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.

Cnidarians have long been considered simple animals in spite of the variety of their complex life cycles and developmental patterns. Several cases of developmental conversion are known, leading to the formation of resting stages or to offspring proliferation. Besides their high regenerative and asexual-reproduction potential, a number of cnidarians can undergo ontogeny reversal, or reverse development: one or more stages in the life cycle can reactivate genetic programs specific to earlier stages, leading to back-transformation and morph rejuvenation. The switch is achieved by a variable combination of cellular processes, such as transdifferentiation, programmed cell death, and proliferation of interstitial cells. The potential for ontogeny reversal has limited ecological meaning and is probably just an extreme example of a more general strategy for withstanding unfavourable periods and allowing temporal persistence of species in the environment.Résumé : Depuis longtemps, on considère les cnidaires comme des animaux simples, malgré la variété de leurs cycles biologiques et de leurs patterns de développement complexes. Il existe plusieurs cas de conversions au cours du déve-loppement qui mènent à la formation de stades de repos ou à la prolifération de rejetons. En plus de leur fort potentiel de régénération et de reproduction asexuée, plusieurs cnidaires peuvent subir un renversement de leur ontogenèse ou un développement inversé : un ou plusieurs stades dans le cycle biologique peuvent réactiver des programmes génétiques spécifiques de stades antérieurs, ce qui donne une rétro-transformation et un rajeunissement des morphes. Le revirement est provoqué par une combinaison variable de processus cellulaires, tels que la transdifférentiation, la mort cellulaire programmée et la prolifération de cellules interstitielles. Le potentiel de renversement ontogénique a une signification écologique limitée; il s'agit probablement d'un état extrême d'une stratégie plus générale pour survivre aux périodes défavorables et permettre à l'espèce de se maintenir dans le temps dans son milieu.[Traduit par la Rédaction] Piraino et al. 1754

The metamorphosis of many marine invertebrate larvae is induced by environmental signals. Upon reception of the cues, internal signals have to be set in motion to convey information to all cells of the larvae. For hydrozoan larvae it was hypothesised that ectodermal neurosensory cells at the anterior part are those cells receptive of the inducer. Recently, it was shown that novel peptides with a common GLWamide terminus are found in Cnidaria. These peptides are located in a specific subset of the anterior sensory cells. It was hypothesised that the neuropeptides represent an internal signal coordinating the metamorphic process. In the current study we present further evidence for this hypothesis. Induction of metamorphosis is very specific for the GLWamide terminus and amidation is essential. The potency to metamorphose is strongly correlated with the presence of GLWamide-immunoreactive cell bodies. Our data fit our hypothesis about a very important role of GLWamides in the initiation of the morphogenetic processes very well.

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Hox and ParaHox (H/P) genes belong to evolutionary-sister clusters that arose through duplication of a ProtoHOX cluster early in animal evolution. In contrast to bilaterians, cnidarians express, beside PG1, PG2 and Gsx orthologs, numerous Hox-related genes with unclear origin. We characterized from marine hydrozoans three novel Hox-related genes expressed at medusa and polyp stages, which include a Pdx/Xlox ParaHox ortholog induced 1 day later than Gsx during embryonic development. To reconstruct H/P genes' early evolution, we performed multiple systematic comparative phylogenetic analyses, which identified derived sequences that blur the phylogenetic picture, recorded dramatically different evolutionary rates between ParaHox and Hox in cnidarians and showed the unexpected grouping of [Gsx-Pdx/Xlox-PG2-PG3] families in a single metagroup distinct from PG1. We propose a novel more parsimonious evolutionary scenario whereby H/P genes originated from a [Gsx-Pdx/Xlox-PG2-PG3]-related ProtoHox gene, the "posterior" and "anterior" H/P genes appearing secondarily. The ProtoHOX cluster would have contained the three Gsx/PG2, Pdx/PG3, Cdx/PG9 paralogs and produced through tandem duplication the primordial HOX and ParaHOX clusters in the Cnidaria-Bilateria ancestor. The stronger constraint on cnidarian ParaHox genes suggests that the primary function of pre-bilaterian H/P genes was to drive cellular evolutionary novelties such as neurogenesis rather than axis specification.