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1.
Biomacromolecules ; 21(8): 3186-3196, 2020 08 10.
Artículo en Inglés | MEDLINE | ID: mdl-32786674

RESUMEN

Neurodegenerative diseases are generally characterized by a progressive loss of neuronal subpopulations, with no available cure to date. One of the main reasons for the limited clinical outcomes of new drug formulations is the lack of appropriate in vitro human cell models for research and validation. Stem cell technologies provide an opportunity to address this challenge by using patient-derived cells as a platform to test various drug formulations, including particle-based drug carriers. The therapeutic efficacy of drug delivery systems relies on efficient cellular uptake of the carrier and can be dependent on its size, shape, and surface chemistry. Although considerable efforts have been made to understand the effects of the physiochemical properties of particles on two-dimensional cell culture models, little is known of their effect in three-dimensional (3D) cell models of neurodegenerative diseases. Herein, we investigated the role of particle size (235-1000 nm), charge (cationic and anionic), and density (1.05 and 1.8 g cm-3) on the interactions of particles with human embryonic stem cell-derived 3D cell cultures of sensory neurons, called sensory neurospheres (sNSP). Templated layer-by-layer particles, with silica or polystyrene cores, and self-assembled glycogen/DNA polyplexes were used. Particles with sizes <280 nm effectively penetrated sNSP. Additionally, effective plasmid DNA delivery was observed up to 6 days post-transfection with glycogen/DNA polyplexes. The findings provide guidance in nanoparticle design for therapies aimed at neurodegenerative diseases, in particular Friedreich's ataxia, whereby sensory neurons are predominantly affected. They also demonstrate the application of 3D models of human sensory neurons in preclinical drug development.


Asunto(s)
Nanopartículas , Humanos , Neuronas , Tamaño de la Partícula , Dióxido de Silicio , Células Madre
2.
Cell Stem Cell ; 29(3): 434-448.e5, 2022 03 03.
Artículo en Inglés | MEDLINE | ID: mdl-35180398

RESUMEN

Midbrain dopamine (mDA) neurons can be replaced in patients with Parkinson's disease (PD) in order to provide long-term improvement in motor functions. The limited capacity for long-distance axonal growth in the adult brain means that cells are transplanted ectopically, into the striatal target. As a consequence, several mDA pathways are not re-instated, which may underlie the incomplete restoration of motor function in patients. Here, we show that viral delivery of GDNF to the striatum, in conjunction with homotopic transplantation of human pluripotent stem-cell-derived mDA neurons, recapitulates brain-wide mDA target innervation. The grafts provided re-instatement of striatal dopamine levels and correction of motor function and also connectivity with additional mDA target nuclei not well innervated by ectopic grafts. These results demonstrate the remarkable capacity for achieving functional and anatomically precise reconstruction of long-distance circuitry in the adult brain by matching appropriate growth-factor signaling to grafting of specific cell types.


Asunto(s)
Dopamina , Células Madre Pluripotentes , Adulto , Dopamina/metabolismo , Terapia Genética , Factor Neurotrófico Derivado de la Línea Celular Glial/metabolismo , Humanos , Mesencéfalo/metabolismo , Células Madre Pluripotentes/metabolismo , Sustancia Negra/metabolismo , Sustancia Negra/trasplante
3.
Brain Res Bull ; 173: 141-149, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-34022288

RESUMEN

Cortical neurospheres (NSPs) derived from human pluripotent stem cells (hPSC), have proven to be a successful platform to investigate human brain development and neuro-related diseases. Currently, many of the standard hPSC neural differentiation media, use concentrations of glucose (approximately 17.5-25 mM) and insulin (approximately 3.2 µM) that are much greater than the physiological concentrations found in the human brain. These culture conditions make it difficult to analyse perturbations of glucose or insulin on neuronal development and differentiation. We established a new hPSC neural differentiation medium that incorporated physiological brain concentrations of glucose (2.5 mM) and significantly reduced insulin levels (0.86 µM). This medium supported hPSC neural induction and formation of cortical NSPs. The revised hPSC neural differentiation medium, may provide an improved platform to model brain development and to investigate neural differentiation signalling pathways impacted by abnormal glucose and insulin levels.


