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1.
Neurol Ther ; 11(3): 981-1042, 2022 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-35610531

RESUMEN

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) resulting in demyelination and neurodegeneration. The therapeutic strategy is now largely based on reducing inflammation with immunosuppressive drugs. Unfortunately, when disease progression is observed, no drug offers neuroprotection apart from its anti-inflammatory effect. In this review, we explore current knowledge on the assessment of neurodegeneration in MS and look at putative targets that might prove useful in protecting the axon from degeneration. Among them, Bruton's tyrosine kinase inhibitors, anti-apoptotic and antioxidant agents, sex hormones, statins, channel blockers, growth factors, and molecules preventing glutamate excitotoxicity have already been studied. Some of them have reached phase III clinical trials and carry a great message of hope for our patients with MS.

2.
Cell Mol Life Sci ; 78(13): 5257-5273, 2021 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-34019104

RESUMEN

In demyelinated lesions, astrocytes, activated microglia and infiltrating macrophages secrete several factors regulating oligodendrocyte precursor cells' behaviour. What appears to be the initiation of an intrinsic mechanism of myelin repair is only leading to partial recovery and inefficient remyelination, a process worsening over the course of the disease. This failure is largely due to the concomitant accumulation of inhibitory cues in and around the lesion sites opposing to growth promoting factors. Here starts a complex game of interactions between the signalling pathways controlling oligodendrocytes migration or differentiation. Receptors of positive or negative cues are modulating Ras, PI3K or RhoGTPases pathways acting on oligodendrocyte cytoskeleton remodelling. From the description of this intricate signalling network, this review addresses the extent to which the modulation of the global response to inhibitory cues may pave the route towards novel therapeutic approaches for myelin repair.


Asunto(s)
Diferenciación Celular , Esclerosis Múltiple/terapia , Oligodendroglía/citología , Regeneración , Remielinización , Animales , Humanos , Oligodendroglía/fisiología
3.
Cells ; 10(3)2021 03 21.
Artículo en Inglés | MEDLINE | ID: mdl-33801063

RESUMEN

Microglial cells are key players in neural pathogenesis and microglial function regulation appears to be pivotal in controlling neuroinflammatory/neurological diseases. Here, we investigated the effects and mechanism of action of neurosteroid allopregnanolone (ALLO) on murine microglial BV-2 cells and primary microglia in order to determine ALLO-induced immunomodulatory potential and to provide new insights for the development of both natural and safe neuroprotective strategies targeting microglia. Indeed, ALLO-treatment is increasingly suggested as beneficial in various models of neurological disorders but the underlying mechanisms have not been elucidated. Therefore, the microglial cells were cultured with various serum concentrations to mimic the blood-brain-barrier rupture and to induce their activation. Proliferation, viability, RT-qPCR, phagocytosis, and morphology analyzes, as well as migration with time-lapse imaging and quantitative morphodynamic methods, were combined to investigate ALLO actions on microglia. BV-2 cells express subunits of GABA-A receptor that mediates ALLO activity. ALLO (10µM) induced microglial cell process extension and decreased migratory capacity. Interestingly, ALLO modulated the phagocytic activity of BV-2 cells and primary microglia. Our results, which show a direct effect of ALLO on microglial morphology and phagocytic function, suggest that the natural neurosteroid-based approach may contribute to developing effective strategies against neurological disorders that are evoked by microglia-related abnormalities.


Asunto(s)
Forma de la Célula , Microglía/citología , Microglía/metabolismo , Neuroprotección , Neuroesteroides/metabolismo , Fagocitosis , Pregnanolona/metabolismo , Animales , Animales Recién Nacidos , Diferenciación Celular/efectos de los fármacos , Línea Celular , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Forma de la Célula/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Ratones Endogámicos C57BL , Microglía/efectos de los fármacos , Modelos Biológicos , Neuroprotección/efectos de los fármacos , Oligodendroglía/efectos de los fármacos , Oligodendroglía/metabolismo , Fagocitosis/efectos de los fármacos , Pregnanolona/química , Subunidades de Proteína/genética , Subunidades de Proteína/metabolismo , Receptores de GABA/genética , Receptores de GABA/metabolismo , Suero
4.
EMBO Mol Med ; 11(11): e10378, 2019 11 07.
Artículo en Inglés | MEDLINE | ID: mdl-31566924

