Development of a reconstituted nerve in a three-dimensional collagen gel under tension populated with fibroblasts and Schwann cells

Abstract

Our aim is to devise an artificially reconstituted nerve segment made of a three-dimensional collagen gel populated with aligned fibroblasts and Schwann cells. Collagen lattices were prepared by mixing concentrated medium, a type I collagen solution and rat Schwann cells (SC), rat neural fibroblasts (nF) or human dermal fibroblasts (dF) and allowed to polymerize at 37 °C. In these free-floating lattices, nF and dF retracted the gel more than SC. All cells appeared to be elongated and oriented at random. Rat cells obtained by enzymatic digestion of nerves undergoing wallerian degeneration retracted the gel at a larger extent than cells from intact nerves. Rectangular lattices restrained at each extremity acquired a paraboloid shape upon retraction by neural or dermal F reflecting the mechanical tension developed by these cells on their support. Adult SC alone produced a faint paraboloid even at high cell density while SC associated with nF developed a paraboloid similar to that obtained with nF alone. The mechanical force developed by dermal F and SC in the restrained lattice was measured by strain gauges and found much higher for F than for SC. In restrained lattices, both types of F were elongated and aligned to the long axis of the gel while SC elongated but not necessarily in a parallel fashion. The central portion of a mixed nF-SC collagen restrained lattice produces a flattened cylindric segment made of longitudinally oriented col-lagen fibrils, F and SC, which could represent a promising material for preparation of nerve grafts. An original plastic mould was devised to allow the preparation of cylindrical segments of free or restrained collagen lattices in view of in vitro and in vivo regeneration studies.