Background3D-scaffolds have been shown to direct cell growth and differentiation in many different cell types, with the formation and functionalisation of the 3D-microenvironment being important in determining the fate of the embedded cells. Here we used a hydrogel-based scaffold to investigate the influences of matrix concentration and functionalisation with laminin on the formation of the scaffolds, and the effect of these scaffolds on human neural progenitor cells cultured within them.MethodsIn this study we used different concentrations of the hydrogel-based matrix PuraMatrix. In some experiments we functionalised the matrix with laminin I. The impact of concentration and treatment with laminin on the formation of the scaffold was examined with atomic force microscopy. Cells from a human fetal neural progenitor cell line were cultured in the different matrices, as well as in a 2D culture system, and were subsequently analysed with antibody stainings against neuronal markers. In parallel, the survival rate of the cells was determined by a live/dead assay.ResultsAtomic force microscopy measurements demonstrated that the matrices are formed by networks of isolated PuraMatrix fibres and aggregates of fibres. An increase of the hydrogel concentration led to a decrease in the mesh size of the scaffolds and functionalisation with laminin promoted aggregation of the fibres (bundle formation), which further reduces the density of isolated fibres. We showed that laminin-functionalisation is essential for human neural progenitor cells to build up 3D-growth patterns, and that proliferation of the cells is also affected by the concentration of matrix. In addition we found that 3D-cultures enhanced neuronal differentiation and the survival rate of the cells compared to 2D-cultures.ConclusionsTaken together, we have demonstrated a direct influence of the 3D-scaffold formation on the survival and neuronal differentiation of human neural progenitor cells. These findings emphasize the importance of optimizing 3D-scaffolds protocols prior to in vivo engraftment of stem and progenitor cells in the context of regenerative medicine.

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The survival of developing dopaminergic neurons has been shown to be modulated by voltage-dependent mechanisms. Manipulation of these mechanisms in human neural progenitor cell cultures could improve the survival of immature dopaminergic neurons, and therefore aid research into pharmacological and cell replacement therapies for Parkinson's disease. Here, we examined the effect of the Na+ channel agonist veratridine on the human fetal neural progenitor ReNcell VM cell line. Neuronal differentiation was determined by immunocytochemistry, whereas patch clamp recordings showed the expression of functional voltage-gated sodium channels. Our results show that veratridine is neuroprotective in human fetal neural progenitor cells, which may benefit studies investigating neuronal development by reducing premature death amongst developing neurons.