RESUMEN

Current treatments in multiple sclerosis (MS) are modulating the inflammatory component of the disease, but no drugs are currently available to repair lesions. Our study identifies in MS patients the overexpression of Plexin-A1, the signalling receptor of the oligodendrocyte inhibitor Semaphorin 3A. Using a novel type of peptidic antagonist, we showed the possibility to counteract the Sema3A inhibitory effect on oligodendrocyte migration and differentiation in vitro when antagonizing Plexin-A1. The use of this compound in vivo demonstrated a myelin protective effect as shown with DTI-MRI and confirmed at the histological level in the mouse cuprizone model of induced demyelination/remyelination. This effect correlated with locomotor performances fully preserved in chronically treated animals. The administration of the peptide also showed protective effects, leading to a reduced severity of demyelination in the context of experimental autoimmune encephalitis (EAE). Hence, the disruption of the inhibitory microenvironmental molecular barriers allows normal myelinating cells to exert their spontaneous remyelinating capacity. This opens unprecedented therapeutic opportunity for patients suffering a disease for which no curative options are yet available.


Asunto(s)
Esclerosis Múltiple/fisiopatología , Proteínas del Tejido Nervioso/metabolismo , Oligodendroglía/fisiología , Receptores de Superficie Celular/metabolismo , Remielinización , Semaforina-3A/metabolismo , Transducción de Señal , Animales , Encéfalo/diagnóstico por imagen , Línea Celular , Movimiento Celular , Proliferación Celular , Modelos Animales de Enfermedad , Imagen por Resonancia Magnética , Ratones Endogámicos C57BL , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Receptores de Superficie Celular/antagonistas & inhibidores
5.
Curr Biol ; 29(17): 2852-2866.e5, 2019 09 09.
Artículo en Inglés | MEDLINE | ID: mdl-31422887

RESUMEN

The spatiotemporal coordination of actin regulators in the lamellipodium determines the dynamics and architecture of branched F-actin networks during cell migration. The WAVE regulatory complex (WRC), an effector of Rac1 during cell protrusion, is concentrated at the lamellipodium tip. Thus, activated Rac1 should operate at this location to activate WRC and trigger membrane protrusion. Yet correlation of Rho GTPase activation with cycles of membrane protrusion previously revealed complex spatiotemporal patterns of Rac1 and RhoA activation in the lamellipodium. Combining single protein tracking (SPT) and super-resolution imaging with loss- or gain-of-function mutants of Rho GTPases, we show that Rac1 immobilizations at the lamellipodium tip correlate with its activation, in contrast to RhoA. Using Rac1 effector loop mutants and wild-type versus mutant variants of WRC, we show that selective immobilizations of activated Rac1 at the lamellipodium tip depend on effector binding, including WRC. In contrast, wild-type Rac1 only displays slower diffusion at the lamellipodium tip, suggesting transient activations. Local optogenetic activation of Rac1, triggered by membrane recruitment of Tiam1, shows that Rac1 activation must occur close to the lamellipodium tip and not behind the lamellipodium to trigger efficient membrane protrusion. However, coupling tracking with optogenetic activation of Rac1 demonstrates that diffusive properties of wild-type Rac1 are unchanged despite enhanced lamellipodium protrusion. Taken together, our results support a model whereby transient activations of Rac1 occurring close to the lamellipodium tip trigger WRC binding. This short-lived activation ensures a local and rapid control of Rac1 actions on its effectors to trigger actin-based protrusion.


