OBJECTIVE To probe the possibility of direct transfer of exogenous gene into peripheral nerve and its following expression in vivo. METHODS The PCMV beta plasmid containing cytomegalovirus (CMV) promoter and Escherichia Coli (E. Coli), beta-Galactosidease (beta-Gal) structural gene (lacZ gene) was constructed and injected into the rabbit sciatic nerve. The control group was injected PBS solution. The injected nerves were sampled and tested by beta-Gal enzyme activity assay of the 5-bromo-4-chloro-3-indolyl-beta-D-galactoside and beta-Gal histochemical stain. RESULTS In the control group, no beta-Gal enzyme activity was detected in the different stages after operation, and beta-Gal histochemical stains showed positive. In the experimental group, enzyme activity could be detected from 2 days to 30 days after operation, and the histochemical stains showed negative. CONCLUSION The exogenous gene can be transferred into peripheral nerve and expressed with bioactivity, thus the gene therapy to accelerate the recovery of nerve is practical.
Nine cases of sciatic nerve from injection hadbeen treated by fasciotomy. The skin temperatureof the diseased limb immediately raised 1-2℃ fol-lowing operation. The cutaneous sensation began torecover 2-3 days after operation. Two cases ofplantar ulcer recovered one month post operation.Five of the eight cases of paralysis of muscle in-nervated by the common peroneal nerve recoveredto normal. The etiology,pathology, and therapeuticmethods of the nerve injury caused by drug injec-tion were discused.
ObjectiveTo observe the changes of microstructure of rats'sciatic nerves with non-freezing cold injury after treated with dexamethasone. MethodsTwelve male Wistar rats were randomly divided into cooling group and treating group.Unilateral sciatic nerves of the rats in the cooling group received cooling treatment with 3-5℃ for 2 hours;while unilateral sciatic nerves of the rats in the treating group received cooling treatment with 3-5℃ for 2 hours and underwent the celiac injection with dexamethasone in addition.The other sciatic nerves were exposed,as the control.The bilateral sciatic nerves of rats in each group were harvested after 24 hours.The microstructure of nerves was examined under the light microscope and electron microscope. ResultsLight microscopic examination revealed extensive myelinated fibre degeneration in form of giant empty axons or shrunken dark axons on the first day after cooled.And the endoneurial capillary lumen was narrowed because of swollen endothelial cells.After the treatment,myelinated fibre degeneration was still similar to that before the treatment,but the endoneurial capillary lumen and endothelial cells were normal.By electron microscopy,sciatic nerves showed extensive myelinated fiber degeneration,and swollen endothelial cells.But unmyelinated fibers and tight junction were preserved on the first days after cooled.Aggregated red cells and platelet thrombus were not found.After the treatment,myelinated fibre degeneration was still similar to that before the treatment.Unmyelinated fibers and tight junction were preserved.The endoneurial capillary lumen and endothelial cells were normal. ConclusionAfter the treatment,the damage of endoneurial capillary had improved,but myelinated fiber degeneration was similar to that before the treatment.It suggested that dexamethasone may only improve the vascular system in non-freezing cold injury of sciatic nerve.
