| Objective:Spinal Cord Injury (SCI) is one of the most devastating forms of trauma cases, with the characteristics of high incidence and high deformity. To promote the nerve regeneration and restoration is the research emphasis in the field after spinal cord injury. It is the key point that collapse of the growth cone can inhibit the axon growth. Spinal cord repair strategies for promoting the axon growth and minimizing inhibitory microenvironment effect are becoming a significant research topic in the field of spinal cord restoration nowadays. It is known that cytoskeleton composed with actin filament and microtubule plays an important role in the process of nerve growth and growth cone orientation. NM II is in the downstream of RhoA/Rock signaling pathway. The activated NM II can promote the collapse of the growth cone and inhibit the axon extension by regulating the structure of cytoskeleton. At the same time, some studies have found that gene silencing or chemical inhibition of NM II can obviously improve the survivability of the mammalian cells. The objective of this study is to deliberate the influence of regulating the structure of growth cone of gene silencing NM II on axon regeneration and nerve cell viability by preparing for rat spinal cord injury model and evaluate the effects on functional recovery through the function score and electrophysiology experiment of rats.Method:80female Wistar rats were randomized to divide into four groups, that is, blank control group, the empty carrier injection control group, siRNA injection group1and siRNA injection group2,20respectively. Preparing the rat thoracic spinal cord injury Model (10g*25mm) using the Impactor Model-II spinal cord compression apparatus. Normal nursing and diet were given to the rats after injury and built lentivirus vectors to make NMII gene silencing in vivo. The rats were killed after injury for12weeks. NMII expression level was detected by Western-blot technique, thus evaluating NMII gene silencing efficiency in vivo. The axon regeneration, microstructure and nerve cells viability in each group was observed with immunohistochemical method including the expression of NF200, GFAP, MAP2and SYN. The influence on neural function repair of NMII gene silencing was observed with the Basso, Beattie and Bresnahan score (BBB score).Results:We succeeded in building the lentivirus vectors. NMII expression level was more than95%in siRNA injection groupl, whereas was80%in siRNA injection group2. The expression of each cells were identified by immunostaining:neurons by NF200, axon by MAP2, growth cone by NeuN and synaptic connections by SYN. They showed a much more apparent high expression level in siRNA injection group1(p<0.05), but astroglia that was positive for GFAP showed no significant difference in four groups (p>0.05). The Basso, Beattie and Bresnahan score(BBB score)showed a better functional recovery of hindlimbs in siRNA injection groupl (p<0.05)Conclusion:After spinal cord injury, except for inhibiting the proliferation of astroglia, NMII gene silencing can promote the viability and regeneration of nerve cells significantly together with the extension of axon, thus forming more synaptic connections and promoting functional recovery of hindlimbs. |
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