Ratas , Ratas WistarRESUMEN
Disease-associated misfolded proteins or proteins damaged due to cellular stress are generally disposed via the cellular protein quality-control system. However, under saturating conditions, misfolded proteins will aggregate. In higher eukaryotes, these aggregates can be transported to accumulate in aggresomes at the microtubule organizing center. The fate of cells that contain aggresomes is currently unknown. Here we report that cells that have formed aggresomes can undergo normal mitosis. As a result, the aggregated proteins are asymmetrically distributed to one of the daughter cells, leaving the other daughter free of accumulated protein damage. Using both epithelial crypts of the small intestine of patients with a protein folding disease and Drosophila melanogaster neural precursor cells as models, we found that the inheritance of protein aggregates during mitosis occurs with a fixed polarity indicative of a mechanism to preserve the long-lived progeny.
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Polaridad Celular , Células Eucariotas/citología , Células Eucariotas/metabolismo , Proteínas/metabolismo , Animales , Células Cultivadas , Cricetinae , Drosophila melanogaster , Humanos , Mitosis , Ácido Poliglutámico/metabolismoRESUMEN
Layer II of the medial entorhinal cortex (MEC LII) contains the largest number of spatially modulated grid cells and is one of the first regions in the brain to express Alzheimer's disease (AD)-related pathology. The most common principal cell type in MEC LII, reelin-expressing stellate cells, are grid cell candidates. Recently we found evidence that γ-aminobutyric acid (GABA)A receptor subunits show a specific distribution in MEC LII, in which GABAA α3 is selectively associated with reelin-positive neurons, with limited association with the other principal cell type, calbindin (CB)-positive pyramidal neurons. Furthermore, the expression of α3 subunit decreases in mice between P15 and P25, which coincides with the emergence of stable grid cell activity. It has been shown that the α3 subunit undergoes specific developmental changes and that it may exert pro-inflammatory actions if improperly regulated. In this review article, we evaluate the changing kinetics of α3-GABAA receptors (GABAARs). during development in relation to α3-subunit expression pattern in MEC LII and conclude that α3 could be closely related to the stabilization of grid cell activity and theta oscillations. We further conclude that dysregulated α3 may be a driving factor in early AD pathology.
RESUMEN
Glioblastomas (GBMs), a type of highly malignant brain tumour, contain various macrophages/microglia that are known as tumour-associated macrophages (TAMs). These TAMs have various roles in tumour biology. Histopathological aspects of TAMs and associations with tumour growth assessed by magnetic resonance imaging (MRI) are poorly described. In the present study, 16 patients that had sufficient tumour tissue and histological hallmarks were examined. The tumours were classified as either slow- (n=7) or fast-growing (n=9) based on the segmented tumour volumes from MRI scans taken at diagnosis and preoperatively. Using cluster of differentiation (CD)68 and ionized calcium-binding adaptor molecule 1 (Iba1) antibodies, the number, morphology, localization and distribution of TAMs in the GBM tissue were studied. TAMs were significantly more immunoreactive for anti-Iba1 (TAMsIba1) compared with anti-CD68 (TAMsCD68; P<0.001). In central tumour areas and around vessels in the infiltration zone there were more TAMsCD68 in slow-growing tumours (P=0.003 and P=0.025, respectively). Central tumour areas contained more TAMs compared with the infiltration zone (P=0.001 for TAMsCD68 and P<0.001 for TAMsIba1). The majority of TAMs exhibited a ramified phenotype in the infiltration zone, whereas central TAMs were mostly amoeboid. TAMs were present in high numbers in most regions of the tumour, whereas there were few in necrotic areas. In conclusion, the present study demonstrated and confirmed that the high numbers of TAMs in GBMs assume a range of morphologies consistent with various activation states, and that slow-growing GBMs seem to contain a TAM-population different to their fast-growing counterparts.
