RESUMEN
The possible use of cell therapies for neurological lesions and disorders is regarded as a very promising strategy. However, many issues related to cell type, tissue donor, expected biological action etc., are still open. In this study human mesenchymal stem cells derived from different fetal and adult tissues were examined in order to explore growth and neurotrophic factor synthesis and biological action, also considering the individual variability of the donors. Cells were derived from different human tissues and characterized according to the guidelines of the International Society for Cellular Therapy. Growth and neurotrophic factor synthesis was evaluated by real time PCR, biological assays and ELISA. It was found that human mesenchymal stem cells produce vascular endothelial-, nerve-growth factor (VEGF, NGF), brain-derived-, ciliary- and glial-derived neurotrophic factors (BDNF, CDGF, GDNF), which are neuroprotective molecules, but the source and the donor influence the synthesis rate. Accordingly, it is suggested that the source and the individual variability are key issues to be considered in the perspective of the clinical use of mesenchymal stem cells in neurological disorders.
Asunto(s)
Células Madre Mesenquimatosas/citología , Factores de Crecimiento Nervioso/biosíntesis , Diferenciación Celular , Separación Celular , Humanos , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/metabolismo , Factores de Crecimiento Nervioso/genética , ARN Mensajero/análisisRESUMEN
Two-dimensional vs three-dimensional culture conditions, such as the presence of extracellular matrix components, could deeply influence the cell fate and properties. In this paper we investigated proliferation, differentiation, survival, apoptosis, growth and neurotrophic factor synthesis of rat embryonic stem cells (RESCs) cultured in 2D and 3D conditions generated using Cultrex® Basement Membrane Extract (BME) and in poly-(L-lactic acid) (PLLA) electrospun sub-micrometric fibres. It is demonstrated that, in the absence of other instructive stimuli, growth, differentiation and paracrine activity of RESCs are directly affected by the different microenvironment provided by the scaffold. In particular, RESCs grown on an electrospun PLLA scaffolds coated or not with BME have a higher proliferation rate, higher production of bioactive nerve growth factor (NGF) and vascular endothelial growth factor (VEGF) compared to standard 2D conditions, lasting for at least 2 weeks. Due to the high mechanical flexibility of PLLA electrospun scaffolds, the PLLA/stem cell culture system offers an interesting potential for implantable neural repair devices.
Asunto(s)
Diferenciación Celular , Proliferación Celular , Células Madre Embrionarias/fisiología , Comunicación Paracrina , Andamios del Tejido/química , Animales , Apoptosis , Factor Neurotrófico Derivado del Encéfalo/genética , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Caspasa 3/metabolismo , Técnicas de Cultivo de Célula , Supervivencia Celular , Células Cultivadas , Medios de Cultivo , Técnicas Electroquímicas , Matriz Extracelular/metabolismo , Expresión Génica , Ácido Láctico/química , Factor de Crecimiento Nervioso/metabolismo , Factor 3 de Transcripción de Unión a Octámeros/genética , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Poliésteres , Polímeros/química , Ratas , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
AIMS: Neurogenesis in adult humans occurs in at least two areas of the brain, the subventricular zone of the telencephalon and the subgranular layer of the dentate gyrus in the hippocampal formation. We studied dentate gyrus subgranular layer neurogenesis in patients subjected to tailored antero-mesial temporal resection including amygdalohippocampectomy due to pharmacoresistant temporal lobe epilepsy (TLE) using the in vitro neurosphere assay. METHODS: Sixteen patients were enrolled in the study; mesial temporal sclerosis (MTS) was present in eight patients. Neurogenesis was investigated by ex vivo neurosphere expansion in the presence of mitogens (epidermal growth factor + basic fibroblast growth factor) and spontaneous differentiation after mitogen withdrawal. Growth factor synthesis was investigated by qRT-PCR in neurospheres. RESULTS: We demonstrate that in vitro proliferation of cells derived from dentate gyrus of TLE patients is dependent on disease duration. Moreover, the presence of MTS impairs proliferation. As long as in vitro proliferation occurs, neurogenesis is maintained, and cells expressing a mature neurone phenotype (TuJ1, MAP2, GAD) are spontaneously formed after mitogen withdrawal. Finally, formed neurospheres express mRNAs encoding for growth (vascular endothelial growth factor) as well as neurotrophic factors (brain-derived neurotrophic factor, ciliary neurotrophic factor, glial-derived neurotrophic factor, nerve growth factor). CONCLUSION: We demonstrated that residual neurogenesis in the subgranular layer of the dentate gyrus in TLE is dependent on diseases duration and absent in MTS.