Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 15 de 15
Filtrar
1.
Chem Res Toxicol ; 33(1): 38-60, 2020 01 21.
Artículo en Inglés | MEDLINE | ID: mdl-31576743

RESUMEN

Despite extensive breakthroughs in chemistry, molecular biology, and genetics in the last decades, the success rates of drug development projects remain low. To improve predictions of clinical efficacy and safety of new compounds, a plethora of 3D culture methods of human cells have been developed in which the cultured cells retain physiologically and functionally relevant phenotypes for multiple weeks. Here, we critically review current paradigms for organotypic cultures of human liver, gut, and kidney such as perfused microchips, spheroids, and hollow fiber bioreactors and discuss their utility for understanding drug pharmacokinetics, metabolism, and toxicity. Furthermore, bioprinting and the microfluidic integration of different tissue models to mimic systemic drug effects are highlighted as promising technological trends. In the last part of the review, we discuss important considerations regarding the choice of culture substratum material to limit adverse effects such as drug absorption while facilitating the phenotypic maintenance of cultured cells. We conclude that recent advances in organotypic and microphysiological culture models of human tissues can improve drug development and contribute to an amelioration of clinical attrition rates. However, further validation, benchmarking, and consolidation efforts are needed to achieve more widespread dissemination and eventually regulatory acceptance of these novel tools.


Asunto(s)
Tracto Gastrointestinal/metabolismo , Riñón/metabolismo , Hígado/metabolismo , Modelos Biológicos , Preparaciones Farmacéuticas/metabolismo , Animales , Evaluación Preclínica de Medicamentos , Efectos Colaterales y Reacciones Adversas Relacionados con Medicamentos , Humanos , Farmacocinética
2.
Arterioscler Thromb Vasc Biol ; 39(11): 2273-2288, 2019 11.
Artículo en Inglés | MEDLINE | ID: mdl-31533473

RESUMEN

OBJECTIVE: Activation of endothelial ß-catenin signaling by neural cell-derived Norrin or Wnt ligands is vital for the vascularization of the retina and brain. Mutations in members of the Norrin/ß-catenin pathway contribute to inherited blinding disorders because of defective vascular development and dysfunctional blood-retina barrier. Despite a vital role for endothelial ß-catenin signaling in central nervous system health and disease, its contribution to central nervous system angiogenesis and its interactions with downstream signaling cascades remains incompletely understood. Approach and Results: Here, using genetically modified mouse models, we show that impaired endothelial ß-catenin signaling caused hypovascularization of the postnatal retina and brain because of deficient endothelial cell proliferation and sprouting. Mosaic genetic analysis demonstrated that endothelial ß-catenin promotes but is not required for tip cell formation. In addition, pharmacological treatment revealed that angiogenesis under conditions of inhibited Notch signaling depends upon endothelial ß-catenin. Importantly, impaired endothelial ß-catenin signaling abrogated the expression of the VEGFR (vascular endothelial growth factor receptor)-2 and VEGFR3 in brain microvessels but not in the lung endothelium. CONCLUSIONS: Our study identifies molecular crosstalk between the Wnt/ß-catenin and the Notch and VEGF-A signaling pathways and strongly suggest that endothelial ß-catenin signaling supports central nervous system angiogenesis by promoting endothelial cell sprouting, tip cell formation, and VEGF-A/VEGFR2 signaling.


Asunto(s)
Encéfalo/irrigación sanguínea , Endotelio Vascular/metabolismo , Neovascularización Fisiológica , Retina/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Vía de Señalización Wnt , beta Catenina/metabolismo , Animales , Proteína Axina/metabolismo , Barrera Hematoencefálica/metabolismo , Proliferación Celular , Células Endoteliales/metabolismo , Ratones Transgénicos , Microcirculación , Receptor Cross-Talk , Receptor Notch1/genética , Receptor Notch1/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética
3.
Arterioscler Thromb Vasc Biol ; 39(7): 1432-1447, 2019 07.
Artículo en Inglés | MEDLINE | ID: mdl-31242033

