Effects Of P(DLLA-co-TMC)/FK506 Microsphere Administration In Different Time On Regeneration After Allogeneil Graft In A Rodent Model

The repair of peripheral nerve injury has been constantly a hot topic of the basic and clinical research, Many scholars study the research tirelessly from different materials, the bridge received a peripheral nerve tissue-engineered graft for repair of peripheral nerves, drugs to promote axonal regeneration and other aspects, But the treatment is still the focus from the autologous nerve graft and drugs in a bid to promote axonal regeneration. However, the existence of autologous nerve graft is restricted for donor source,the district sensory and motor dysfunction of the shortcomings of . As the use of immunosuppressive agents, Allogenic nerve graft has been a great development . At present in the experimental stage of the new immunosuppressant FK506 has been recognized as a drug of first choice in this area. In this study, FK506, and select P (DLLA-co-TMC) copolymer material into P (DLLA-co-TMC) / FK506 nanospheres, In conditions of nerve allografts, approach FK506 nanospheres'role in promoting nerve regeneration and the best delivery time.【Objectives】1. The Preparation and Properties research of P (DLLA-co-TMC) / FK506 Nanoparticles.2. study the effects of Allogeneic transplantation of P (DLLA-co-TMC) / FK506 nanosphere to promote nerve regeneration and the best use of time【Methods】1. A single emulsion - solvent evaporation method (O / W) Preparation of P (DLLA-co-TMC) / FK506 nanospheres, Weigh accurately 100 mg P (DLLA-co-TMC) and a certain amount of FK506 dissolved in 4 ml dichloromethane, Stirring slowly with the syringe into the polymer solution 40 ml 2% PVA (w / v) aqueous solution, Solvent evaporation under vacuum at room temperature stirring 4 ~ 6 h, Microspheres obtained for centrifugal separation, washed with distilled water several times, try to remove the surface of PVA microspheres, Freeze-dried (ALPHA2-4 freeze dryer, CHRIST) 48 h, to be a white powder P (DLLA-co-TMC) drug-loaded microspheres stored at room temperature dryer spare.2. Allograft SD rat tibial nerve, Immediately after operation, respectively, 24 hours,3 days would be P (DLLA-co-TMC) / FK506 nanosphere placed locally around the nerve anastomosis, Press release rate a 1㎎ / kg / d free for 30 days, Observed partial response and parcel of connective tissue at different time points after operation, Determination of ipsilateral triceps muscle wet weight, in order to know muscle atrophy and motor function recovery.3. HE sections of the transplanted nerve staining, light microscopy observation of the growth of nerve and determination of the number of myelinated fibers.4. Was drawn 5 days before the transplant distal nerve beyond 5 mm injected into true blue (true blue) retrograde labeling neurons were observed under a fluorescence microscope to measure labeled the total number of neurons.5. All right tibial nerve sciatic nerve branches are outside the department in the opening cut, with Medtronic EMG recording electrodes placed in the analyzer calf triceps muscle belly, using acupuncture needles inserted in rat tail and grounded, stimulate electrodes placed on the distal and proximal tibial nerve was stimulated, respectively, arrive at the affected side of the nerve conduction velocity and their respective contralateral comparison.【Results】1. With P (DLLA-co-TMC) / as the encapsulation material, use of O / W load FK506 was prepared polymer microspheres to study the rate of drug concentration on the impact of the load, and determine the mass ratio of polymer and drug 100:10 .2. FK506 nanospheres can inhibit allograft rejection and improved allograft tibial nerve of the local micro-environment.3. Immediate use of FK506 nanosphere group and treatment group 24 hours after 8,12 weeks in the triceps muscle of calf to promote recovery of similar effect, better than the 3 days treatment group and non-treatment group, there are significant differences.and in the three days treatment group and non-treatment group differences were statistically significant.4. FK506 nanospheres promoted the proliferation of Schwann cells, the determination of the number of myelinated nerve fibers in the immediate treatment group and 24-hour treatment group were significantly more than 3 days group and non-treatment group ; and 3 days group and non-treatment group were significantly different, 4,8,12 weeks after operation.5. FK506 nanospheres have a protective effect on neurons, can promote their recovery, true blue mark the number of motor neurons in the immediate treatment group and 24-hour treatment group were significantly more than 3 days group and non - treatment group; and 3 days group and non-treatment group were significantly different, 4,8,12 weeks after operation.6. FK506 nanospheres promote electrophysiological recovery, 12 weeks after operation,the tibial nerve conduction velocity in treatment group than the non-treatment group which greatly fast, but the immediate treatment group and 24-hour treatment group were faster than the 3 treatment group, the difference there are statistically significant.【Conclusion】1. P (DLLA-co-TMC) / FK506 nanosphere the best mass ratio of 100:102. FK506 nanosphere degradation of absorption is good, release rate in line with the law of regeneration after nerve injury.3. Topical FK506 nanospheres significant role in promoting nerve regeneration, but also required fewer doses. 4. Early (24 hours) the use of FK506 to promote nerve regeneration is more evident, delayed three days later decreased...