Asunto(s)
Movimiento Celular , Extensiones de la Superficie Celular/metabolismo , Fibroblastos/metabolismo , Neuropéptidos/metabolismo , Seudópodos/metabolismo , Proteína de Unión al GTP rac1/metabolismo , Animales , Embrión de Mamíferos/metabolismo , Ratones , Proteína de Unión al GTP rhoA/metabolismo
6.
Small GTPases ; 10(2): 99-110, 2019 03.
Artículo en Inglés | MEDLINE | ID: mdl-28287334

RESUMEN

Cell migration, a key feature of embryonic development, immunity, angiogenesis, and tumor metastasis, is based on the coordinated regulation of actin dynamics and integrin-mediated adhesion. Rho GTPases play a major role in this phenomenon by regulating the onset and maintenance of actin-based protruding structures at cell leading edges (i.e. lamellipodia and filopodia) and contractile structures (i.e., stress fibers) at their trailing edge. While spatio-temporal analysis demonstrated the tight regulation of Rho GTPases at the migration front during cell locomotion, little is known about how the main regulators of Rho GTPase activity, such as GAPs, GEFs and GDIs, play a role in this process. In this review, we focus on a major negative regulator of RhoA, p190RhoGAP-A and its close isoform p190RhoGAP-B, which are necessary for efficient cell migration. Recent studies, including our, demonstrated that p190RhoGAP-A localization and activity undergo a complex regulatory mechanism, accounting for the tight regulation of RhoA, but also other members of the Rho GTPase family, at the cell periphery.


Asunto(s)
Movimiento Celular , Proteínas Activadoras de GTPasa/química , Proteínas Activadoras de GTPasa/metabolismo , Proteínas de Unión al GTP rho/metabolismo , Animales , Humanos , Isoenzimas/metabolismo
7.
J Cell Biol ; 214(7): 859-73, 2016 09 26.
Artículo en Inglés | MEDLINE | ID: mdl-27646271

RESUMEN

Spatiotemporal regulation of RhoGTPases such as RhoA is required at the cell leading edge to achieve cell migration. p190RhoGAP (p190A) is the main negative regulator of RhoA and localizes to membrane protrusions, where its GTPase-activating protein (GAP) activity is required for directional migration. In this study, we investigated the molecular processes responsible for p190A targeting to actin protrusions. By analyzing the subcellular localization of truncated versions of p190A in hepatocellular carcinoma cells, we identified a novel functional p190A domain: the protrusion localization sequence (PLS) necessary and sufficient for p190A targeting to leading edges. Interestingly, the PLS is also required for the negative regulation of p190A RhoGAP activity. Further, we show that the F-actin binding protein cortactin binds the PLS and is required for p190A targeting to protrusions. Lastly, we demonstrate that cancer-associated mutations in PLS affect p190A localization and function, as well as tumor cell migration. Altogether, our data unveil a new mechanism of regulation of p190A in migrating tumor cells.


Asunto(s)
Movimiento Celular , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/metabolismo , Mutación/genética , Neoplasias/genética , Neoplasias/patología , Actinas/metabolismo , Animales , Línea Celular Tumoral , Extensiones de la Superficie Celular/metabolismo , Cortactina/metabolismo , Factores de Intercambio de Guanina Nucleótido/química , Humanos , Ratones , Unión Proteica , Dominios Proteicos , Seudópodos/metabolismo , Proteínas Represoras , Sus scrofa , Transfección
8.
PLoS One ; 10(9): e0137311, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-26340347

RESUMEN

The NG2 proteoglycan is characteristically expressed by oligodendrocyte progenitor cells (OPC) and also by aggressive brain tumours highly resistant to chemo- and radiation therapy. Oligodendrocyte-lineage cells are particularly sensitive to stress resulting in cell death in white matter after hypoxic or ischemic insults of premature infants and destruction of OPC in some types of Multiple Sclerosis lesions. Here we show that the NG2 proteoglycan binds OMI/HtrA2, a mitochondrial serine protease which is released from damaged mitochondria into the cytosol in response to stress. In the cytosol, OMI/HtrA2 initiates apoptosis by proteolytic degradation of anti-apoptotic factors. OPC in which NG2 has been downregulated by siRNA, or OPC from the NG2-knockout mouse show an increased sensitivity to oxidative stress evidenced by increased cell death. The proapoptotic protease activity of OMI/HtrA2 in the cytosol can be reduced by the interaction with NG2. Human glioma expressing high levels of NG2 are less sensitive to oxidative stress than those with lower NG2 expression and reducing NG2 expression by siRNA increases cell death in response to oxidative stress. Binding of NG2 to OMI/HtrA2 may thus help protect cells against oxidative stress-induced cell death. This interaction is likely to contribute to the high chemo- and radioresistance of glioma.


