GENETIC MODIFICATION OF HUMAN SURAL NERVE SEGMENTS BY A LENTIVIRAL VECTOR ENCODING NERVE GROWTH FACTOR

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Abstract
Autologous nerve grafts are used to treat severe peripheral nerve injury, but recovery of nerve function after grafting is rarely complete. Exogenous application of neurotrophic factors may enhance regeneration, but thus far the application of neurotrophic factors has been hampered by fast degradation after local application and unwanted side effects after systemic application. These problems may be overcome with the use of lentiviral (LV) vectors that direct sustained local transgene expression in cells. Human sural nerve segments were either submerged in or injected with LV vectors encoding green fluorescent protein and cultured in vitro. Production of nerve growth factor (NGF) by nerve segments after injection of LV-NGF was quantified. The effect of NGF produced by LV-transduced fibroblasts derived from human sural nerve segments was assessed on neurite outgrowth in vitro. The injection of vector into nerve segments is a more effective way to deliver the vector than submersion of the nerve in vector-containing medium, leading to large numbers of transduced fibroblasts over a significant extent inside the nerve. The injection of LV-NGF leads to a gradual increase of NGF production, reaching a plateau after 4 days. LV-NGF-transduced human fibroblasts promote neurite outgrowth in vitro. We have developed a method to transduce cells in human sural nerve segments with LV vector. This approach holds promise as a powerful novel adjuvant therapy for peripheral nerve surgery and can be performed without changing the routine practice of nerve grafting.