Bone marrow- and adipose-derived stem cells show expression of myelin mRNAs and proteins.

AIMS: PNS myelin is formed by Schwann cells (SCs). In this study, we applied an in vitro model to study myelin formation, using bone marrow mesenchymal stem cells and adipose-derived stem cells differentiated into SC-like cells and co-cultured with dissociated adult dorsal root ganglia neurons. METHODS: Immunocytochemistry, reverse transcription-PCR and western blotting techniques were used to investigate the expression of myelin proteins at both the transcriptional and translational level. RESULTS: Transcripts for protein zero, peripheral myelin protein 22 and myelin basic protein were detected in differentiated stem cells following co-culture with neuronal cells. Furthermore, protein zero, peripheral myelin protein 22 and myelin basic proteins were recognized in the co-cultures. These results were consistent with immunostaining of myelin proteins and with observation by electron microscopy. CONCLUSION: Both types of adult stems cells differentiated into SC-like cells have potential to myelinate neuronal cells during regeneration, being functionally identical to SCs of the PNS.

Characterisation of human mesenchymal stem cells following differentiation into Schwann cell-like cells.

Cell-based therapies provide a clinically applicable and available alternative to nerve autografts. Our previous studies have characterised rat-derived mesenchymal stem cells (MSC) and here we have investigated the phenotypic, molecular and functional characteristics of human-derived MSC (hMSC) differentiated along a Schwann cell lineage. The hMSC were isolated from healthy human donors and the identity of the undifferentiated hMSC was confirmed by the detection of MSC specific cells surface markers. The hMSC were differentiated along a glial cell lineage using an established cocktail of growth factors including glial growth factor-2. Following differentiation, the hMSC expressed the key Schwann cell (SC) markers at both the transcriptional and translational level. More importantly, we show the functional effect of hMSC on neurite outgrowth using an in vitro co-culture model system with rat-derived primary sensory neurons. The number of DRG sprouting neurites was significantly enhanced in the presence of differentiated hMSC; neurite length and density (branching) were also increased. These results provide evidence that hMSC can undergo molecular, morphological and functional changes to adopt a SC-like behaviour and, therefore, could be suitable as SC substitutes for nerve repair in clinical applications.

Schwann cell mediated trophic effects by differentiated mesenchymal stem cells.

Mesenchymal stem cells were isolated from the bone marrow of rats and differentiated to provide a functional substitute for slow growing Schwann cells for peripheral nerve regeneration. To assess the properties of the differentiated mesenchymal stem cell, the cells were co-cultured with dorsal root ganglia and the secretion of the neurotrophic factors and the neurite outgrowth was evaluated. The neurite outgrowth of the dorsal root ganglia neurons was enhanced in co-culture with the differentiated stem cells compared to the undifferentiated stem cells. Differentiated stem cells like Schwann cells were responsible for the stimulation of longer and branched neurites. Using enzyme-linked immunosorbant assays and blocking antibodies, we have shown that this effect is due to the release of brain derived neurotrophic factor and nerve growth factor, which were up-regulated in differentiated mesenchymal stem cells following co-culture. The relevance of the tyrosine kinase receptors was confirmed by the selective tyrosine kinase inhibitor, K252a which abolished the neurite outgrowth of the dorsal root ganglia neurons when co-cultured with the differentiated mesenchymal stem cells similar to Schwann cells. The results of the study further support the notion that mesenchymal stem cells can be differentiated and display trophic influences as those of Schwann cells.

Growth factors in mesenchymal stem cells following glial-cell differentiation.

Schwann cells are essential facilitators of peripheral nerve regeneration following injury, as they provide physical support and guidance. In vitro these supporting cells are slow-growing and hence are not well suited to a tissue-engineering approach to nerve repair. We have differentiated rat bone-marrow-derived mesenchymal stem cells into Schwann-cell-like cells using a cocktail of growth factors, including glial growth factor-2. Qualitative reverse transcription-PCR, Western-blotting and immunocytochemical approaches were used to investigate the mRNA transcript levels and protein expression of glial cell markers and neurotrophic factors in differentiated mesenchymal stem cells compared with the levels found in Schwann cells (which acted as a positive control). The results showed that differentiated mesenchymal stem cells expressed transcripts and proteins for the specific glial growth receptor 2, erbB3 and neurotrophic factors, nerve growth factor, brain-derived neurotrophic factor, glial-derived neurotrophic factor and leukaemia inhibitory factor. Expression of these growth factors provides further evidence that differentiated mesenchymal stem cells appear to have cellular and molecular characteristics similar to those of Schwann cells.

Adipose-derived stem cells differentiate into a Schwann cell phenotype and promote neurite outgrowth in vitro.

Experimentally, peripheral nerve repair can be enhanced by Schwann cell transplantation but the clinical application is limited by donor site morbidity and the inability to generate a sufficient number of cells quickly. We have investigated whether adult stem cells, isolated from adipose tissue, can be differentiated into functional Schwann cells. Rat visceral fat was enzymatically digested to yield rapidly proliferating fibroblast-like cells, a proportion of which expressed the mesenchymal stem cell marker, stro-1, and nestin, a neural progenitor protein. Cells treated with a mixture of glial growth factors (GGF-2, bFGF, PDGF and forskolin) adopted a spindle-like morphology similar to Schwann cells. Immunocytochemical staining and western blotting indicated that the treated cells expressed the glial markers, GFAP, S100 and p75, indicative of differentiation. When co-cultured with NG108-15 motor neuron-like cells, the differentiated stem cells enhanced the number of NG108-15 cells expressing neurites, the number of neurites per cell and the mean length of the longest neurite extended. Schwann cells evoked a similar response whilst undifferentiated stem cells had no effect. These results indicate adipose tissue contains a pool of regenerative stem cells which can be differentiated to a Schwann cell phenotype and may be of benefit for treatment of peripheral nerve injuries.