Studies On The Repair Of Acute Spinal Cord Injury By Transplantation Of Olfactory Ensheating Glia

It has been proved that olfactory ensheating glia (OEG) can promote the regeneration of spinal cord axons by excreting several factors and forming myelin sheathes for regenerating axons. There were some experiments about the functional recovery of OEG transplantation on the complete spinal cord transection, semisection and electrolytic lesions, but there were little research on the OEG transplantation in spinal contusion. Because there are some remaining nerve fibres in spinal contusion, a transection at the contusion area was made to ensure all the nerve fibers were cut completely within the area. Objectives: One of the aims of the current study was to search for a simple method for OEG culture and purification. The other was to observe OEG survival and repair in vivo for spinal cord contusion and transection after the OEG transplantation.Methods: 1. Cell culture: The primary OEG was cultivated with the olfactory bulb of Wistar neonate rats, and then the purification was done by free serum medium. As a result, the majority of neuron and other mixed cells were eliminated. Because only OEG express both GFAP and P75 in the CNS, we can identify it by this characteristic.2. Animal experiment: The T9-10 spinal cords contusion of 4 months Wistar rats was made with the NYU impactor, then complete transection was performed in the middle of the contusion area in Group A and Group B. OEG labeled by Hoechst was transplanted in group A (20 rats). In group B (20 rats), DMEM were injected. In Group C (10 rats), T9-10 laminectomies were done without cord contusion and transection. The functional recoveryof the spinal cord injury was observed with BBB locomotion score in each double weeks. The tissue sections were done 3 months postoperatively. HE staining, silver staining and NF immunohistochemical staining were performed respectively to observe the pathological changes and axon regeneration. The survival of OEG, which was labeled by Hoechst, was observed under the fluorescence microscope. Results:1. OEG proliferated quickly by free serum medium, and cell count increased by 16 times of primary culture through 9 days cultivation. Its purity is 85%.2. The experiment showed that locomotion behaviour had improved 4 weeks postoperatively. The BBB locomotion score of group A is higher than that of group B in all periods, statistically significant difference in 12 week.3. Nerve fiber innervated injury area in the group A and B. In the cephal lesion, the number of nerve fibers of group A was more than that of group B(P(0.001), but less than that of group C(P(0.001). 4. In group A, the number of nerve fiber of the caudal lesions was less than that of the cephal lesion(P(0.001). In the group B, there were few nerve fibers in the caudal part of lesion.5. A great number of OEG labeled by Hoechst were observed around spinal injury area through the fluorescence microscope. OEG can migrate to 2cm far from the injury area.6.There was positive correlation between the number of regeneration axons and the recovery of locomotion function (r=0.9816).Conclusions:1. The cell culture of free serum medium can obtain a great number of high purity OEG. Furthermore, the method was simple and economical. 2. After transplantation, OEG can survive for at least 3 months in vivo, and can migrate from the injection sites.3. For spinal cord contusion and transection, OEG can promote the axons regeneration and the recovery of locomotion function in our study.