Transcriptional characterization of Wnt and Notch signaling pathways in neuronal differentiation of human adipose tissue-derived stem cells.
ABSTRACT Since the nervous system has limited self-repair capability, a great interest in using stem cells is generated to repair it. The adipose tissue is an abundant source of stem cells and previous reports have shown the differentiation of them in neuron-like cells when cultures are enriched with growth factors involved in neurogenesis. Regarding this, it could be thought that a functional parallelism between neurogenesis and neuronal differentiation of human adipose stem cells (hASCs) exists. For this reason, we investigated the putative involvement of Notch and Wnt pathways in neuronal differentiation of hASCs through real-time PCR. We found that both Wnt and Notch signaling are present in proliferating hASCs and that both cascades are downregulated when cells are differentiated to a neuronal phenotype. These results are in concordance with previous works where it was found that both pathways are involved in the maintenance of the proliferative state of stem cells, probably through inhibition of the expression of cell-fate-specific genes. These results could support the notion that hASCs differentiation into neuron-like cells represents a regulated process analogous to what occurs during neuronal differentiation of NSCs and could partially contribute to elucidate the molecular mechanisms involved in neuronal differentiation of adult human nonneural tissues.
- [Show abstract] [Hide abstract]
ABSTRACT: Notch signaling plays critical roles in stem cells by regulating cell fate determination and differentiation. The aim of this study was to evaluate the participation of Notch signaling in neurogenic commitment of human periodontal ligament-derived mesenchymal stem cells (hPDLSCs) and to examine the ability to control differentiation of these cells using modified surfaces containing affinity immobilized Notch-ligands. Neurogenic induction of hPDLSCs was performed via neurosphere formation. Cells were aggregated and form spheres as early 1 day in culture. In addition, the induced cells exhibited increased mRNA and protein expression of neuronal markers i.e. β3-tubulin and neurofilament. During neuronal differentiation, a significant increase of Hes1 and Hey1 mRNA expression was noted. Using pharmacological inhibition (γ-secretase inhibitor) or genetic manipulation (overexpression of dominant negative mastermind-like transcription co-activators), neurosphere formation was attenuated and a marked decrease in neurogenic mRNA expression was observed. To confirm the role of Notch signaling in neuronal differentiation of hPDLSCs, the Notch ligand, Jagged-1, is bound to the surface using an affinity immobilization technique. The hPDLSC cultured on a Jagged-1modified surface had increased expression of Notch signaling target genes, Hes-1 and Hey-1, confirming the activity and potency of surface-bound Jagged-1. Further, hPDLSC on surface-bound Jagged-1 under serum free conditions showed multiple long and thin neurite-like extensions, and an increase in the expression of neurogenic mRNA markers was observed. Pretreatment of the cells with γ-secretase inhibitor, DAPT, prior to seeding on the Jagged-1 modified surface blocked development of the neurite-like morphology. Together, the results in this study suggest the involvement of Notch signaling in neurogenic commitment of hPDLSCs.Stem cells and development 02/2013; · 4.15 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: Multipotent human adipose-derived stromal/stem cells (hADSCs) hold a great promise for cell-based therapy for many devastating human diseases such as spinal cord injury, stroke et al. If exogenous hADSCs can be cultured in a three-dimensional scaffold with effective proliferation and differentiation capacity, it will better mimic the in vivo environment, which will have profound impact on the therapeutic application of hADSCs. In this study, a group of elastic-dominant, porous bioscaffolds from photocurable chitosan and gelatin were fabricated and proven to be bio-compatible with both hADSCs and hADSC-derived neuron-like cells (hADSC-NLCs) in vitro. The identity of harvested hADSCs was confirmed by their positive immunostaining of mesenchymal stem cell surface markers CD29, CD44, and CD105 , and alsopositive expression of stem markers Sox-2, Oct-4, c-Myc, Nanog and Klf-4.. Theirmultipotency was further confirmed by tri-lineage differentiation of hADSCs towards adipocyte, osteoblast and chondrocyte. It was found that hADSCs could be conditioned to differentiate into neurons in vitro as determined by immunostaining the markers of Tuj1, MAP2, NeuN and Synapsin. The hADSCs and hADSC-NLCs were proven to be biocompatible with 3D scaffold, which actually facilitated the proliferation and differentiation of hADSCs in vitro,by MTT assay and their neuronal gene expression profiling. Moreover, hADSC-NLCs, which were mixed with 3D scaffold and transplanted into traumatic brain injury mouse model, survived in vivo and led to the better repair of the damaged brain area. The immunohistochemical studies revealed that 3D scaffold indeed improved the viability of transplanted cells, their ability to incorporate into the in vivo neural circuit and their capacity for tissue repair. This study indicates that hADSCs would have great therapeutic application potential as seeding cells for in vivo transplantation to treat various neurological diseases when co-applied with porous chitosan/gelatin bioscaffolds.Tissue Engineering Part A 11/2013; · 4.64 Impact Factor