RESUMEN

Objective- The Wnt/ß-catenin pathway orchestrates development of the blood-brain barrier, but the downstream mechanisms involved at different developmental windows and in different central nervous system (CNS) tissues have remained elusive. Approach and Results- Here, we create a new mouse model allowing spatiotemporal investigations of Wnt/ß-catenin signaling by induced overexpression of Axin1, an inhibitor of ß-catenin signaling, specifically in endothelial cells ( Axin1 iEC- OE). AOE (Axin1 overexpression) in Axin1 iEC- OE mice at stages following the initial vascular invasion of the CNS did not impair angiogenesis but led to premature vascular regression followed by progressive dilation and inhibition of vascular maturation resulting in forebrain-specific hemorrhage 4 days post-AOE. Analysis of the temporal Wnt/ß-catenin driven CNS vascular development in zebrafish also suggested that Axin1 iEC- OE led to CNS vascular regression and impaired maturation but not inhibition of ongoing angiogenesis within the CNS. Transcriptomic profiling of isolated, ß-catenin signaling-deficient endothelial cells during early blood-brain barrier-development (E11.5) revealed ECM (extracellular matrix) proteins as one of the most severely deregulated clusters. Among the 20 genes constituting the forebrain endothelial cell-specific response signature, 8 ( Adamtsl2, Apod, Ctsw, Htra3, Pglyrp1, Spock2, Ttyh2, and Wfdc1) encoded bona fide ECM proteins. This specific ß-catenin-responsive ECM signature was also repressed in Axin1 iEC- OE and endothelial cell-specific ß-catenin-knockout mice ( Ctnnb1-KOiEC) during initial blood-brain barrier maturation (E14.5), consistent with an important role of Wnt/ß-catenin signaling in orchestrating the development of the forebrain vascular ECM. Conclusions- These results suggest a novel mechanism of establishing a CNS endothelium-specific ECM signature downstream of Wnt-ß-catenin that impact spatiotemporally on blood-brain barrier differentiation during forebrain vessel development. Visual Overview- An online visual overview is available for this article.


Asunto(s)
Matriz Extracelular/fisiología , Prosencéfalo/irrigación sanguínea , Vía de Señalización Wnt/fisiología , beta Catenina/fisiología , Animales , Proteína Axina/fisiología , Barrera Hematoencefálica , Masculino , Ratones , Ratones Endogámicos C57BL , Transducción de Señal/fisiología , Remodelación Vascular , Pez Cebra
4.
Arterioscler Thromb Vasc Biol ; 39(7): 1402-1418, 2019 07.
Artículo en Inglés | MEDLINE | ID: mdl-31242036

RESUMEN

Objective- Pathological neovascularization is crucial for progression and morbidity of serious diseases such as cancer, diabetic retinopathy, and age-related macular degeneration. While mechanisms of ongoing pathological neovascularization have been extensively studied, the initiating pathological vascular remodeling (PVR) events, which precede neovascularization remains poorly understood. Here, we identify novel molecular and cellular mechanisms of preneovascular PVR, by using the adult choriocapillaris as a model. Approach and Results- Using hypoxia or forced overexpression of VEGF (vascular endothelial growth factor) in the subretinal space to induce PVR in zebrafish and rats respectively, and by analyzing choriocapillaris membranes adjacent to choroidal neovascular lesions from age-related macular degeneration patients, we show that the choriocapillaris undergo robust induction of vascular intussusception and permeability at preneovascular stages of PVR. This PVR response included endothelial cell proliferation, formation of endothelial luminal processes, extensive vesiculation and thickening of the endothelium, degradation of collagen fibers, and splitting of existing extravascular columns. RNA-sequencing established a role for endothelial tight junction disruption, cytoskeletal remodeling, vesicle- and cilium biogenesis in this process. Mechanistically, using genetic gain- and loss-of-function zebrafish models and analysis of primary human choriocapillaris endothelial cells, we determined that HIF (hypoxia-induced factor)-1α-VEGF-A-VEGFR2 signaling was important for hypoxia-induced PVR. Conclusions- Our findings reveal that PVR involving intussusception and splitting of extravascular columns, endothelial proliferation, vesiculation, fenestration, and thickening is induced before neovascularization, suggesting that identifying and targeting these processes may prevent development of advanced neovascular disease in the future. Visual Overview- An online visual overview is available for this article.