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Autofagia , Médula Ósea/patología , Células Endoteliales/patología , Leucemia Mieloide Aguda/patología , Síndromes Mielodisplásicos/patología , Médula Ósea/irrigación sanguínea , Células Endoteliales/química , Células Endoteliales/ultraestructura , Humanos , Leucemia Mieloide Aguda/metabolismo , Lisosomas/ultraestructura , Microscopía Electrónica , Síndromes Mielodisplásicos/metabolismo , Vacuolas/ultraestructura , Factor A de Crecimiento Endotelial Vascular/análisisRESUMEN
The corticotropin-releasing factor (CRF) type 2 receptor has three splice variants alpha, beta, and gamma. In the rodent brain only CRFR2alpha is present. In the cerebellum, CRF-R2alpha has two different isoforms: a full-length form (fl) and truncated (tr). Both forms CRF-R2 have a unique cellular distribution. During postnatal cerebellar development, the expression patterns of tr and fl isoforms are changing. This suggests that, CRF and the related peptide urocortin (UCN) could play distinct roles in the immature and adult cerebellum, acting via different receptors subtypes. This review focuses on differences in the distribution of each isoform of CRF-R2 in view of their relationship to CRF and UCN release sites and their possible functional implications. Moreover, it includes novel findings of molecular pathways activating CRF-R2 isoforms through which CRF and UCN excert their specific actions.
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Cerebelo/fisiología , Receptores de Hormona Liberadora de Corticotropina/fisiología , Animales , Axones/fisiología , Ratones , Modelos Moleculares , Conformación Proteica , Isoformas de Proteínas/análisis , Isoformas de Proteínas/fisiología , Células de Purkinje/fisiología , Receptores de Hormona Liberadora de Corticotropina/análisis , Receptores de Hormona Liberadora de Corticotropina/químicaRESUMEN
GABAergic parvalbumin-expressing (PV+) interneurons provide powerful inhibitory modulation of grid cells in layer II of the medial entorhinal cortex (MEC LII). However, the molecular machinery through which PV+ cells regulate grid cell activity is poorly defined. PV+ interneurons impart inhibitory modulation primarily via GABA-A receptors (GABAARs). GABAARs are pentameric ion channels assembled from a repertoire of 19 subunits. Multiple subunit combinations result in a variety of receptor subtypes mediating functionally diverse postsynaptic inhibitory currents. Whilst the broad expression patterns of GABAAR subunits within the EC have been reported, those expressed by individual MEC LII cell types, in particular grid cells candidates, stellate and pyramidal cells, are less well described. Stellate and pyramidal cells are distinguished by their selective expression of reelin (RE+) and calbindin (CB+) respectively. Thus, the overall aim of this study was to provide a high resolution analysis of the major (α and γ) GABAAR subunits expressed in proximity to somato-dendritic PV+ boutons, on RE+ and CB+ cells, using immunohistochemistry, confocal microscopy and quantitative RT-PCR (qPCR). Clusters immunoreactive for the α1 and γ2 subunits decorated the somatic membranes of both RE+ and CB+ cells and were predominantly located in apposition to clusters immunoreactive for PV and vesicular GABA transporter (VGAT), suggesting expression in GABAergic synapses innervated by PV interneurons. Although intense α2 subunit-immunopositive clusters were evident in hippocampal fields located in close proximity to the EC, no specific signal was detected in MEC LII RE+ and CB+ profiles. Immunoreactivity for the α3 subunit was detected in all RE+ somata. In contrast, only a sub-population of CB+ cells was α3 immunopositive. These included CB-α3 cells which were both PV+ and PV-. Furthermore, α3 subunit mRNA and immunofluorescence decreased significantly between P 15 and P 25, a period implicated in the functional maturation of grid cells. Finally, α5 subunit immunoreactivity was detectable only on CB+ cells, not on RE+ cells. The present data demonstrates that physiologically distinct GABAAR subtypes are selectively expressed by CB+ and RE+ cells. This suggests that PV+ interneurons could utilize distinct postsynaptic signaling mechanisms to regulate the excitability of these different, candidate grid cell sub-populations.
RESUMEN
Grid cells in layer II of the medial entorhinal cortex (MEC LII) generate multiple regular firing fields in response to the position and speed of an individual within the environment. They exhibit a protracted postnatal development and, in the adult, show activity differences along the dorsoventral axis (DVA). Evidence suggests parvalbumin-positive (PV+) interneurons, most of which are perisomatic-targeting cells, play a crucial role in generation of the hexagonal grid cell activity pattern. We therefore hypothesized that the development and organization of PV+ perisomatic terminals in MEC LII reflect the postnatal emergence of the hexagonal firing pattern and dorsoventral differences seen in grid cell activity. We used immuno-electron microscopy to examine the development of PV+ perisomatic terminals and their target somata within dorsal and ventral MEC LII in rats of postnatal day (P)10, P15, and P30. We demonstrate that in dorsal and ventral MEC LII, the cross-sectional area of somata and number and density of perisomatic PV+ terminals increase between P10 and P15. A simultaneous decrease was observed in cross-sectional area of PV+ terminals. Between P15 and P30, both MEC regions showed an increase in PV+ terminal size and percentage of PV+ terminals containing mitochondria, which may enable grid cell activity to emerge and stabilize. We also report that dorsal somata are larger and apposed by more PV+ terminals than ventral somata at all stages, suggesting a protracted maturation in the ventral portion and a possible gradient in soma size and PV+ basket innervation along the DVA in the adult.