Asunto(s)
Antígenos/metabolismo , Neoplasias Encefálicas/metabolismo , Regulación Neoplásica de la Expresión Génica , Glioblastoma/metabolismo , Proteínas Mitocondriales/metabolismo , Proteoglicanos/metabolismo , Serina Endopeptidasas/metabolismo , Animales , Anticuerpos Neutralizantes/farmacología , Antígenos/genética , Apoptosis/efectos de los fármacos , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Cerebelo/efectos de los fármacos , Cerebelo/metabolismo , Cerebelo/patología , Citosol/efectos de los fármacos , Citosol/metabolismo , Glioblastoma/genética , Glioblastoma/patología , Serina Peptidasa A2 que Requiere Temperaturas Altas , Humanos , Peróxido de Hidrógeno/farmacología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Proteínas Mitocondriales/genética , Estrés Oxidativo , Cultivo Primario de Células , Unión Proteica , Proteoglicanos/antagonistas & inhibidores , Proteoglicanos/genética , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Serina Endopeptidasas/genética , Transducción de Señal
9.
Acta Neuropathol ; 129(2): 279-95, 2015 02.
Artículo en Inglés | MEDLINE | ID: mdl-25500713

RESUMEN

The contribution of microglia to ischemic cortical stroke is of particular therapeutic interest because of the impact on the survival of brain tissue in the ischemic penumbra, a region that is potentially salvable upon a brain infarct. Whether or not tissue in the penumbra survives critically depends on blood flow and vessel perfusion. To study the role of microglia in cortical stroke and blood vessel stability, CX3CR1(+/GFP) mice were subjected to transient middle cerebral artery occlusion and then microglia were investigated using time-lapse two-photon microscopy in vivo. Soon after reperfusion, microglia became activated in the stroke penumbra and started to expand cellular protrusions towards adjacent blood vessels. All microglia in the penumbra were found associated with blood vessels within 24 h post reperfusion and partially fully engulfed them. In the same time frame blood vessels became permissive for blood serum components. Migration assays in vitro showed that blood serum proteins leaking into the tissue provided molecular cues leading to the recruitment of microglia to blood vessels and to their activation. Subsequently, these perivascular microglia started to eat up endothelial cells by phagocytosis, which caused an activation of the local endothelium and contributed to the disintegration of blood vessels with an eventual break down of the blood brain barrier. Loss-of-microglia-function studies using CX3CR1(GFP/GFP) mice displayed a decrease in stroke size and a reduction in the extravasation of contrast agent into the brain penumbra as measured by MRI. Potentially, medication directed at inhibiting microglia activation within the first day after stroke could stabilize blood vessels in the penumbra, increase blood flow, and serve as a valuable treatment for patients suffering from ischemic stroke.


Asunto(s)
Isquemia Encefálica/fisiopatología , Encéfalo/irrigación sanguínea , Encéfalo/fisiopatología , Microglía/fisiología , Accidente Cerebrovascular/fisiopatología , Animales , Barrera Hematoencefálica/patología , Barrera Hematoencefálica/fisiopatología , Encéfalo/patología , Isquemia Encefálica/patología , Receptor 1 de Quimiocinas CX3C , Línea Celular , Modelos Animales de Enfermedad , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Transgénicos , Microglía/patología , Fagocitosis/fisiología , Receptores de Quimiocina/genética , Receptores de Quimiocina/metabolismo , Accidente Cerebrovascular/patología
10.
PLoS Biol ; 12(11): e1001993, 2014 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-25387269