ObjectiveTo fabricate salidroside/collagen/polycaprolactone (PCL) nerve conduit composite and to investigate the effect of composite nerve conduits for repairing sciatic nerve defect. MethodsThe salidroside microspheres were prepared by W/O/W method, and the sustained release rate of microspheres was detected. The microspheres containing 10, 20, and 40 μg salidroside were mixed with collagen to prepare the nerve conduit core layer by freeze-drying method. The shell layer of collagen/PCL scaffold material was fabricated by electrospinning technology. The genipin cross-linked salidroside/collagen/PCL nerve conduit composite was prepared. The structure of nerve conduit was observed before and after cross-linked by scanning electron microscope. Thirty-eight Wistar rats were used to make the right sciatic nerve defect model of 15 mm in length, and randomly divided into groups A, B, C, D (n=9), and group E (n=2), then defect was repaired with the collagen/PCL conduit in group A, autologous nerve in group E, the 10, 20, and 40 μg/mL salidroside/collagen/PCL conduit in groups B, C, and D, respectively. The survival of rats was observed. The sciatic functional index (SFI) was evaluated at 1, 3, and 6 months after operation. At 6 months, the tissue of defect area was harvested for the general, electrophysiology, histological, and immunohistochemical[S-100 and peripheral myelin protein 0(P0)] staining observations. ResultsSalidroside microspheres showed burst release at 3 days, and then it tended to be stable at 13 days and lasted for 16 days, with a cumulative release rate of 76.59%. SEM showed that the disordered fiber of nerve conduit shell layer after crosslinking became conglutination, shrinkage, and density, and had void. The channels of core layer were clearly visible before and after crosslinking. The rats had no infection or death after operation. The SFI of group E was significantly higher than that of groups A, B, C, and D at 1, 3, and 6 months (P<0.05); it was significantly higher in groups B, C, and D than group A (P<0.05), but no significant difference was found among groups B, C, and D at 1 month (P>0.05); there was no significant difference in SFI among groups A, B, C, and D at 3 months (P>0.05); SFI was significantly higher in group C than groups A, B, and D and in groups A and B than group D (P<0.05), but no significant difference between groups A and B (P>0.05) at 6 months. In addition, no significant difference was shown among different time points in the other groups (P>0.05) except groups C and E at 1, 3, and 6 months (P<0.05). The general observation showed that good connection with the thick nerve in groups B and C, and connection with the fine nerves in groups A and D. The conduit materials obviously degraded. Nerve electrophysiological examination showed that the latency/conduction velocity of groups C and E were significantly lower than those of groups A, B, and D (P<0.05), but difference was not significant between groups C and E, and among groups A, B, and D (P>0.05). The histological observation showed that the nerve fiber tissue of groups B, C, and E was obviously more than that of groups A and D, and group C was similar to group E in the nerve fiber arrangement, and the core layer material of each group was completely degraded. Immunohistochemical staining showed that S-100 and P0 proteins expressed in all groups; and the expression level of groups B, C, and E was significantly higher than that of groups A and D, and gradually increased (P<0.05); difference in S-100 expression level was not significant between groups A and D (P>0.05), and P0 expression level of group A was significantly lower than that of group D (P<0.05). ConclusionSalidroside/collagen/PCL nerve conduit can promote sciatic nerve defect repair.
Objective To observe the result of reconstructing quadriceps femoris function in the paraplegia rats by using the 7th cervical nerve root (C7) transposition with autologous and allogeneic neural transplantation. Methods Twenty16-week-old SPF male Wistar rats were adopted to prepare frozen sciatic nerve. Thirty-six Wistar rats were divided into 2 groups (group A and group B, n=18). The left paraplegia model was establ ished with left spinal cord hemisection by the micro scissors under the operation microscope. After the model establ ishment, the homolateral autologous sciatic nerve was bridged with the femoral nerve root by the translocation of C7 in group A, while the allogeneic sciatic nerve was bridged with the femoral nerve root by the translocation of C7 in group B. At 16 weeks and 24 weeks after operation, 9 rats in each group were selected for the neuroelectric-physiological test and then the histomorphology of the nerves was observed under the microscope and the electron microscope. The fresh weight recovery rate of quadriceps femoris was calculated. Results At 16 and 24 weeks after operation, the nerve action-evoked potential (NAP) was (1.14 ± 0.07) mV and (1.21 ± 0.07) mV in group A, and (0.87 ± 0.06) mV and (0.99 ± 0.05) mV in group B; the nerve conduction velocity (NCV) was (17.34 ± 2.15) m/s and (19.00 ± 3.02) m/s in group A, and (11.23 ± 1.45) m/s and (12.54 ± 1.59) m/s in group B, respectively, indicating significant differences (P lt; 0.05) between 2 groups. At 16 and 24 weeks after operation, HE staining and Bielschowsky staining showed that group A had a large number of nerve fiber regeneration, with a regular arrange of axons; while group B had l ittle nerve fiber regeneration with a scattered arrange of axons. At 24 weeks after operation, images in TEM showed a large number of regeneration myel inated nerve fibers and a small number of unmyel inated nerve fibers through the transplanted nerve in two groups. At 16 weeks after operation, the number of myel inated nerve fibers in group A and group B was (438 ± 79) and (196 ± 31) / vision, the areas of myel inated nerve fiberswere (5 596.00 ± 583.94) and (4 022.63 ± 615.75) μm2 / vision; after 24 weeks, the number of myel inated nerve fibers in groups A and B were (642 ± 64) and (321 ± 75)/vision, the areas of myel inated nerve fibers were (6 689.50 ± 1 142.10) and ( 4 733.00 ± 982.22) μm2/vision, indicating significant differences between two groups (P lt; 0.05). There was no statistically significant difference (P gt; 0.05) in the wet weight recovery rate of quadriceps between group A and group B at 16 weeks (87.96% ± 4.93% vs. 86.47% ± 7.47%) and at 24 weeks after operation (90.10% ± 4.22% vs. 87.66% ± 3.14%). Conclusion C7 transposition combined with autograft and allograft of sciatic nerve can reconstruct the partial function of the quadriceps femoris in paraplegia rats. The effect of graft is better than that of graft obviously.