Asunto(s)
Encéfalo/metabolismo , Diferenciación Celular/fisiología , Glucosa/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Encéfalo/citología , Medios de Cultivo , Humanos , Células Madre Pluripotentes Inducidas/citología
4.
Stem Cells Transl Med ; 10(8): 1157-1169, 2021 08.
Artículo en Inglés | MEDLINE | ID: mdl-33734599

RESUMEN

Friedreich ataxia (FRDA) is an autosomal recessive disease characterized by degeneration of dorsal root ganglia (DRG) sensory neurons, which is due to low levels of the mitochondrial protein Frataxin. To explore cell replacement therapies as a possible approach to treat FRDA, we examined transplantation of sensory neural progenitors derived from human embryonic stem cells (hESC) and FRDA induced pluripotent stem cells (iPSC) into adult rodent DRG regions. Our data showed survival and differentiation of hESC and FRDA iPSC-derived progenitors in the DRG 2 and 8 weeks post-transplantation, respectively. Donor cells expressed neuronal markers, including sensory and glial markers, demonstrating differentiation to these lineages. These results are novel and a highly significant first step in showing the possibility of using stem cells as a cell replacement therapy to treat DRG neurodegeneration in FRDA as well as other peripheral neuropathies.


Asunto(s)
Ataxia de Friedreich , Células Madre Pluripotentes Inducidas , Enfermedades del Sistema Nervioso Periférico , Ataxia de Friedreich/metabolismo , Ataxia de Friedreich/terapia , Ganglios Espinales , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Células Receptoras Sensoriales
5.
Biomater Sci ; 8(9): 2398-2403, 2020 May 07.
Artículo en Inglés | MEDLINE | ID: mdl-32270790

RESUMEN

Increasing frataxin protein levels through gene therapy is envisaged to improve therapeutic outcomes for patients with Friedreich's ataxia (FRDA). A non-viral strategy that uses submicrometer-sized multilayered particles to deliver frataxin-encoding plasmid DNA affords up to 27 000-fold increase in frataxin gene expression within 2 days in vitro in a stem cell-derived neuronal model of FRDA.


Asunto(s)
ADN/administración & dosificación , Ataxia de Friedreich , Proteínas de Unión a Hierro/genética , Modelos Biológicos , Plásmidos , Células Receptoras Sensoriales/metabolismo , Línea Celular Tumoral , Humanos , Células Madre Pluripotentes Inducidas/citología , Frataxina
6.
Methods Mol Biol ; 1976: 37-47, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-30977063

RESUMEN

There are a vast range of diseases and disorders that are neurocristopathic in origin, including Hirschsprung's disease, pheochromocytoma, familial dysautonomia, craniofacial disorders, and melanomas. Having a source of human neural crest cells is highly valuable for investigating potential treatments for such diseases. This chapter describes a robust and well-characterized protocol for deriving neural crest from human pluripotent stem cells (hPSCs), which can then be differentiated to neuronal and non-neuronal lineages. The protocol is adapted to suit hPSC maintenance as a monolayer bulk culture or as manual-passaged colonies, which makes it widely applicable to researchers that may use different systems for hPSC maintenance.


Asunto(s)
Cresta Neural/citología , Neuronas/citología , Células Madre Pluripotentes/citología , Animales , Técnicas de Cultivo de Célula , Diferenciación Celular , Humanos
7.
Int J Biochem Cell Biol ; 100: 61-68, 2018 07.
Artículo en Inglés | MEDLINE | ID: mdl-29772357

RESUMEN

Sensory neurons of the dorsal root ganglia (DRG) are the primary responders to stimuli inducing feelings of touch, pain, temperature, vibration, pressure and muscle tension. They consist of multiple subpopulations based on their morphology, molecular and functional properties. Our understanding of DRG sensory neurons has been predominantly driven by rodent studies and using transformed cell lines, whereas less is known about human sensory DRG neurons simply because of limited availability of human tissue. Although these previous studies have been fundamental for our understanding of the sensory system, it is imperative to profile human DRG subpopulations as it is becoming evident that human sensory neurons do not share the identical molecular and functional properties found in other species. Furthermore, there are wide range of diseases and disorders that directly/indirectly cause sensory neuronal degeneration or dysfunctionality. Having an in vitro source of human DRG sensory neurons is paramount for studying their development, unique neuronal properties and for accelerating regenerative therapies to treat sensory neuropathies. Here we review the major studies describing generation of DRG sensory neurons from human pluripotent stem cells and fibroblasts and the gaps that need to be addressed for using in vitro-generated human DRG neurons to model human DRG tissue.