RESUMEN

The role of glia in modulating neuronal network activity is an important question. Oligodendrocyte precursor cells (OPC) characteristically express the transmembrane proteoglycan nerve-glia antigen 2 (NG2) and are unique glial cells receiving synaptic input from neurons. The development of NG2+ OPC into myelinating oligodendrocytes has been well studied, yet the retention of a large population of synapse-bearing OPC in the adult brain poses the question as to additional functional roles of OPC in the neuronal network. Here we report that activity-dependent processing of NG2 by OPC-expressed secretases functionally regulates the neuronal network. NG2 cleavage by the α-secretase ADAM10 yields an ectodomain present in the extracellular matrix and a C-terminal fragment that is subsequently further processed by the γ-secretase to release an intracellular domain. ADAM10-dependent NG2 ectodomain cleavage and release (shedding) in acute brain slices or isolated OPC is increased by distinct activity-increasing stimuli. Lack of NG2 expression in OPC (NG2-knockout mice), or pharmacological inhibition of NG2 ectodomain shedding in wild-type OPC, results in a striking reduction of N-methyl-D-aspartate (NMDA) receptor-dependent long-term potentiation (LTP) in pyramidal neurons of the somatosensory cortex and alterations in the subunit composition of their α-amino-3-hydroxy-5-methyl-4-isoxazolepr opionicacid (AMPA) receptors. In NG2-knockout mice these neurons exhibit diminished AMPA and NMDA receptor-dependent current amplitudes; strikingly AMPA receptor currents can be rescued by application of conserved LNS protein domains of the NG2 ectodomain. Furthermore, NG2-knockout mice exhibit altered behavior in tests measuring sensorimotor function. These results demonstrate for the first time a bidirectional cross-talk between OPC and the surrounding neuronal network and demonstrate a novel physiological role for OPC in regulating information processing at neuronal synapses.


Asunto(s)
Proteínas ADAM/metabolismo , Secretasas de la Proteína Precursora del Amiloide/metabolismo , Antígenos/metabolismo , Proteínas de la Membrana/metabolismo , Neuronas/metabolismo , Oligodendroglía/fisiología , Proteoglicanos/metabolismo , Proteína ADAM10 , Animales , Línea Celular , Matriz Extracelular/metabolismo , Potenciación a Largo Plazo , Masculino , Ratones , Ratones Noqueados , Plasticidad Neuronal , Estructura Terciaria de Proteína , Células Piramidales/metabolismo , Receptores de Glutamato/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Filtrado Sensorial , Sinapsis/metabolismo
11.
PLoS One ; 9(2): e89423, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-24586768

RESUMEN

Oligodendrocytes are the myelinating glial cells of the central nervous system. In the course of brain development, oligodendrocyte precursor cells migrate, scan the environment and differentiate into mature oligodendrocytes with multiple cellular processes which recognize and ensheath neuronal axons. During differentiation, oligodendrocytes undergo dramatic morphological changes requiring cytoskeletal rearrangements which need to be tightly regulated. The non-receptor tyrosine kinase Fyn plays a central role in oligodendrocyte differentiation and myelination. In order to improve our understanding of the role of oligodendroglial Fyn kinase, we have identified Fyn targets in these cells. Purification and mass-spectrometric analysis of tyrosine-phosphorylated proteins in response to overexpressed active Fyn in the oligodendrocyte precursor cell line Oli-neu, yielded the adaptor molecule p130Cas. We analyzed the function of this Fyn target in oligodendroglial cells and observed that reduction of p130Cas levels by siRNA affects process outgrowth, the thickness of cellular processes and migration behavior of Oli-neu cells. Furthermore, long term p130Cas reduction results in decreased cell numbers as a result of increased apoptosis in cultured primary oligodendrocytes. Our data contribute to understanding the molecular events taking place during oligodendrocyte migration and morphological differentiation and have implications for myelin formation.


Asunto(s)
Movimiento Celular/fisiología , Supervivencia Celular/fisiología , Proteína Sustrato Asociada a CrK/metabolismo , Oligodendroglía/metabolismo , Proteínas Proto-Oncogénicas c-fyn/metabolismo , Animales , Axones/metabolismo , Células Cultivadas , Ratones , Oligodendroglía/citología , Fosforilación
12.
Mol Neurobiol ; 50(2): 482-93, 2014 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-24390567

RESUMEN

Resident progenitor cells expressing nerve/glial antigen 2 (NG2) such as oligodendrocyte precursor cells (OPC) and pericytes persist in the adult brain. The transmembrane proteoglycan NG2 regulates migration of both these cell types in response to growth factors or specific components of the extracellular matrix. This role of NG2 is linked to the control of cell polarity. The polarization of OPC toward an acute lesion in the brain is impaired in NG2-deficient mice, supporting this concept. A review of the signaling pathways impinged on by NG2 reveals key proteins of cell polarity: phosphatidylinositol 3-kinase, focal adhesion kinase, Rho GTPases, and polarity complex proteins. In the scope of cell migration, I discuss here how the interplay of NG2 with signaling transmitted by extracellular cues can control the establishment of cell polarity, and I propose a model to integrate the apparent opposite effects of NG2 on cellular dynamics.