Asunto(s)
Neovascularización Patológica/etiología , Remodelación Vascular/fisiología , Adulto , Animales , Humanos , Hipoxia , Subunidad alfa del Factor 1 Inducible por Hipoxia/fisiología , Degeneración Macular/etiología , Factor A de Crecimiento Endotelial Vascular/fisiología , Receptor 2 de Factores de Crecimiento Endotelial Vascular/fisiología , Pez Cebra
5.
iScience ; 27(4): 109346, 2024 Apr 19.
Artículo en Inglés | MEDLINE | ID: mdl-38500830

RESUMEN

Neonatal hypoxia-ischemia (HI) is a major cause of perinatal death and long-term disabilities worldwide. Post-ischemic neuroinflammation plays a pivotal role in HI pathophysiology. In the present study, we investigated the temporal dynamics of microglia (CX3CR1GFP/+) and infiltrating macrophages (CCR2RFP/+) in the hippocampi of mice subjected to HI at postnatal day 9. Using inflammatory pathway and transcription factor (TF) analyses, we identified a distinct post-ischemic response in CCR2RFP/+ cells characterized by differential gene expression in sensome, homeostatic, matrisome, lipid metabolic, and inflammatory molecular signatures. Three days after injury, transcriptomic signatures of CX3CR1GFP/+ and CCR2RFP/+ cells isolated from hippocampi showed a partial convergence. Interestingly, microglia-specific genes in CX3CR1GFP/+ cells showed a sexual dimorphism, where expression returned to control levels in males but not in females during the experimental time frame. These results highlight the importance of further investigations on metabolic rewiring to pave the way for future interventions in asphyxiated neonates.

6.
Stem Cells ; 30(5): 865-75, 2012 May.
Artículo en Inglés | MEDLINE | ID: mdl-22290867

RESUMEN

Secreted Frizzled related proteins (sFRPs) are a family of proteins that modulate Wnt signaling, which in turn regulates multiple aspects of ventral midbrain (VM) and dopamine (DA) neuron development. However, it is not known which Wnt signaling branch and what aspects of midbrain DA neuron development are regulated by sFRPs. Here, we show that sFRP1 and sFRP2 activate the Wnt/planar-cell-polarity/Rac1 pathway in DA cells. In the developing VM, sFRP1 and sFRP2 are expressed at low levels, and sFRP1-/- or sFRP2-/- mice had no detectable phenotype. However, compound sFRP1-/-;sFRP2-/- mutants revealed a Wnt/PCP phenotype similar to that previously described for Wnt5a-/- mice. This included an anteroposterior shortening of the VM, a lateral expansion of the Shh domain and DA lineage markers (Lmx1a and Th), as well as an accumulation of Nurr1+ precursors in the VM. In vitro experiments showed that, while very high concentrations of SFRP1 had a negative effect on cell survival, low/medium concentrations of sFRP1 or sFRP2 promoted the DA differentiation of progenitors derived from primary VM cultures or mouse embryonic stem cells (ESCs), mimicking the effects of Wnt5a. We thus conclude that the main function of sFRP1 and sFRP2 is to enhance Wnt/PCP signaling in DA cells and to regulate Wnt/PCP-dependent functions in midbrain development. Moreover, we suggest that low-medium concentrations of sFRPs may be used to enhance the DA differentiation of ESCs and improve their therapeutic application.


Asunto(s)
Neuronas Dopaminérgicas/metabolismo , Células Madre Embrionarias/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Proteínas de la Membrana/metabolismo , Mesencéfalo/embriología , Proteínas del Tejido Nervioso/metabolismo , Animales , Neuronas Dopaminérgicas/citología , Relación Dosis-Respuesta a Droga , Células Madre Embrionarias/citología , Péptidos y Proteínas de Señalización Intercelular/genética , Péptidos y Proteínas de Señalización Intercelular/farmacología , Proteínas de la Membrana/genética , Proteínas de la Membrana/farmacología , Mesencéfalo/citología , Ratones , Ratones Noqueados , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/farmacología , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología , Proteínas Wnt/genética , Proteínas Wnt/metabolismo
7.
Commun Biol ; 5(1): 27, 2022 01 11.
Artículo en Inglés | MEDLINE | ID: mdl-35017663

