Femke Mathot

10 General discussion and future perspectives 177 Translation to bigger nerve gap models Rats are often used as animal-model to perform fundamental studies prior to translation to a bigger animal. We choose to do so as well as they can be easily housed, are relatively cheap, they can endure anesthesia and surgery quite well, plenty of rat-specific reagents (needed for qPCR analysis for example) are available and most nerve studies have been performed in rats making results easy comparable to other studies. The major disadvantage of using a rat-model is their described superposed neuroregenerative capacities compared to humans, particularly due to the limited nerve gap that can be obtained. 50, 51 After fundamental studies have been performed to clarify details of the neuroregenerative capacity of MSCs in vivo, we suggest to translate studies to a bigger animal model like the rabbit. Needless to say, considering alternative animal- free study options and critically assessing the necessity and size of another animal-study must precede future animal-studies. Surgical angiogenesis As described in several chapters of this thesis, MSCs demonstrated to improve functional outcomes of peripheral nerve repair due to release of trophic factors that, among other things, enhance angiogenesis. Angiogenesis is postulated to be a crucial element in nerve regeneration and therefore other or complementing strategies to enhance angiogenesis are estimated to be valuable future research subjects. A vital, well vascularized wound bed has shown to be essential for regenerating tissues, providing trophic factors and enhancing outgrowth of blood vessels. 68, 69 A vascularized fat flap that envelopes an implanted decellularized nerve allograft is hypothesized to improve axon regeneration. The blood vessels and the stromal cells (MSC relatives) that are present in the fat tissue might lead to less fibrosis, enhanced vascularization and improved regeneration. Moreover, it would be interesting to study whether this surgical angiogenesis technique can work synergistic with the MSC-seeding technique presented in this thesis. One could hypothesize that a vascularized wound bed leads to more extensive delivery of trophic factors to the MSCs, resulting in extended survival times or enhanced activity/ vitality of MSCs. On their turn, the trophic factors of the MSCs might decrease the immune response resulting in less fibrosis of the fat flap and potentially stimulate the outgrowth of blood vessels from the flap inside the nerve graft. Perhaps this also influences the distribution and orientation of vessels inside the nerve. The hypothesized synergistic effect of MSCs and surgical angiogenesis is illustrated in figure 1 . The effect on peripheral nerve regeneration of stem-cell therapy combined with surgical angiogenesis deserves to be studied in future in vitro and in vivo settings. Immunomodulation Adjuvant therapies like immunomodulation have also demonstrated future potential to improve outcomes of peripheral nerve regeneration. Tacrolimus (FK506) is a regularly used immunosuppressant, often chronically systemically administered after organ transplantation. Besides, it demonstrated to have neuroprotective characteristics by influencing pathways that lead to enhanced GAP43 and NGF synthesis, which hypothetically results in accelerated Wallerian degeneration and axon regeneration. 70 Although this suggests a role for Tacrolimus in peripheral nerve regeneration, substantial systemic doses of Tacrolimus not only results in undesirable immunosuppression, but also in neurotoxicity, inducing peripheral neuropathy.