Femke Mathot

2 Stem cell differentiation in peripheral nerve repair 35 in the outer zone. 48 Dynamic seeding has been successful in vascular tissue engineering and resulted in a more efficient and uniform distribution of cells compared to static seeding with seeding efficiencies ranging from 38% to 90%. 49 This strategy was applied to a nerve-model by Rbia and colleagues. They non-traumatically seeded MSCs on the surface of a processed nerve allograft with the use of a bioreactor. This resulted in a uniform distribution of MSCs that were adhered to the nerve graft. 50 The cells did not migrate into the nerve allograft and the interaction between the MSCs and the nerve surface resulted in an upregulation of neurotrophic factors that potentially enhance nerve regeneration within the nerve graft. 50 Overall, dynamic seeding results in a uniform distribution of MSCs on nerve allografts that enables the cells to interact with the nerve ultrastructure with a high efficiency without harming the cells nor the nerve ultrastructure. To date, this is the most promising delivery method of MSCs to allograft nerves and might form the bridge towards individualized peripheral nerve repair in clinical practice. Jesuraj and colleagues used a concentration of 1 x10^5 cells/5uL to inject and compared it to a concentration of 10^6 cells/5uL. Their analysis revealed an injection efficiency of 10% for the 1x10^6 cells (100.000 cells) and 40% for the 1x10^5 cells (40.000 cells) of which only the larger dose was trackable by in vivo fluorescence. 38 Thompson and colleagues also soaked or injected their 10mm allograft segments with 1x10^6 cells/5uL, but did not report a total efficiency. 48 Rbia and colleagues used 1x10^6 cells to dynamically seed their nerve segments and reported a seeding efficiency of 89.2%, suggesting that almost 900.000 cells were attached to the surface of the 10mm nerve segment before in vivo implementation. 50 Wang and colleagues also used 1x10^6 MSCs, diluted in 1mL fluid, to inject in the gastrocnemius muscle. 46 Despite the wide variety of delivery efficiencies, there seems to be consensus that at least 1x10^6 MSCs need to be presented to the nerve graft to generate noticeable biological effects. However, no studies on the optimal dosing of MSCs have been reported. Dynamic seeding of MSCs appears to be a reliable, effective and well-studied delivery method (see table 1 ). None of the methods of administration reported to date have been specifically tested on differentiated MSCs, so direct comparisons between undifferentiated and differentiated MSCs with respect to their delivery efficiency are lacking. This could be a potential decisive factor as it has been emphasized that differentiation of MSC may decrease their potential to attach to surfaces. 51 It is essential that delivery methods are tested on differentiated MSCs as well as undifferentiated MSCs as the impact on clinical application is significant with respect to cost and time. Differentiated MSCs versus undifferentiated MSCs in vitro The in vitro capabilities of differentiated MSCs in peripheral nerve repair have been extensively evaluated. Kingham and colleagues found that differentiated MSCs significantly extended the number and the lengthof formedneuritesbymotor neuron-likecells compared toundifferentiated MSCs. 11 In another study of Kingham, enhanced expression was observed for nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), glial cell-derived neurotrophic factor (GDNF), vascular endothelial growth factor-A (VEGF-A) and angiopoietin-1 in differentiated MSC compared to undifferentiated MSCs. 17 ELISA analysis demonstrated enhanced secretion