RESUMEN

Despite successful combination antiretroviral therapy (cART), persistent low-grade immune activation together with inflammation and toxic antiretroviral drugs can lead to long-lasting metabolic flexibility and adaptation in people living with HIV (PLWH). Our study investigated alterations in the plasma metabolic profiles by comparing PLWH on long-term cART(>5 years) and matched HIV-negative controls (HC) in two cohorts from low- and middle-income countries (LMIC), Cameroon, and India, respectively, to understand the system-level dysregulation in HIV-infection. Using untargeted and targeted LC-MS/MS-based metabolic profiling and applying advanced system biology methods, an altered amino acid metabolism, more specifically to glutaminolysis in PLWH than HC were reported. A significantly lower level of neurosteroids was observed in both cohorts and could potentiate neurological impairments in PLWH. Further, modulation of cellular glutaminolysis promoted increased cell death and latency reversal in pre-monocytic HIV-1 latent cell model U1, which may be essential for the clearance of the inducible reservoir in HIV-integrated cells.


Asunto(s)
Fármacos Anti-VIH/uso terapéutico , Glutamina/metabolismo , Infecciones por VIH , Metaboloma , Adulto , Células Cultivadas , Metabolismo Energético/genética , Metabolismo Energético/fisiología , Femenino , Glucólisis/genética , Glucólisis/fisiología , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/metabolismo , Infecciones por VIH/fisiopatología , Humanos , Masculino , Metaboloma/genética , Metaboloma/fisiología , Metabolómica , Persona de Mediana Edad , Biología de Sistemas
8.
Cell Rep ; 31(9): 107699, 2020 06 02.
Artículo en Inglés | MEDLINE | ID: mdl-32492415

RESUMEN

Cranial irradiation (IR), an effective tool to treat malignant brain tumors, triggers a chronic pro-inflammatory microglial response, at least in the adult brain. Using single-cell and bulk RNA sequencing, combined with histology, we show that the microglial response in the juvenile mouse hippocampus is rapid but returns toward normal within 1 week. The response is characterized by a series of temporally distinct homeostasis-, sensome-, and inflammation-related molecular signatures. We find that a single microglial cell simultaneously upregulates transcripts associated with pro- and anti-inflammatory microglial phenotypes. Finally, we show that juvenile and adult irradiated microglia are already transcriptionally distinct in the early phase after IR. Our results indicate that microglia are involved in the initial stages but may not be responsible for driving long-term inflammation in the juvenile brain.


Asunto(s)
Neoplasias Encefálicas/radioterapia , Microglía/metabolismo , Radiación Ionizante , Envejecimiento , Animales , Quimiocinas/genética , Quimiocinas/metabolismo , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Regulación hacia Abajo/efectos de la radiación , Femenino , Hipocampo/metabolismo , Hipocampo/patología , Ratones , Ratones Endogámicos C57BL , Microglía/citología , Microglía/efectos de la radiación , Análisis de Secuencia de ARN , Análisis de la Célula Individual , Regulación hacia Arriba/efectos de la radiación
9.
Sci Signal ; 10(487)2017 07 11.
Artículo en Inglés | MEDLINE | ID: mdl-28698213

RESUMEN

The blood-brain barrier is a dynamic interface that separates the brain from the circulatory system, and it is formed by highly specialized endothelial cells. To explore the molecular mechanisms defining the unique nature of vascular development and differentiation in the brain, we generated high-resolution gene expression profiles of mouse embryonic brain endothelial cells using translating ribosome affinity purification and single-cell RNA sequencing. We compared the brain vascular translatome with the vascular translatomes of other organs and analyzed the vascular translatomes of the brain at different time points during embryonic development. Because canonical Wnt signaling is implicated in the formation of the blood-brain barrier, we also compared the brain endothelial translatome of wild-type mice with that of mice lacking the transcriptional cofactor ß-catenin (Ctnnb1). Our analysis revealed extensive molecular changes during the embryonic development of the brain endothelium. We identified genes encoding brain endothelium-specific transcription factors (Foxf2, Foxl2, Foxq1, Lef1, Ppard, Zfp551, and Zic3) that are associated with maturation of the blood-brain barrier and act downstream of the Wnt-ß-catenin signaling pathway. Profiling of individual brain endothelial cells revealed substantial heterogeneity in the population. Nevertheless, the high abundance of Foxf2, Foxq1, Ppard, or Zic3 transcripts correlated with the increased expression of genes encoding markers of brain endothelial cell differentiation. Expression of Foxf2 and Zic3 in human umbilical vein endothelial cells induced the production of blood-brain barrier differentiation markers. This comprehensive data set may help to improve the engineering of in vitro blood-brain barrier models.