Asunto(s)
Antígenos/metabolismo , Movimiento Celular/fisiología , Polaridad Celular/fisiología , Oligodendroglía/citología , Proteoglicanos/metabolismo , Animales , Humanos , Oligodendroglía/metabolismo , Transducción de Señal/fisiología , Células Madre/citología
13.
J Neurosci ; 33(26): 10858-74, 2013 Jun 26.
Artículo en Inglés | MEDLINE | ID: mdl-23804106

RESUMEN

The transmembrane proteoglycan NG2 is expressed by oligodendrocyte precursor cells (OPC), which migrate to axons during developmental myelination and remyelinate in the adult after migration to injured sites. Highly invasive glial tumors also express NG2. Despite the fact that NG2 has been implicated in control of OPC migration, its mode of action remains unknown. Here, we show in vitro and in vivo that NG2 controls migration of OPC through the regulation of cell polarity. In stab wounds in adult mice we show that NG2 controls orientation of OPC toward the wound. NG2 stimulates RhoA activity at the cell periphery via the MUPP1/Syx1 signaling pathway, which favors the bipolar shape of migrating OPC and thus directional migration. Upon phosphorylation of Thr-2256, downstream signaling of NG2 switches from RhoA to Rac stimulation. This triggers process outgrowth through regulators of front-rear polarity and we show using a phospho-mimetic form of NG2 that indeed NG2 recruits proteins of the CRB and the PAR polarity complexes to stimulate Rac activity via the GEF Tiam1. Our findings demonstrate that NG2 is a core organizer of Rho GTPase activity and localization in the cell, which controls OPC polarity and directional migration. This work also reveals CRB and PAR polarity complexes as new effectors of NG2 signaling in the establishment of front-rear polarity.


Asunto(s)
Antígenos/fisiología , Movimiento Celular/fisiología , Polaridad Celular/fisiología , Proteínas del Tejido Nervioso/fisiología , Oligodendroglía/fisiología , Proteoglicanos/fisiología , Proteínas de Unión al GTP rho/fisiología , Antígenos/genética , Movimiento Celular/genética , Forma de la Célula/genética , Forma de la Célula/fisiología , Quimiotaxis/fisiología , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/fisiología , Humanos , Fosforilación , Proteoglicanos/genética , ARN/biosíntesis , ARN/genética , Interferencia de ARN , Transducción de Señal/genética , Transducción de Señal/fisiología , Células Madre , Proteína 1 de Invasión e Inducción de Metástasis del Linfoma-T , Treonina/metabolismo , Proteínas de Uniones Estrechas/genética , Proteínas de Uniones Estrechas/fisiología , Proteínas de Unión al GTP rac/metabolismo
14.
J Cell Physiol ; 225(1): 186-95, 2010 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-20458741

RESUMEN

Cellular differentiation relies on both physical and chemical environmental cues. The bipotential mouse embryonic liver (BMEL) cells are early progenitors of liver epithelial cells with an apparently infinite proliferative potential. These cells, which remain undifferentiated in a monolayer culture, differentiate upon release from geometrical constraints imposed by growth on a stiff plastic plate. In a complex three dimensional environment of a Matrigel extracellular matrix, BMEL cells form two types of polarized organoids of distinct morphologies: cyst-like structures suggesting cholangiocyte-type organization or complex organoids, reminiscent of liver parenchyma and associated with acquisition of hepatocyte-specific phenotypic markers. The choice of the in vitro differentiation lineage is governed by Transforming Growth Factor-beta (TGF-beta) signaling. Our results suggest that morphological cues initiate the differentiation of early hepatic precursors and confirm the inhibitory role of TGF-beta on hepatocytic lineage differentiation.


