Schwann cells loaded neural tissue engineering scaffold combined pulsed electromagnetic field in vivo promote sciatic nerve regeneration in rats

Abstract

Objective Under proper pulsed electromagnetic field(PMF, 50 Hz, 2 mT, 4 h)exposure, to observe the regenerative effects of Schwann cells(SCs)loaded nerve scaffold bridging 12 mm sciatic nerve defects in rats. Methods SD male rats were divided randomly into five groups as follows: Autograft group, Scaffold group, Scaffold+PMF group, Scaffold+SCs group, and Scaffold+SCs+PMF group. Then, the collagen-chitosan nerve scaffolds were fabricated by I type collagen and chitosan in condensation, and SCs were loaded into the nerve scaffold to form neural tissue engineering material. Then, the 12 mm sciatic nerve defects of rats were bridged using the material. After surgery, the rats were exposed under PMF 4 hours every day. Three, seven and fourteen days after surgery, the survival of SCs in nerve scaffold was tested. Four, eight and twelve weeks after surgery, the neurologic function recovery and morphometric analysis were performed. Meanwhile, RT-PCR test of neurotrophic factors was analyzed one and three weeks after operation. Results Three, 7, and 14 days after surgery, SCs transplanted into scaffolds were in good condition, the number of Live-labeled SCs in nerve scaffold under PMF were 85.28%±2.47%, 88.41%±1.39%, and 92.73%± 2.36%, which were significantly higher than that without PMF, respectively. Twelve weeks after surgery, Scaffold+SCs+PMF group was observed massive axons regenerated into scaffolds by TEM, and the total area of regenerated axons, the total number of myelinated axons, the mean diameter of the myelinated axons, and the degree of myelination(G-ratio)were in the similar range between Scaffold+SCs+PMF group and Autograft group, which were significantly higher than those in the remaining groups. Twelve weeks after surgery, the morphological analysis of gastrocnemius muscles showed that the percentage of muscle fiber area in the Scaffold+ SCs+PMF group was 84.95%±3.92%, which was in a similar range of Autograft group(86.52%±4.38%), and was significantly higher than the remaining groups. Four, 8, and 12 weeks after surgery, the number of FG-labeled motoneurons and sensory neurons were in the similar range between the Scaffold+SCs+PMF group and Autograft group, which were significantly higher than those in the remaining groups. Four, 8, and 12 weeks after surgery, the peak amplitude of CMAP, latency of CMAP latency and NCV value were in the similar range between the Scaffold+SCs+PMF group and Autograft group, which were significantly higher than those in the remaining groups. RT-PCR results showed the mRNA levels of BDNF, GDNF and VEGF in the Scaffold+SCs+PMF group were significantly higher than those in other groups without Autograftc group. Conclusion Under proper PMF exposure, SCs loaded nerve scaffold bridging the sciatic nerve defects could enhance functional recovery of the injured nerve, and promote nerve regeneration in rats. Key words: Sciatic nerve; Schwann cells; Tissue engineering; Electromagnetic fields