An Optimized Collagen-Fibrin Blend Engineered Neural Tissue Promotes Peripheral Nerve Repair

Tissue engineering approaches in nerve regeneration often aim to improve results by bridging nerve defects with conduits that mimic key features of the nerve autograft. One such approach uses Schwann cell self-alignment and stabilization within collagen gels to generate engineered neural tissue (EngNT). In this study, we investigated whether a novel blend of fibrin and collagen could be used to form EngNT, as before EngNT design a beneficial effect of fibrin on Schwann cell proliferation was observed. A range of blend formulations was tested in terms of mechanical behavior (gel formation, stabilization, swelling, tensile strength, and stiffness), and lead formulations were assessed in vitro. A 90% collagen 10% fibrin blend was found to promote SCL4.1/F7 Schwann cell viability and supported the formation of aligned EngNT, which enhanced neurite outgrowth in vitro (NG108 cells) compared to formulations with higher and lower fibrin content. Initial in vivo tests in an 8 mm rat sciatic nerve model using rolled collagen-fibrin EngNT rods revealed a significantly enhanced axonal count in the midsection of the repair, as well as in the distal part of the nerve after 4 weeks. This optimized collagen-fibrin blend therefore provides a novel way to improve the capacity of EngNT to promote regeneration following peripheral nerve injury

Improved osteogenic vector for non-viral gene therapy

Therapeutic compensation of deficient bone regeneration is a challenging task and a topic of on-going search for novel treatment strategies. One promising approach for improvement involves non-viral gene delivery using the bone morphogenetic protein-2 (BMP-2) gene to provide transient, local and sustained expression of the growth factor. However, since efficiency of non-viral gene delivery is low, this study focused on the improvement of a BMP-2 gene expression system, aiming for compensation of poor transfection efficiency. First, the native BMP-2 gene sequence was modified by codon optimisation and altered by inserting a highly truncated artificial intron (96 bp). Transfection of multiple cell lines and rat adipose-derived mesenchymal stem cells with plasmids harbouring the improved BMP-2 sequence led to a several fold increased expression rate and subsequent osteogenic differentiation. Additionally, comparing expression kinetics of elongation factor 1 alpha (EF1α) promoter with a state of the art CMV promoter revealed significantly higher BMP-2 expression when under the influence of the EF1α promoter. Results obtained by quantification of bone markers as well as osteogenic assays showed reduced sensitivity to promoter silencing effects of the EF1α promoter in rat adipose-derived mesenchymal stem cells. Finally, screening of several protein secretion signals using either luciferase or BMP-2 as reporter protein revealed no superior candidates for potential replacement of the native BMP-2 secretion signal. Taken together, by enhancing the exogenous BMP-2 expression system, low transfection efficiencies in therapeutic applications can be compensated, making safe non-viral systems even more suitable for tissue regeneration approaches