Abstract
We present here a fast protocol that could be used to obtain highly purified cultures of maximal proliferating adult rat Schwann cells. These adult rat Schwann cells can be transfected in a nonbiological way using the physical transfection method of electroporation. Schwann cells are decisive in recovery of peripheral nerves after injury. In a clinical context, the use of enriched adult Schwann cells is necessary for autologous cell transplantation within nerve transplants for peripheral nerve repair. Different parameters such as tissue preparation, culture conditions, and protocols for enrichment, elevation of proliferation rates, and transfection were evaluated in cell cultures harvested from adult rat peripheral nerves. Cell preparation from in vivo predegenerated adult rat sciatic nerves combined with the use of melanocyte growth medium supplemented with forskolin, fibroblast growth factor (FGF)-2, and pituitary extract as a selective, serum-free culture medium, with a secondary cell-enrichment step using specific detachment, resulted in highly enriched cultures of adult rat Schwann cells (>90%) with enhanced proliferation rates (>or=40%). About 20% of these adult Schwann cells could be modified genetically using an optimized electroporation protocol.
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