Experimental Study Of ChABC And GDNF And Nogo-A Antibody Combined Treatment For Injuried Spinal Cord Of Rat

Objective: 1. To determine whether Nogo-A antibody has the effect of inhibititor-neutralizing in vitro. 2. To study the effect of strategy combined with ChABC , GDNF and Nogo-A antibody on the regeneration and functional recovery of transected spinal cord. Method: 1. To determine whether Nogo-A has the effect of inhibitor-neutralizing, one standard bioassays that is spreading of 3T3 fibroblasts in vitro is used. Fibroblast spreading is strongly inhibited on culture dishes coated with CNS myelin or a protein extract enriched in Nogo-A(q-pool). When the inhibitory activity is neutralized by Nogo-A antibody, 3T3 spreading is restored to a large extent. 2. Ninety-two adult SD female rats, ranging from 200g to 250g, are randomly divided into 10 groups: (1)normal control group(n=6); (2) sham operation group(n=6) in which the PE-10 tube is placed into subarachnoid space and the spinal cord is remained intact with treatment of NS(saline)by perfusion; (3) spinal cord transection group(n=10) in which the spinal cord is transected at T7-8 completely, then PE-10 tube is placed in subarachnoid space below the injury level for perfusion of NS(saline); (4)spinal cord transection combined with treatment of ChABC by perfusion (n=10); (5)spinal cordtransection combined with treatment of GDNF by perfusion (n-10); (6)spinal cord transection combined with treatment of Nogo-A antibody by perfusion (n=10); (7)spinal cord transection combined with treatment of ChABC and GDNF by perfusion (n=10); (8)spinal cord transection combined with treatment of ChABC and Nogo-A antibody by perfusion (n=10); (9)spinal cord transection combined with treatment of GDNF and Nogo-A antibody by perfusion (n=10); (lO)spinal cord transection combined with treatment of ChABC, GDNF and Nogo-A antibody by perfusion (n=10). The normative nursing project is employed to decrease both the complication and the death rate of rats following transection of spinal cord. Behavioral evaluation, SEP(somatosensory evoked potential), BDA tracing, Immunohistochemical analysis: NF-200 , GFAP,GAP-43 immunohistochemistry are adopted to assess the animals, surviving for 6m after operation. The behavioral evaluation are analyzed by two-way ANOVA. NF-200, GAP-43 and GFAP positive expression are analyzed quantitatively with a computer image analysis system, and the results are analyzed respectively by one-way ANOVA. Results: 1., Nogo-A antibody exhibit strong neutralizing activities in the 3T3 fibroblast spreading at a concentration of 18ug/ml and 30ug/ml (p <0.05). 2. The Mean Score of Behavioral evaluation of hindlimbs is 53.5 in the normal group, but it is only 11.0±1.47 in transected spinal cord injury group treated only with saline after 4w. There is an apparent increase of scores in every treatment group compared with that of transected spinal cord rats only. (p<0.05). The scores are significantly different between AGN group and other treatment groups, (p<0.05). 3. SEP: 6m after operation, the latent period of SEP are delayed in AGN group, whereas other transectedgroups has no sign of wave recorded. The differences are significant (p< 0.05); at the same time, the pathologic changes of spinal cord, including the decrease of neurons, the hyperplasia of gliacyte and a large number of cavities are obvious in the injury site. In addition, the phenomena are more obvious in gray matter than in white matter, and also more serious in the caudal than in the rostral. 4.NF-200: Compared with the control group, NF-200 expression increase significantly in A, AG, AN, AGN groups at the 24th week; compared with G,N,GN groups, expression increase significantly in AGN group(p<0.05). There is no significant difference between control group and G, N, GN groups respectively. 5.GAP-43: Compared with the control group, expression of GAP-43 in A, AG, AN, AGN groups increase significantly (p<0.05). Expression is higher in AGN group than that of other treatment groups(p<0.05). 6.GFAP: Compared with every treatment group, GFAP expression in control group increase significantly(p<0.05);There is no significant difference among the treatment groups. Conclusions: 1. Nogo-A antibody has the effect of inhibitor-neutralizing in vitro. 2. The results show that treatment combined with ChABC, GDNF and Nogo-A antibody can promote the regeneration of injuried spinal cord and the recovery of hind limb function more evidently than other treatment groups. 3. ChABC, GDNF, Nogo-A antibody used alone or combined with each other is helpful for the regeneration of nerve fibers. But there is no functional regeneration of nerve fiber. 4. The functional recovery of rat with transected spinal cord may be related to functional compensation of caudal spinal cordo...