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Corteza Entorrinal/crecimiento & desarrollo , Parvalbúminas/metabolismo , Terminales Presinápticos/fisiología , Animales , Recuento de Células , Corteza Entorrinal/ultraestructura , Masculino , Terminales Presinápticos/ultraestructura , Ratas Long-EvansRESUMEN
Understanding what are the mechanisms that strengthen, stabilize and restrict synaptic innervation is a relevant topic in glutamate receptor delta 2 (GluRdelta2)-related research. It also involves targeting and selection of afferent input during formation of the neuronal circuitry in the cerebellar cortex and its functioning. This review will focus on the role of GluRdelta2, one of the main players in this field. Special emphasis will be placed on the processes that regulate the rapid translocation from climbing fibres to parallel fibres of GluRdelta2 and the role of GluRdelta2 in the reduction of supernumerary climbing fibre contacts on a single Purkinje cell. Furthermore, GluRdelta2 knockout mice show ataxia and impaired motor coordination, suggesting that the presence of GluRdelta2 plays an important role in controlling cerebellar functioning.
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Cerebelo/citología , Cerebelo/crecimiento & desarrollo , Plasticidad Neuronal/fisiología , Neuronas/fisiología , Receptores de Glutamato/fisiología , Animales , Ratones , Ratones NoqueadosRESUMEN
In this study the effects of hypergravity were analyzed on cerebellar Purkinje cells during early development in rats. The cerebellum is a key structure in the control and the adaptation of posture and anti-gravity activities. This holds particularly when external conditions are modified. Three groups of rats were conceived, born and reared in hypergravity (2g). At postnatal day 5 (P5), P10 or P15, they were exposed to normal gravity and at P40, the cerebella were investigated on the expression of calbindin-D28k and inositol-3-phosphate (IP3) in Purkinje cells. Control animals were bred in the same conditions but at 1g. Immunoreactivity of Purkinje cells was studied in lobules III and IX of the vermis. Lobule IX of the vermis is one of the targets of primary otolithic vestibular projections, and lobule III served as a control, being much less related with vestibular inputs. The results show that hypergravity induces a decrease in calbindin and IP3 labeling in 20% of Purkinje cells of lobule IX without any change in lobule III. Animals transferred from 2g to 1g at P5 or P10 showed the most pronounced effects and much less at P15. This study demonstrates that early development of the cerebellum is highly sensitive to changes in gravity. Ages until P10 are critical for the development of vestibulo-cerebellar connections, and in particularly the calcium signaling in Purkinje cells.
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Hipergravedad/efectos adversos , Fosfatos de Inositol/biosíntesis , Células de Purkinje/metabolismo , Células de Purkinje/fisiología , Proteína G de Unión al Calcio S100/biosíntesis , Envejecimiento/fisiología , Animales , Conducta Animal/fisiología , Calbindina 1 , Calbindinas , Células Cultivadas , Centrifugación , Cerebelo/citología , Cerebelo/crecimiento & desarrollo , Femenino , Inmunohistoquímica , Masculino , RatasRESUMEN
To investigate whether the type of programmed cell death of myelodysplastic erythroid cells depends on their cellular context, we performed studies on cells from patients with low-risk myelodysplastic syndromes. We compared erythroid cells (and their precursor cells) from the mononuclear cell fraction with those from the hematon fraction, which are compacted complexes of hematopoietic cells surrounded by their own micro-environment. In directly fixed materials, erythroblasts exhibited signs of autophagy with limited apoptosis (<3%) based on ultrastructural characteristics and immunogold labeling for activated caspase-3. After 24 h in culture, myelodysplastic erythroblasts exhibited a significant increase in apoptosis (22 ± 7% vs. 3 ± 2%, p = 0.001). In contrast, the myelodysplastic erythroblasts from the hematon fraction did not exhibit an increased tendency toward apoptosis after culture (7 ± 3.3% vs. 1.8 ± 2.3%), which was in line with results for normal bone marrow cells. The same dependency on the micro-environment was noted for immature erythroid progenitor cells. Myelodysplastic hematons exhibited distinct numbers of erythroid burst-forming units in association with an extensive network of stromal cells, whereas small numbers of erythroid burst-forming units were generated from the myelodysplastic mononuclear cells compared with normal mononuclear cells (10.2 ± 9 vs. 162 ± 125, p < 0.001). Co-culture of erythroid myelodysplastic cells in the presence of growth factors (vascular endothelial growth factor, leukemia inhibitory factor) or on the MS-5 stromal layer did not restore the expansion of erythroid precursor cells. These data indicate that surviving myelodysplastic erythroid progenitors become more vulnerable to programmed cell death when they are detached from their own micro-environment.
