Inducing And Activation Of Schwann Cells And Construction Of Nerve Bridge

Self- neural transplantation is the first method after injury of peripheral nerve ,but limited donator ,sense of donor site , motor dysfunction, long time of operation are all the defects and are difficulted in satisfying with long segment neurologic defect. Therefore, finding ideal seed cells and cage materials to construct functional tissue engineering peripheral nerve is the main method for solving suffering ofneurologic defect. It is important how to obtain mass of segmentation and proliferation of SCs and choice suitable cage materials for constructing functional tissue engineering peripheral nerve. SCs are induced and stimulated by cell cultural biological agent IL-1β in this experiment. To grope the best action condition of segmentation and proliferation of SCs after inducing and obtain ideal seed cells. Meanwhile, as new organism degradation materials, human hair keratins can make up some detects of synthetic depolymerize , for example, PGA, PLA and PLGA, etc. Because these synthetic depolymerizes have pykno- degradation , collapse, cage body sink, local higher acidity products. Implanting tissue engineering peripheral nerve bridge into the defect of sciatic nerve , and to explore the best experimental condition of HHKs three dimentional cultured with SCs after inducing by IL-16 for construction of tissue engineering peripheral nerve bridge.[Objectives]1. To explore IL-16 expression of macrophages activated by self-neural homogenate in vitro culture.2. To explore IL-18 induce and activate on SCs and the best action concentration of IL-1B.3. To explore the establishment of three dimentional culture of HHKs and SCs before or after inducing by IL-1B for construction of tissue engineering peripheral nerve bridge.[Methods]1. Conditioned medium of macrophages inactivated can be obtained by injecting in abdominal cavity without sciatic nerve injury. Cut distal tip of sciatic nerve in one-month SD rat, fore-degeneration after two days, to make homogenate of nerve and inject it into itself abdominal cavity. Three days later, to draw fluid in abdominal cavity to culture macrophages. The level of IL-18 in the conditioned media was determined by an enzyme-linked immunoadsordent assay.2. Screened the best concentration of IL-18 which promoted proliferation of Schwann cells by MTT. To detect the proliferation of Schwann cells by incorporated radioactive methyl-3H into culture. Enzyme-linked immunoadsordent assay (ELISA) was detected in group which effected on NGF excretion of Schwann cells.3. SCs were not induced by IL-18 and were purified by primary culture and were marked by Brdu, which were three dimentional cultured with HHKs which were decorated by ECM for construction of tissue engineering peripheral nerve bridge, and implant it into the defect of sciatic nerve , down skin, skeletal muscle of SD rat . Observing morphology of SCs were cultured with HHKs by scanning electron microscope and evaluated SCs in immunocytochemistry staining with anti-S-100. Observing the growth of SCs in vivo which marked by Brdu in vitro with immunocytochemistry staining through paraffin section.4. SCs were induced by IL-18 and were three dimentional cultured with HHKs which were decorated by ECM for construction of tissue engineering peripheral nervebridge, and implant it into the defect of sciatic nerve of SD rat. Observing the growth and proliferation of SCs and positive expression of NGF by hybridization in situ. [Results]1. Changing of cell's ultrastructure occurred and IL-6 was expressed in induced macrophage cells.2. IL-6 can be more promoted segmentation and proliferation of Schwann cells and higher expression of NGF.3. SCs were not induced by IL-8 and were cultured with HHKs which were not modificated by ECM, SCs were all killed. SCs were not induced by IL-6 and were cultured with HHKs which were modificated by ECM, SCs were not all killed, the parts of them can live and sticky on the HHK, and implant it into the defect of sciatic nerve, down skin, skeletal muscle of SD rat, SCs can be conglutinated on HHKs and grew well.down skin, skeletal muscle of SD rat were occured in artificial nerve4. Induced SCs can be more satisfier than un-induced SCs for seed cells of tissue engineering, and lots of SCs can be seen survived and proliferated on HHKs. Implanting it into the defect of sciatic nerve , four weeks later , positive expression of NGF of induced SCs in the defect of sciatic nerve by hybridization in situ and HHKs' degradation in vivo.[conclusion]l.IL-18 expression was highly through inducing and activating macrophages which swallowed self-neural homogenate.2. Schwann cells can be expressed NGF by itself, while 2.0ng/ml IL-16 can be more promoted segmentation and proliferation and higher expression of Schwann cells.3. Induced SCs can be more better than other methods in three dimentional culturingwith HHKs which were modificated by ECM and to construct peripheral nerve oftissue engineering.