Objective To observe and evaluate the expression and significance of Nogo66 receptor (NgR) mRNA in adult ratsprime;optic nerve. Methods Optic and sciatic nerves of 8 adult rats were used to make the sections, which were divided into 3 groups: optic-nerve experimental group, sciatic-nerve control group, and optic-nerve negative control group. In situ hybridization was used to observe the expression of NgR mRNA in optic nerve and sciatic nerve. Results The expression of NgR mRNA in the 8 rats was positive in optic nerve and negative in sciatic nerve. The positive signals were arranged along the long axis of optic nerve. Conclusion The expression of NgR mRNA is positive in optic nerve while negative in sciatic nerve in adult rats, which suggests that the positive expression and distribution of NgR may be related to the poor regenerate ability of optic nerves. (Chin J Ocul Fundus Dis, 2005,21:246-248)
OBJECTIVE To analysis the clinical characters of gluteal sciatic nerve injuries and investigate the treatment options. METHODS From October 1962 to June 1997, 190 patients with gluteal sciatic nerve injuries were adopted in this retrospective study. In these cases, the sciatic nerve injuries were caused by injection in 164 patients(86.32%), stab injury in 14 patients, pelvic fracture and hip dislocation in 11 patients, and contusion injury in 1 patient. Among them, 15 cases were treated by conservative method and the other 175 cases were operated. According to the observation during the operations, the injuries were occurred at the region of gluteal muscle in 146 cases, at the region of piriform muscle in 26 cases, and at the region of pelvic cavity in 3 cases. Then neurolysis was performed in 160 cases, epineurial neurorrhaphy in 12 cases and nerve grafting in 2 cases, and nerve exploration but no repair in 1 case. Late stage functional reconstruction of the foot and ankle was carried out in 23 cases. RESULTS One hundred and fifty-one patients were followed up 8.5 years in average. The occurrence of excellent and good nerve recovery was 56.95% and the occurrence of excellent and good functional reconstruction of late stage was 78.26%. CONCLUSION The gluteal sciatic nerve injury has since been challenging because of the tremendous difficulty in treatment and the poor outcome. The injury situation at the different region was closely related to the regional anatomy. According to this study, it is advised that the surgical treatment should be carried out actively. Neurolysis should be performed as soon as possible in the cases of injection injury. Epineurial neurorrhaphy should be performed in the cases of nerve rupture. In case of the gluteal sciatic nerve injury which caused by pelvic fracture or hip dislocation, the reduction and decompression is suggested in the early stage, and exploration and nerve repair is indicated in the late stage. The functional reconstruction of foot and ankle should be carried out in the late stage for the improvement of the limb function.