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Células Precursoras Eritroides/fisiología , Síndromes Mielodisplásicos/fisiopatología , Microambiente Tumoral , Anciano , Anciano de 80 o más Años , Apoptosis , Supervivencia Celular , Células Cultivadas , Células Precursoras Eritroides/patología , Femenino , Citometría de Flujo , Humanos , Leucocitos Mononucleares/patología , Masculino , Persona de Mediana Edad , Reacción en Cadena en Tiempo Real de la Polimerasa , Factores de RiesgoRESUMEN
Lipid droplets (LDs) hypertrophy in adipocytes is the main cause of energy metabolic system dysfunction, obesity and its afflictions such as T2D. However, the role of adipocytes in linking energy metabolic disorders with insulin regulation is unknown in humans. Human adipocytes constitutively synthesize and secrete insulin, which is biologically functional. Insulin concentrations and release are fat mass- and LDs-dependent respectively. Fat reduction mediated by bariatric surgery repairs obesity-associated T2D. The expression of genes, like PCSK1 (proinsulin conversion enzyme), GCG (Glucagon), GPLD1, CD38 and NNAT, involved in insulin regulation/release were differentially expressed in pancreas and adipose tissue (AT). INS (insulin) and GCG expression reduced in human AT-T2D as compared to AT-control, but remained unchanged in pancreas in either state. Insulin levels (mRNA/protein) were higher in AT derived from prediabetes BB rats with destructed pancreatic ß-cells and controls than pancreas derived from the same rats respectively. Insulin expression in 10 human primary cell types including adipocytes and macrophages is an evidence for extrapancreatic insulin-producing cells. The data suggest a crosstalk between AT and pancreas to fine-tune energy metabolic system or may minimize the metabolic damage during diabetes. This study opens new avenues towards T2D therapy with a great impact on public health.
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Adipocitos/metabolismo , Insulina/metabolismo , Gotas Lipídicas/metabolismo , Adipocitos/citología , Adipocitos/patología , Animales , Biomarcadores , Diferenciación Celular/genética , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/metabolismo , Perfilación de la Expresión Génica , Humanos , Insulina/genética , Gotas Lipídicas/patología , Transporte de Proteínas , ARN Mensajero/genética , RatasRESUMEN
Urocortin belongs to the family of corticotropin-releasing factor (CRF)-like peptides, which play an important role in sensorimotor coordination. CRF induces locomotor activity, and urocortin has an inhibitory effect. Here, we document the regional and subcellular localization of urocortin in the developing rat cerebellum to compare it with CRF. During the first postnatal week, urocortin immunoreactivity (UCN-ir), within the white matter and cerebellar cortex, was strongest in vermal lobules I, II, IX, and X, closely followed by lobules IV, V, and VIII; lobules VI and VII showed the weakest labeling. Cortical immunoreactivity was in the form of puncta that encircled Purkinje cell somata. By postnatal day (PD) 12, UCN-ir had increased appreciably in all lobules. In Purkinje cells, labeling was spread throughout their somata and proximal dendrites. By PD 15, labeling in lobules I-IV appeared to wane, yet still prevailed in the central and posterior lobules. This anterior-to-posterior gradient persisted through to adulthood. The study shows that urocortin and CRF have similar regional distribution profiles during development, suggesting synergistic roles within the vestibulocerebellum. The onset of the adult distributional pattern of urocortin at the stage when rats are capable of fluent walking patterns further strengthens the correlation between CRF-like peptides and postural control. An important difference between urocortin and CRF is the localization of urocortin, and not CRF, within Purkinje cells, implying that urocortin probably has an additional role in modulating the signals emanating from the cerebellar cortex to the deep cerebellar nuclei.