Asunto(s)
Encéfalo/embriología , Embrión de Mamíferos/embriología , Desarrollo Embrionario/fisiología , Células Endoteliales/metabolismo , Perfilación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica/fisiología , Animales , Encéfalo/citología , Embrión de Mamíferos/citología , Células Endoteliales/citología , Ratones , Ratones Transgénicos
10.
Stem Cells Dev ; 23(17): 1991-2003, 2014 Sep 01.
Artículo en Inglés | MEDLINE | ID: mdl-24803261

RESUMEN

During development of the central nervous system, trophic, together with genetic, cues dictate the balance between cellular proliferation and differentiation. Subsequent to the birth of new neurons, additional intrinsic and extrinsic signals regulate the connectivity of these cells. While a number of regulators of ventral midbrain (VM) neurogenesis and dopaminergic (DA) axon guidance are known, we identify a number of novel roles for the secreted glycoprotein, Wnt7a, in this context. We demonstrate a temporal and spatial expression of Wnt7a in the VM, indicative of roles in neurogenesis, differentiation, and axonal growth and guidance. In primary VM cultures, and validated in Wnt7a-deficient mice, we show that the early expression within the VM is important for regulating VM progenitor proliferation, cell cycle progression, and cell survival, thereby dictating the number of midbrain Nurr1 precursors and DA neurons. During early development of the midbrain DA pathways, Wnt7a promotes axonal elongation and repels DA neurites out of the midbrain. Later, Wnt7a expression in the VM midline suggests a role in preventing axonal crossing while expression in regions flanking the medial forebrain bundle (thalamus and hypothalamus) ensured appropriate trajectory of DA axons en route to their forebrain targets. We show that the effects of Wnt7a in VM development are mediated, at least in part, by the ß-catenin/canonical pathways. Together, these findings identify Wnt7a as a new regulator of VM neurogenesis and DA axon growth and guidance.


Asunto(s)
Axones/fisiología , Mesencéfalo/embriología , Neurogénesis , Proteínas Wnt/fisiología , Animales , Ciclo Celular , Forma de la Célula , Supervivencia Celular , Células Cultivadas , Neuronas Dopaminérgicas/metabolismo , Regulación del Desarrollo de la Expresión Génica , Mesencéfalo/citología , Mesencéfalo/metabolismo , Ratones Noqueados , Morfogénesis , Fibras Nerviosas/fisiología , Células-Madre Neurales/fisiología , Neuritas/fisiología , Especificidad de Órganos , Ratas Sprague-Dawley
11.
PLoS One ; 5(3): e9647, 2010 Mar 19.
Artículo en Inglés | MEDLINE | ID: mdl-20333299

RESUMEN

Nerve Growth Factor (NGF)-induced neuronal differentiation requires the activation of members of the Rho family of small GTPases. However, the molecular mechanisms through which NGF regulates cytoskeletal changes and neurite outgrowth are not totally understood. In this work, we identify the Rac1-specific guanine exchange factor (GEF) Tiam1 as a novel mediator of NGF/TrkA-dependent neurite elongation. In particular, we report that knockdown of Tiam1 causes a significant reduction in Rac1 activity and neurite outgrowth induced by NGF. Physical interaction between Tiam1 and active Ras (Ras-GTP), but not tyrosine phosphorylation of Tiam1, plays a central role in Rac1 activation by NGF. In addition, our findings indicate that Ras is required to associate Tiam1 with Rac1 and promote Rac1 activation upon NGF stimulation. Taken together, these findings define a novel molecular mechanism through which Tiam1 mediates TrkA signaling and neurite outgrowth induced by NGF.


Asunto(s)
Factores de Intercambio de Guanina Nucleótido/metabolismo , Proteínas de Neoplasias/metabolismo , Factor de Crecimiento Nervioso/metabolismo , Neuritas/metabolismo , Receptor trkA/metabolismo , Animales , Células COS , Diferenciación Celular , Chlorocebus aethiops , Humanos , Neuronas/metabolismo , Células PC12 , Ratas , Transducción de Señal , Proteína 1 de Invasión e Inducción de Metástasis del Linfoma-T , Proteína de Unión al GTP rac1/metabolismo , Proteínas ras/metabolismo
12.
Exp Cell Res ; 312(9): 1626-36, 2006 May 15.
Artículo en Inglés | MEDLINE | ID: mdl-16510140