Asunto(s)
Ganglios Espinales/citología , Enfermedades del Sistema Nervioso Periférico/patología , Células Madre Pluripotentes/citología , Animales , Diferenciación Celular , Fibroblastos/citología , Humanos , Células Receptoras Sensoriales/citología
8.
Sci Rep ; 8(1): 603, 2018 01 12.
Artículo en Inglés | MEDLINE | ID: mdl-29330377

RESUMEN

The dorsal root ganglia (DRG) consist of a multitude of sensory neuronal subtypes that function to relay sensory stimuli, including temperature, pressure, pain and position to the central nervous system. Our knowledge of DRG sensory neurons have been predominantly driven by animal studies and considerably less is known about the human DRG. Human embryonic stem cells (hESC) are valuable resource to help close this gap. Our previous studies reported an efficient system for deriving neural crest and DRG sensory neurons from hESC. Here we show that this differentiation system gives rise to heterogeneous populations of sensory neuronal subtypes as demonstrated by phenotypic and functional analyses. Furthermore, using microelectrode arrays the maturation rate of the hESC-derived sensory neuronal cultures was monitored over 8 weeks in culture, showing their spontaneous firing activities starting at about 12 days post-differentiation and reaching maximum firing at about 6 weeks. These studies are highly valuable for developing an in vitro platform to study the diversity of sensory neuronal subtypes found within the human DRG.


Asunto(s)
Técnicas de Cultivo de Célula/métodos , Células Madre Embrionarias Humanas/citología , Sistema Nervioso Periférico/citología , Células Receptoras Sensoriales/citología , Biomarcadores/metabolismo , Diferenciación Celular , Línea Celular , Células Madre Embrionarias Humanas/metabolismo , Humanos , Microelectrodos , Fenotipo , Células Receptoras Sensoriales/metabolismo
9.
Acta Histochem ; 117(4-5): 329-38, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-25747736

RESUMEN

Multipotent mesenchymal stromal cells, also known as mesenchymal stem cells (MSC), can be isolated from bone marrow or other tissues, including fat, muscle and umbilical cord. It has been shown that MSC behave in vitro as stem cells: they self-renew and are able to differentiate into mature cells typical of several mesenchymal tissues. Moreover, the differentiation toward non-mesenchymal cell lineages (e.g. neurons) has been reported as well. The clinical relevance of these cells is mainly related to their ability to spontaneously migrate to the site of inflammation/damage, to their safety profile thanks to their low immunogenicity and to their immunomodulation capacities. To date, MSCs isolated from the post-natal bone marrow have represented the most extensively studied population of adult MSCs, in view of their possible use in various therapeutical applications. However, the bone marrow-derived MSCs exhibit a series of limitations, mainly related to their problematic isolation, culturing and use. In recent years, umbilical cord (UC) matrix (i.e. Wharton's jelly, WJ) stromal cells have therefore emerged as a more suitable alternative source of MSCs, thanks to their primitive nature and the easy isolation without relevant ethical concerns. This review seeks to provide an overview of the main biological properties of WJ-derived MSCs. Moreover, the potential application of these cells for the treatment of some known dysfunctions in the central and peripheral nervous system will also be discussed.


Asunto(s)
Células Madre Adultas/metabolismo , Diferenciación Celular , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/metabolismo , Enfermedades Neurodegenerativas/terapia , Neuronas/metabolismo , Adulto , Humanos , Enfermedades Neurodegenerativas/metabolismo , Enfermedades Neurodegenerativas/patología
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