Identifying the optimal developmental age of human pluripotent stem cell-derived midbrain dopaminergic progenitors for transplantation in a rodent model of Parkinson's disease.

Donor cell age can have a significant impact on transplantation outcomes. Despite the rapid advancement of human pluripotent stem cell (hPSC)-derived dopaminergic (DA) progenitors to the clinic for transplantation into Parkinson's Disease (PD), surprisingly limited data exists regarding the influence of cellular age on neural graft survival, composition, and integration. Here we examined the impact of transplanting ventral midbrain (VM) progenitors at varying days of differentiation (from day 13-30) into a rodent PD model, comparing two hPSC lines (an embryonic and an induced pluripotent cell line, hESC and hiPSC, respectively). Both hPSC lines expressed GFP under the promoter PITX3 enabling specific tracking of graft-derived DA neurons. Post-mortem analysis at 6 months revealed larger grafts from Day19 (D19), D22 and D25 progenitors, yet contained a higher proportion of non-DA and poorly specified (FOXA2-) cells. While D13 and D30 progenitors yielded smaller grafts. D13-derived grafts had the highest DA neuron proportion and proportionally more GIRK2+ DA neurons, the subpopulation critical for motor function. These younger progenitor grafts maintained their capacity to innervate developmentally relevant DA targets, with increased innervation capacity per DA neuron, collectively resulting in restoration of motor deficits with equal or greater proficiency than older donor cells. While donor age effects were reproducible for a given hPSC line and trends were similar between the two hPSC lines, grafts of D13 hiPSC-derived progenitors showed a 6-fold greater density of DA neurons compared to D13 hESC-derived grafts, highlighting between-line variability. These findings show that hPSC-derived VM donor age has a direct impact on graft survival, composition and maturation, and that careful assessment, on a line-to-line basis is required prior to translation.

Using minimalist self-assembling peptides as hierarchical scaffolds to stabilise growth factors and promote stem cell integration in the injured brain.

Neurotrophic growth factors are effective in slowing progressive degeneration and/or promoting neural repair through the support of residual host and/or transplanted neurons. However, limitations including short half-life and enzyme susceptibility of growth factors highlight the need for alternative strategies to prolong localised delivery at a site of injury. Here, we establish the utility of minimalist N-fluorenylmethyloxycarbonyl (Fmoc) self-assembling peptides (SAPs) as growth factor delivery vehicle, targeted at supporting neural transplants in an animal model of Parkinson's disease. The neural tissue-specific SAP, Fmoc-DIKVAV, demonstrated sustained release of glial cell line derived neurotrophic factor, up to 172 hr after gel loading. This represents a significant advance in drug delivery, because its lifetime in phosphate buffered saline was less than 1 hr. In vivo transplantation of neural progenitor cells, together with our growth factor-loaded material, into the injured brain improved graft survival compared with cell transplants alone. We show for the first time the use of minimalist Fmoc-SAP in an in vivo disease model for sustaining the delivery of neurotrophic growth factors, facilitating their spatial and temporal delivery in vivo, whilst also providing an enhanced niche environment for transplanted cells.

Biofunctionalisation of polymeric scaffolds for neural tissue engineering.

Patients who experience injury to the central or peripheral nervous systems invariably suffer from a range of dysfunctions due to the limited ability for repair and reconstruction of damaged neural tissue. Whilst some treatment strategies can provide symptomatic improvement of motor and cognitive function, they fail to repair the injured circuits and rarely offer long-term disease modification. To this end, the biological molecules, used in combination with neural tissue engineering scaffolds, may provide feasible means to repair damaged neural pathways. This review will focus on three promising classes of neural tissue engineering scaffolds, namely hydrogels, electrospun nanofibres and self-assembling peptides. Additionally, the importance and methods for presenting biologically relevant molecules such as, neurotrophins, extracellular matrix proteins and protein-derived sequences that promote neuronal survival, proliferation and neurite outgrowth into the lesion will be discussed.

Macrophages and Microglia Produce Local Trophic Gradients That Stimulate Axonal Sprouting Toward but Not beyond the Wound Edge.

Following injury to the mammalian CNS, axons sprout in the vicinity of the wound margin. Growth then ceases and axons fail to cross the lesion site. In this study, using dopaminergic sprouting in the injured striatum as a model system, we have examined the relationship of periwound sprouting fibers to reactive glia and macrophages. In the first week after injury we find that sprouting fibers form intimate relationships with activated microglia as they traverse toward the wound edge. Once at the wound edge, complicated plexuses of fibers form around individual macrophages. Axons, however, fail to grow further into the interior of the wound despite the presence of many macrophages in this location. We find that the expression of BDNF by activated microglia progressively increases as the wound edge is approached, while GDNF expression by macrophages is highest at the immediate wound margin. In contrast, the expression of both factors is substantially reduced within the macrophage-filled interior of the wound. Our data suggest that periwound sprouting fibers grow toward the wound margin along an increasing trophic gradient generated by progressively microglial and macrophage activation. Once at the wound edge, sprouting ceases over macrophages at the point of maximal neurotrophic factor expression and further axonal growth into the relatively poor trophic environment of the wound core fails to occur.