Femke Mathot

3 MSC-seeding on processed nerve allografts 45 INTRODUCTION When it is impossible to repair an injured peripheral nerve by direct coaptation of the nerve ends, there are several options that can be used to restore the nerve’s function. These include interposition nerve autografts, processed nerve allografts or bio-absorbable hollow conduits. While many studies have compared these options, the autologous nerve remains the gold standard in clinical practice, especially for restoration of major motor nerves. 1, 2 The limited availability and length of autografts requires the development of a replacement nerve that results in equal functional outcomes. 3 Although their regenerating potency still needs to be improved, processed allografts have shown to be a promising option. 2, 4-6 Stem cells are believed to be an important control element in tissue regeneration by producing proteins and molecules that enhance angiogenesis, inhibit scar formation, stimulate tissue regeneration and have immunomodulatory effects. 7, 8 Their use has demonstrated potential to provide the needed extra biological support to processed nerve allografts. 9-13 Adipose tissue is a valuable source for mesenchymal stem cells (MSCs) from the stromal vascular fraction as it is easily accessible and contains relatively large amounts of rapidly proliferating MSCs. 14-18 Schwann cells, the original facilitators of nerve regeneration, have been confirmed as even better providers of biological support to processed nerve allografts than MSCs. 19 However, their harvest requires the sacrifice of autologous nerve tissue. Another option to obtain Schwann-like cells is to chemically differentiate MSCs into Schwann-like cells. 20-23 Several in-vitro studies have demonstrated the potential of differentiated Schwann-like MSCs in peripheral nerve repair, resulting in increased neurite outgrowth of motor neurons compared to undifferentiated MSCs. 19, 24, 25 The trophic function of MSCs that results in enhanced gene expression and production of neurotrophic growth factors 24, 26 , does not require delivery of MSCs inside the nerve allograft. Because microinjection and soaking of MSCs damage both the cells and the allograft while reducing the number of uniformly attached cells, dynamic seeding offers a promising cell delivery strategy that has not been evaluated for differentiated MSCs. 27-29 The purpose of this study was to determine the ability of MSCs differentiated into Schwann- like cells to be dynamically seeded on decellularized nerve allografts and to compare their seeding potential to that of control MSCs. This was investigated by three different sub- studies that aimed to evaluate (I) the influence of processed nerve grafts on the viability of differentiated MSCs, (II) the seeding potential of Schwann-like MSCs on processed nerve grafts and temporal optimization of seeding duration, and (III) the survivability, distribution and migration of differentiated MSCs after seeding. MATERIALS AND METHODS This study was approved by the IACUC institutional review committee and the Institutional