Asunto(s)
Diferenciación Celular/fisiología , Linaje de la Célula , Forma de la Célula , Hígado/citología , Transducción de Señal/fisiología , Células Madre/citología , Factor de Crecimiento Transformador beta/metabolismo , Animales , Biomarcadores/metabolismo , Células Cultivadas , Colágeno/metabolismo , Combinación de Medicamentos , Humanos , Laminina/metabolismo , Hígado/fisiología , Ratones , Organoides/citología , Organoides/metabolismo , Proteoglicanos/metabolismo , Células Madre/fisiología , Técnicas de Cultivo de Tejidos , Factor de Crecimiento Transformador beta/genética
15.
Mol Biosyst ; 6(4): 648-61, 2010 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-20237642

RESUMEN

Movement of individual cells and of cellular cohorts, chains or sheets requires physical forces that are established through interactions of cells with their environment. In vivo, migration occurs extensively during embryonic development and in adults during wound healing and tumorigenesis. In order to identify the molecular events involved in cell movement, in vitro systems have been developed. These have contributed to the definition of a number of molecular pathways put into play in the course of migratory behaviours, such as mesenchymal and amoeboid movement. More recently, our knowledge of migratory modes has been enriched by analyses of cells exploring and moving through three-dimensional (3D) matrices. While the cells' morphologies differ in 2D and 3D environments, the basic mechanisms that put a cellular body into motion are remarkably similar. Thus, in both 2D and 3D, the polarity of the migrating cell is initially defined by a specific subcellular localization of signalling molecules and components of molecular machines required for motion. While the polarization can be initiated either in response to extracellular signalling or be a chance occurrence, it is reinforced and sustained by positive feedback loops of signalling molecules. Second, adhesion to a substratum is necessary to generate forces that will propel the cell engaged in either mesenchymal or ameboid migration. For collective cell movement, intercellular coordination constitutes an additional requirement: a cell cohort remains stationary if individual cells pull in opposite directions. Finally, the availability of space to move into is a general requirement to set cells into motion. Lack of free space is probably the main obstacle for migration of most healthy cells in an adult multicellular organism. Thus, the requirements for cell movement are both intrinsic to the cell, involving coordinated signalling and interactions with molecular machines, and extrinsic, imposed by the physicochemical nature of the environment. In particular, the geometry and stiffness of the support act on a range of signalling pathways that induce specific cell migratory responses. These issues are discussed in the present review in the context of published work and our own data on collective migration of hepatocyte cohorts.


Asunto(s)
Movimiento Celular/fisiología , Animales , Adhesión Celular/fisiología , Polaridad Celular/fisiología , Hepatocitos/fisiología , Humanos , Imagenología Tridimensional , Técnicas In Vitro , Mecanotransducción Celular , Modelos Biológicos , Transducción de Señal , Biología de Sistemas
16.
Hepatology ; 50(5): 1370-9, 2009 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-19711428

RESUMEN

UNLABELLED: An unresolved question regarding the physiopathology of hepatitis C virus (HCV) infection is the remarkable efficiency with which host defenses are neutralized to establish chronic infection. Modulation of an apoptotic response is one strategy used by viruses to escape immune surveillance. We previously showed that HCV proteins down-regulate expression of BH3-only Bcl2 interacting domain (Bid) in hepatocytes of HCV transgenic mice. As a consequence, cells acquire resistance to Fas-mediated apoptosis, which in turn leads to increased persistence of experimental viral infections in vivo. This mechanism might participate in the establishment of chronic infections and the resulting pathologies, including hepatocellular carcinoma. We now report that Bid is also down-regulated in patients in the context of noncirrhotic HCV-linked tumorigenesis and in the HCV RNA replicon system. We show that the nonstructural HCV viral protein NS5A is sufficient to activate a calpain cysteine protease, leading to degradation of Bid. Moreover, pharmacological inhibitors of calpains restore both the physiological levels of Bid and the sensitivity of cells toward a death receptor-mediated apoptotic signal. Finally, human HCV-related tumors and hepatocytes from HCV transgenic mice that display low Bid expression contain activated calpains. CONCLUSION: Calpains activated by HCV proteins degrade Bid and thus dampen apoptotic signaling. These results suggest that inhibiting calpains could lead to an improved efficiency of immune-mediated elimination of HCV-infected cells.


