Development of a decellularization method to produce nerve allografts using less invasive detergents and hyper/hypotonic solutions

Abstract

Background Here, we describe a novel method of processing decellularized nerve grafts using osmotic effects of hypotonic and hypertonic solutions and Triton X-100 (a nonionic detergent) and CHAPS (an amphoteric detergent). Materials and methods To evaluate decellularization, the devised method and Hudson's method were compared with respect to remaining cellular components (as assessed by H&E staining and S-100 immunoreactivity) and extracellular matrix structural integrity (as assessed by H&E staining and laminin immunoreactivity) by using rat sciatic nerves. In addition, a 1.5-cm sciatic nerve gap rat model was treated by implanting decellularized nerve grafts prepared using the devised method, Hudson's method, or an autograft to evaluate nerve regeneration. Nerve histomorphometry of distal stumps and wet muscle mass were evaluated at 12 weeks after implantation. Results The devised method produced outcomes similar to those of Hudson's method in terms of cellular component removal, but the devised method was significantly better in terms of ECM preservation. Histomorphometric study showed that the devised method produced significantly fewer nerve fiber and axonal densities than autografting, but much more than Hudson's method. The wet muscle mass of the devised method was also significantly lower than that of autografting, but much higher than that of Hudson's method. Conclusion The described process for producing decellularized nerve grafts yielded better outcomes with respect to peripheral nerve regeneration than the established ionic detergent-based methods in a rat model. This study indicates that decellularized nerve grafts produced in this manner show favorable nerve regeneration used for bridging nerve gaps.