Emma East

Alignment of Astrocytes Increases Neuronal Growth in Three-Dimensional Collagen Gels and Is Maintained Following Plastic Compression to Form a Spinal Cord Repair Conduit

After injury to the spinal cord, reactive astrocytes form a glial scar consisting of highly ramified cell processes that constitute a major impediment to repair, partly due to their lack of orientation and guidance for regenerating axons. In some nonmammalian vertebrates, successful central nervous system regeneration is attributed to the alignment of reactive glia, which guide axons across the lesion site. Here, a three-dimensional mammalian cell-seeded collagen gel culture system was used to explore the effect of astrocyte alignment on neuronal growth. Astrocyte alignment was mapped within tethered rectangular gels and was significantly greater at the edge and middle of the gels compared to the control unaligned regions. When neurons were seeded on and within astrocyte gels, neurite length was greatest in the areas of astrocyte alignment. There was no difference in expression of astrocyte reactivity markers between aligned and control areas. Having established the potential utility of astrocyte alignment, the aligned gels were plastic compressed, transforming them into mechanically robust implantable devices. After compression, astrocytes remained viable and aligned and supported neurite outgrowth, yielding a novel method for assembling aligned cellular constructs suitable for tissue engineering and highlighting the importance of astrocyte alignment as a possible future therapeutic intervention for spinal cord repair.

A versatile 3D culture model facilitates monitoring of astrocytes undergoing reactive gliosis

A major impediment to CNS repair is the glial scar, which forms following damage and is composed mainly of ramified, ‘reactive’ astrocytes that inhibit neuronal regrowth. The transition of astrocytes into this reactive phenotype (reactive gliosis) is a potential therapeutic target, but glial scar formation has proved difficult to study in monolayer cultures because they induce constitutive astrocyte activation. Here we demonstrate a 3D collagen gel system in which primary rat astrocytes were maintained in a persistently less reactive state than comparable cells in monolayer, resembling their status in the undamaged CNS. Reactivity, proliferation and viability were monitored and quantified using confocal, fluorescence and time‐lapse microscopy, 3D image analysis, RT–PCR and ELISA. To assess the potential of this system as a model of reactive gliosis, astrocytes in 3D were activated with TGFβ1 to a ramified, reactive phenotype (elevated GFAP, Aquaporin 4, CSPG, Vimentin and IL‐6 secretion). This provides a versatile system in which astrocytes can be maintained in a resting state, then be triggered to undergo reactive gliosis, enabling real‐time monitoring and quantitative analysis throughout and providing a powerful new tool for research into CNS damage and repair. Copyright

A 3D in vitro model reveals differences in the astrocyte response elicited by potential stem cell therapies for CNS injury

AIM This study aimed to develop a 3D culture model to test the extent to which transplanted stem cells modulate astrocyte reactivity, where exacerbated glial cell activation could be detrimental to CNS repair success. MATERIALS & METHODS The reactivity of rat astrocytes to bone marrow mesenchymal stem cells, neural crest stem cells (NCSCs) and differentiated adipose-derived stem cells was assessed after 5 days. Schwann cells were used as a positive control. RESULTS NCSCs and differentiated Schwann cell-like adipose-derived stem cells did not increase astrocyte reactivity. Highly reactive responses to bone marrow mesenchymal stem cells and Schwann cells were equivalent. CONCLUSION This approach can screen therapeutic cells prior to in vivo testing, allowing cells likely to trigger a substantial astrocyte response to be identified at an early stage. NCSCs and differentiated Schwann cell-like adipose-derived stem cells may be useful in treating CNS damage without increasing astrogliosis.