RESUMEN

Wnt1 and -5a have been shown to modulate the proliferation and differentiation of midbrain dopaminergic (DA) neurons. However, it is not known whether other Wnts or which Frizzled (Fz) receptors are expressed in the developing midbrain. We found that 13 out of 19 Wnts, all 10 Fzs, and several intracellular Wnt signaling modulators, including Axin, FRAT, Naked, Par-1, and Ltap are developmentally regulated between embryonic days (E) 10.5 and 15.5. Next, we studied whether Fzs are differentially expressed in different cell types and examined neuronal-progenitor- or glial-enriched cultures and DA neurons isolated from TH-GFP reporter mice. We found that Fz8 is expressed at high levels in DA neurons at E11.5 and E13.5. Fz6 and -7 are the predominant transcripts in glial precursors, and Fz9, which is absent in DA neurons at E11.5, is the main receptor expressed in neuronal precursors. We therefore examined the function of Fz9 in DA cells and found that overexpression of Fz9 reduced Wnt5a- but not Wnt3a-induced hyperphosphorylation of Dishevelled. Thus, our results show that Fzs are developmentally regulated and differentially expressed in VM precursors, DA neurons, and glia. These findings suggest that Fz expression contributes to provide specificity to Wnt-mediated effects.


Asunto(s)
Regulación del Desarrollo de la Expresión Génica , Mesencéfalo/metabolismo , Transducción de Señal/genética , Proteínas Wnt/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Proteína Axina , Línea Celular , Células Cultivadas , Proteínas Dishevelled , Femenino , Receptores Frizzled/genética , Péptidos y Proteínas de Señalización Intracelular/genética , Masculino , Mesencéfalo/citología , Mesencéfalo/embriología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas del Tejido Nervioso/genética , Fosfoproteínas/metabolismo , Fosforilación/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/farmacología , Ratas , Receptores de Neurotransmisores/genética , Proteínas Recombinantes/farmacología , Proteínas Represoras/genética , Transfección , Proteínas Wnt/farmacología , Proteína Wnt-5a
13.
J Neurochem ; 99(1): 343-52, 2006 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-16987254

RESUMEN

In the present study, we investigated the involvement of rhombomere 1 patterning proteins in the regulation of the major noradrenergic centre of the brain, the locus coeruleus. Primary cultures of rat embryonic day 13.5 locus coeruleus were treated with fibroblast growth factor-8, noggin and members of the bone morphogenetic and Wnt protein families. We show that bone morphogenetic proteins 2, 5 and 7 increase and noggin decreases the number of tyrosine hydroxylase-positive locus coeruleus neurons. Interestingly, from all Wnts expressed in the first rhombomere by embryonic day 12.5 in the mice, we only found expression of wnt5a mRNA in the vicinity of the locus coeruleus. In agreement with this finding, from all Wnts studied in vitro, only Wnt5a increased the number of tyrosine hydroxylase-positive neurons in locus coeruleus cultures. Finally, we also found that fibroblast growth factor-8 increased the number of tyrosine hydroxylase-positive cells in locus coeruleus cultures. Neither of the identified factors affected the survival of tyrosine hydroxylase-positive locus coeruleus noradrenergic neurons or the proliferation of their progenitors or neurogenesis. Instead, our results suggest that these patterning signals of rhombomere 1 may work to promote the differentiation of noradrenergic progenitors at later stages of development.


Asunto(s)
Proteínas Morfogenéticas Óseas/fisiología , Desarrollo Embrionario/fisiología , Factor 8 de Crecimiento de Fibroblastos/fisiología , Locus Coeruleus/embriología , Neuronas/fisiología , Proteínas Wnt/fisiología , Animales , Apoptosis , Técnicas de Cultivo de Célula/métodos , Diferenciación Celular , Separación Celular , Medios de Cultivo Condicionados , Femenino , Etiquetado Corte-Fin in Situ , Locus Coeruleus/citología , Neuronas/citología , Embarazo , Ratas , Ratas Sprague-Dawley
14.
Development ; 133(3): 495-505, 2006 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-16410412