Abstract In case of sciatic nerve injury, there is degeneration of neuron in the corresponding segment of spinal cord. To study whether NGF could protect the dorsal root ganglia in this situation, the following experiments were performed: 72 SD mice were divided into 2 groups. In each mouse, the sciatic nerve was sectioned at the middle of the right thigh, and then,the proximal end of the sciatic nerve was inserted into a one ended silastic tube. The NGF 0.15ml (contain 2.5S NGF 0.15mg) was injected into the tubes of the experimental group, while a equal amount of normal saline was injected into the tubes of the control group. After 1, 3, 5, 9, 20 and 30 days, 6 mice of each groupwere sacrificed respectively, and 5th to 6th lumbar segments of the spinal cords were resected for examination. By histochemical study, the activity of fluoride resistant acid phosphatase (FRAP) of each animal was detected. The results showed: (1) Excision of the sciatic nerve led to decrease of FRAP activity, it suggested that the injury of sciatic nerve could damage the dorsal root ganglia; (2) The use of exogenous NGF could protect the FRAP activity. It was concluded that NGF played an important role in protecting the dorsal root ganglia in peripheral nerve injury, in vivo.
Objective To establ ish the methods to get high activity, high purity, and adequate Schwann cells (SCs), and to provide sufficient seed cells for the peripheral nerve repair. Methods Six 5-day-old, male or female, Sprague Dawley rats were selected and the sciatic nerve (control group) and dorsal root gangl ion (DRG) (ex perimental group) were harvested.Then the sciatic nerves and DRG were digested by co-enzyme and dispersed by medium containing serum to isolate SCs. Freshlyisolated SCs from rats were cultured, purified and subcultured. The 1st generation of SCs were chosen to draw the growth curve of SCs by the counting method and to detect the prol iferation of SCs by MTT assay at 8 days of culture, the purity of SCs by immunocytochemistry of anti-S-100 and the brain-derived neurotrophic factor (BDNF) concentration by ELISA. Results A total of 36-43 DRGs could be obtained in each rat. The number of obtained single SC in experimental group [(7.5 ± 0.6）× 106] was significantly higher than that in control group [(3.5 ± 0.4）× 106 ] (t=13.175, P=0.000). SCs reached logarithm prol iferation phase at 3 days. With time, the cell number and the prol iferation absorbance (A) value of 2 groups all showed upward trend. The number and A value of experimental group were significantly higher than those of control group (P lt; 0.05). The SCs purity of experimental group (92.08% ± 3.45%) was significantly higher than that of control group (77.50% ± 3.57%) (t=6.689, P=0.001).The concentrations of BDNF at 3 days and 5 days in experimental group were significantly higher than those of control group (P lt; 0.05). Conclusion The sufficient amount, high purity, and viabil ity of SCs from DRGs can meet the needs of studies on peripheral nerve repairment.
ObjectiveTo investigate the regularity of myelin degeneration and regeneration and the difference of axonal density between tibial nerve and common peroneal nerve after sciatic nerve injury repair in rhesue monkey. MethodsNine adult rhesue monkeys (male or female, weighing 3.5-4.5 kg) were selected to establish the model of rat sciatic nerve transaction injury. The tibial nerve and common peroneal nerve of 5 mm in length were harvested at 5 mm from injury site as controls in 3 monkeys; the distal tibial nerve and common peroneal nerve were repaired with 9-0 suture immediately in the other 6 monkeys. And the gross observation and neural electrophysiological examination were performed at 3 and 8 weeks after repair respectively. Then, distal tibial nerve and common peroneal nerve at anastomotic site were harvested to observe the myelin sheath changes, and to calculate the number of axon counts and axonal density by staining with Luxol Fast Blue. ResultsAtrophy of the lower limb muscle and various degrees of plantar ulcer were observed. Gross observation showed nerve enlargement at anastomosis site, the peripheral connective tissue hyperplasia, and obvious adhesion. The compound muscle action potential (CMAP) of tibial nerve and common peroneal nerve could not be detected at 3 weeks; the CMAP amplitude of common peroneal nerve was less than that of the tibial nerve at 8 weeks. Different degrees of axonal degeneration was shown in the tibial nerve and common peroneal nerve, especially in the common peroneal nerve. The average axonal density of common peroneal nerve was lower than that of tibial nerve at 3 weeks (13.2% vs. 44.5%) and at 8 weeks (10.3% vs. 35.3%) after repair. ConclusionThe regeneration of tibial nerve is better and faster than that of common peroneal nerve, and gastrocnemius muscle CMAP recovers quicker, and amplitude is higher, which is the reason of better recovery of tibial nerve.