Functional nerve regeneration after reconstructive nerve surgery remains unsatisfying. In this study, vascular endothelial growth factor (VEGF) gene therapy combined with a hyaluronic acid (HA)-enriched microenvironment in nerve regeneration was investigated. Sciatic nerve was transected, and end-to-end neurorrhaphy was performed on 32 male Sprague-Dawley rats, which were randomly divided into four groups (n = 8 per group): nerve coaptation without treatment (group I); nerve coaptation covered with HA film sheath (group II); nerve coaptation with intramuscular VEGF gene in plasmid injection (group III); and nerve coaptation combined with HA film sheath and intramuscular VEGF gene in plasmid injection (group IV). Contralateral sciatic nerves were used as control. VEGF expression was verified from gluteal muscle biopsies surrounding the sciatic nerve by reverse transcriptase-PCR. Electrophysiological, histopathological, and electron microscopic evaluations were performed after 4 weeks. Mean peak amplitude of groups I-IV and nonoperated sciatic nerve were 4.5 +/- 0.6 mV, 6.4 +/- 0.4 mV, 6.7 +/- 0.5 mV, 8.5 +/- 0.4 mV, and 9.8 +/- 0.5 mV, respectively. Mean myelinated axonal counts of groups I-IV and nonoperated sciatic nerve were 105 +/- 24, 165 +/- 19, 181 +/- 22, 271 +/- 23, and 344 +/- 17, respectively. Treatment with HA film sheath coverage combined with intramuscular VEGF gene in plasmid injection yielded statistically significant higher peak amplitudes and myelinated axonal counts (P < 0.001). In addition, significantly less scar formation with HA administration (groups II and IV; P < 0.001) was found. Thus, it was found that VEGF might crucially regulate nerve regeneration processes and that HA can reduce the scar formation. This study showed that the combination of HA film sheath and VEGF gene may synergistically promote peripheral nerve regeneration. (c) 2013 Wiley Periodicals, Inc. Microsurgery 34:209-216, 2014.