Asunto(s)
Apoptosis/fisiología , Calpaína/metabolismo , Carcinoma Hepatocelular/patología , Hepacivirus/metabolismo , Neoplasias Hepáticas/patología , Transducción de Señal/fisiología , Proteínas Virales/metabolismo , Adulto , Anciano , Animales , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/metabolismo , Carcinoma Hepatocelular/metabolismo , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Hepatitis C Crónica/patología , Hepatitis C Crónica/fisiopatología , Humanos , Hígado/metabolismo , Hígado/patología , Neoplasias Hepáticas/metabolismo , Masculino , Ratones , Ratones Transgénicos , Persona de Mediana Edad , Replicón/fisiología , Proteínas no Estructurales Virales/metabolismo
17.
Mol Biol Cell ; 19(3): 945-56, 2008 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-18094041

RESUMEN

Transforming growth factor beta (TGF-beta) has a strong impact on liver development and physiopathology, exercised through its pleiotropic effects on growth, differentiation, survival, and migration. When exposed to TGF-beta, the mhAT3F cells, immortalized, highly differentiated hepatocytes, maintained their epithelial morphology and underwent dramatic alterations of adhesion, leading to partial or complete detachment from a culture plate, followed by readhesion and spreading. These alterations of adhesive behavior were caused by sequential changes in expression of the alpha5beta1 integrin and of its ligand, the fibronectin. The altered specificity of anchorage to the extracellular matrix gave rise to changes in cells' collective motility: cohorts adhering to fibronectin maintained a persistent, directional motility, with ezrin-rich pathfinder cells protruding from the tips of the cohorts. The absence of adhesion to fibronectin prevented the appearance of polarized pathfinders and lead to random, oscillatory motility. Our data suggest a novel role for TGF-beta in the control of collective migration of epithelial cohorts.


Asunto(s)
Movimiento Celular/efectos de los fármacos , Fibronectinas/metabolismo , Hepatocitos/citología , Hepatocitos/efectos de los fármacos , Factor de Crecimiento Transformador beta/farmacología , Animales , Apoptosis/efectos de los fármacos , Adhesión Celular/efectos de los fármacos , Comunicación Celular/efectos de los fármacos , Línea Celular , Membrana Celular/efectos de los fármacos , Membrana Celular/metabolismo , Extensiones de la Superficie Celular/efectos de los fármacos , Extensiones de la Superficie Celular/metabolismo , Epitelio/efectos de los fármacos , Perfilación de la Expresión Génica , Regulación de la Expresión Génica/efectos de los fármacos , Hepatocitos/metabolismo , Humanos , Integrina alfa5/genética , Integrina alfa5/metabolismo , Mesodermo/efectos de los fármacos , Ratones , Subunidades de Proteína/metabolismo , Proteína de Unión al GTP rac1/metabolismo
18.
Neurobiol Aging ; 28(7): 1088-98, 2007 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-16769156

RESUMEN

Brain-derived neurotrophic factor (BDNF) is a key regulator of neuronal plasticity in adult rat brain and its effects are mediated through TrkB receptors. BDNF and its receptors are also localized in the pituitary, but their expressions throughout the rat lifespan are poorly known. Here we analyzed levels of BDNF and the different subtypes of TrkB receptors (mRNA and proteins) in the rat pituitary at different stages of life. BDNF immunoreactivity was expressed in folliculo-stellate cells from the anterior pituitary and in the intermediate lobe. TrkB.FL and TrkB.T1 receptors were strongly and essentially expressed in the intermediate lobe similar to the alpha-MSH localization pattern. These receptors begun decreasing at middle-age but TrkB.T2 was not detected in the pituitary at any age. Finally, in vitro alpha-MSH release from the intermediate lobe was correlated with the receptor content throughout the lifespan. The present results demonstrate the presence of BDNF in folliculo-stellate cells and indicated that receptors, rather than BDNF itself, are impaired with aging. These changes can contribute to explain age-dependent endocrine changes.


Asunto(s)
Envejecimiento/metabolismo , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Hipófisis/metabolismo , Receptor trkB/metabolismo , Animales , Animales Recién Nacidos , Regulación del Desarrollo de la Expresión Génica/fisiología , Isoformas de Proteínas/metabolismo , Ratas , Ratas Sprague-Dawley , Distribución Tisular
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