RESUMEN

Proneural genes are crucial regulators of neurogenesis and subtype specification in many areas of the nervous system; however, their function in dopaminergic neuron development is unknown. We report that proneural genes have an intricate pattern of expression in the ventricular zone of the ventral midbrain, where mesencephalic dopaminergic neurons are generated. Neurogenin 2 (Ngn2) and Mash1 are expressed in the ventral midline, while Ngn1, Ngn2 and Mash1 are co-localized more laterally in the ventricular zone. Ngn2 is also expressed in an intermediate zone immediately adjacent to the ventricular zone at the ventral midline. To examine the function of these genes, we analyzed mutant mice in which one or two of these genes were deleted (Ngn1, Ngn2 and Mash1) or substituted (Mash1 in the Ngn2 locus). Our results demonstrate that Ngn2 is required for the differentiation of Sox2(+) ventricular zone progenitors into Nurr1(+) postmitotic dopaminergic neuron precursors in the intermediate zone, and that it is also likely to be required for their subsequent differentiation into tyrosine hydroxylase-positive dopaminergic neurons in the marginal zone. Although Mash1 normally has no detectable function in dopaminergic neuron development, it could partially rescue the generation of dopaminergic neuron precursors in the absence of Ngn2. These results demonstrate that Ngn2 is uniquely required for the development of midbrain dopaminergic neurons.


Asunto(s)
Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Dopamina/metabolismo , Mesencéfalo/citología , Mesencéfalo/embriología , Proteínas del Tejido Nervioso/metabolismo , Neuronas/fisiología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Diferenciación Celular/fisiología , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Femenino , Proteínas de Homeodominio/metabolismo , Hibridación in Situ , Proteínas con Homeodominio LIM , Masculino , Ratones , Ratones Noqueados , Proteínas del Tejido Nervioso/genética , Neuronas/citología , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares , Factores de Transcripción SOXB1 , Células Madre/citología , Células Madre/fisiología , Transactivadores/genética , Transactivadores/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Tirosina 3-Monooxigenasa/metabolismo
15.
Proc Natl Acad Sci U S A ; 100(22): 12747-52, 2003 Oct 28.
Artículo en Inglés | MEDLINE | ID: mdl-14557550

RESUMEN

The Wnts are a family of glycoproteins that regulate cell proliferation, fate decisions, and differentiation. In our study, we examined the contribution of Wnts to the development of ventral midbrain (VM) dopaminergic (DA) neurons. Our results show that beta-catenin is expressed in DA precursor cells and that beta-catenin signaling takes place in these cells, as assessed in TOPGAL [Tcf optimal-promoter beta-galactosidase] reporter mice. We also found that Wnt-1, -3a, and -5a expression is differentially regulated during development and that partially purified Wnts distinctively regulate VM development. Wnt-3a promoted the proliferation of precursor cells expressing the orphan nuclear receptor-related factor 1 (Nurr1) but did not increase the number of tyrosine hydroxylase-positive neurons. Instead, Wnt-1 and -5a increased the number of rat midbrain DA neurons in rat embryonic day 14.5 precursor cultures by two distinct mechanisms. Wnt-1 predominantly increased the proliferation of Nurr1+ precursors, up-regulated cyclins D1 and D3, and down-regulated p27 and p57 mRNAs. In contrast, Wnt-5a primarily increased the proportion of Nurr1+ precursors that acquired a neuronal DA phenotype and up-regulated the expression of Ptx3 and c-ret mRNA. Moreover, the soluble cysteine-rich domain of Frizzled-8 (a Wnt inhibitor) blocked endogenous Wnts and the effects of Wnt-1 and -5a on proliferation and the acquisition of a DA phenotype in precursor cultures. These findings indicate that Wnts are key regulators of proliferation and differentiation of DA precursors during VM neurogenesis and that different Wnts have specific and unique activity profiles.


Asunto(s)
Proteínas del Citoesqueleto/fisiología , Regulación del Desarrollo de la Expresión Génica/fisiología , Mesencéfalo/fisiología , Neuronas/fisiología , Proteínas Proto-Oncogénicas/genética , Transactivadores/fisiología , Proteínas de Pez Cebra , Animales , Diferenciación Celular , Células Cultivadas , Proteínas del Citoesqueleto/genética , Femenino , Masculino , Ratones , Mitógenos/fisiología , Neuronas/citología , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transactivadores/genética , Proteínas Wnt , Proteína Wnt-5a , Proteína Wnt1 , Proteína wnt2 , beta Catenina
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA