Objective To explore an effect of the artificial nerve graft wrapped in the pedicled greater omentum on the early revascularization and an effectof the increased blood supply to the artificial nerve graft on the nerve regeneration. Methods Seventy-five rabbits were randomized into 3 groups, in which there were 2 experimental groups where the rabbits were made to abridge respectively with the artificial nerve grafts wrapped in the pedicled greater omentum (Group A) and with the artificial nerve grafts only (Group B), and the control group where the rabbits were abridged with the autologous nerve (Group C).On the 3rd, 7th and 14th days after operation, the evans blue bound to albumin (EBA) was injected into the vessels in all the grafts to show their revascularization. Twelve weeks after operation the nerve regeneration was evaluated with theelectrophysiological and histological observations on the serial sections, and was evaluated also with the transmission electron microscope. Results The artificial nerve grafts wrapped in the pedicled greater omentum in Group A and the autologous nerve grafts in Group C showed a beginning of revascularization on the3rd day after operation, and the revascularization was increased on the 7th and14th days. Compared with Groups A and C, the artificial nerve grafts in Group Bshowed a delayed revascularization on the7th day after operation. At 12 weeks after operation, there were no significant differences in the motor never conduction velocity, density of the regenerated myelinated nerve fibers, myelin sheath thickness, and diameter between Group A and Group C(Pgt;0.05). However, both Group A and Group C were superior to Group B in the above variables, with significant differences(Plt;0.05). Conclusion Utilization of the pedicled greater omentum to wrapthe artificialnerve grafts can promote an early revascularization of the artificial nerve graft and an early nerve regeneration of the artificial nerve graft because of an enhanced blood supply to the nerve graft.
OBJECTIVE To observe the degeneration and regeneration of the Meissner’s corpuscles after implanted sensory nerve into the denervated monkey’s fingers under electron microscope. METHODS The two finger nerves of the monkey’s fingers were denervated. Afterwards, one finger nerve was cut off, and the other was reimplanted into the denervated finger. After 1, 3, 5, 8 and 12 months, the finger skin was cut off and observed under electron microscope. RESULTS The degenerative changes of nerve ending in Meissner’s corpuscles were observed after 1 month of denervation, and the basic structure of the corpuscles had no obvious changes. After 3 months, the axons of corpuscles were disappeared, and the volume of corpuscles was shrunk. The basic structure of nerves was disappeared, and the lemmocyte and neurolemma plate were changed after 5 months. The collagen fibrils in the corpuscles were gradually increased in 8 months, the endoneurial structure and interneurial matrix were completely disappeared and replaced by collagen fibrils in 12 months. After 3 months of nerve implantation, unmyelinated nerve fibers were appeared and grew into the corpuscles. A part of corpuscles innervated in 5 months. Most of corpuscles innervated and myelinated nerve fibers were observed in 8 months. And in 12 months, corpuscles innervated to normal level. CONCLUSION The implantative sensory nerve by means of reinnervating the original corpuscles and regenerating new corpuscles could innervate the degenerative Meissner’s corpuscles.
OBJECTIVE Following the delayed repair of peripheral nerve injury, the cell number of anterior horn of the spinal cord and its ultrastructural changes, motorneuron and its electrophysiological changes were investigated. METHODS In 16 rabbits the common peroneal nerves of both sides being transected one year later were divided into four groups randomly: the degeneration group and regeneration of 1, 3 and 5 months groups. Another 4 rabbits were used for control. All transected common peroneal nerves underwent epineural suture except for the degeneration group the electrophysiological examination was carried out at 1, 3 and 5 months postoperatively. Retrograde labelling of the anterior horn cells was demonstrated and the cells were observed under light and electronmicroscope. RESULTS 1. The number of labelled anterior horn cell in the spinal cord was 45% of the normal population after denervation for one year (P lt; 0.01). The number of labelled cells increased steadily from 48% to 57% and 68% of normal values at 1, 3 and 5 months following delayed nerve repair (P lt; 0.01). 2. The ultrastructure of the anterior horn cells of the recover gradually after repair. 3. With the progress of regeneration the latency become shortened, the conduction velocity was increased, the amplitude of action potential was increased. CONCLUSION Following delayed repair of injury of peripheral nerve, the morphology of anterior horn cells of spinal cord and electrophysiological display all revealed evidence of regeneration, thus the late repair of injury of peripheral nerve was valid.
Objective To investigate the effect of tetramethylpyrazine (TMP) with a certain concentration added to vitrification solution on peripheral nerve allografts regeneration. Methods Forty-eight healthy clean SD male rats were selected as donors, and 96 healthy clean Wistar male rats as recipients, all rats being 3 months old and weighing 200-250 g. The sciatic nerves segments of 15 mm were removed from the donors, then randomly divided into 4 groups according to vitrificationsolution containing TMP. No TMP was used in group A as the control group; 100 mg/L, 200 mg/L and 400 mg/L TMP were used in group B, group C and group D, respectively. Then them were cryo-preserved at — 196 ℃ for 3 weeks. Nerve defect of 10 mm in length was made in the sciatic nerves of recipients. After rewarming, the allografts were transplanted to the corresponding rats. The gross appearance, the morphological and electrophysiological changes, the image analysis of axons and motor end-plate were detected at 4, 8, 12 and 16 weeks. Results All rates survived to the end of the experiment. The adhesion and edema of allografts in group A and group B were obvious 4 weeks after operation; then adhesion and edema was obvious in group A and were improved in the other groups 8 weeks after operation. Adhesion was observed in groups A and B; no adhesion was observed in groups C and D at 12 weeks. The number of regeneration nerve, the latent, the ampl itude, the nerve conduction velocity, the medullary sheath/μm2, the medullary sheath density/μm2 and the image analysis of axons and motor end-plate in groups A and B were significantly lower than those in groups C and D (P lt; 0.01); and there were no significant differences between groups C and D (P gt; 0.05). The observation of transmission electron microscope showed that medullated nerve fibers and myel in sheath of groups C and D were thicker than groups A and B, layers of groups C and D were clear. Conclusion The vitrification solution with 200 mg/L tetramethylpyrazine has protective effect on regeneration of peripheral nerve allografts．
Objective To investigate the feasibility of selenium-methylselenocysteine (SMC) to promote peripheral nerve regeneration and its mechanism of action. Methods Rat Schwann cells RSC96 cells were randomly divided into 5 groups, which were group A (without any treatment, control group), group B (adding 100 μmol/L H2O2), group C (adding 100 μmol/L H2O2+100 μmol/L SMC), group D (adding 100 μmol/L H2O2+200 μmol/L SMC), group E (adding 100 μmol/L H2O2+400 μmol/L SMC); the effect of SMC on cell proliferation was detected by MTT method, and the level of oxidative stress was detected by immunofluorescence for free radicals [reactive oxygen species (ROS)] after determining the appropriate dose group. Thirty-six 4-week-old male Sprague Dawley rats were randomly divided into 3 groups, namely, the sham operation group (Sham group), the sciatic nerve injury group (PNI group), and the SMC treatment group (SMC group), with 12 rats in each group; the rats in the PNI group were fed with food and water normally after modelling operation, and the rats in the SMC group were added 0.75 mg/kg SMC to the drinking water every day. At 4 weeks after operation, the sciatic nerves of rats in each group were sampled for neuroelectrophysiological detection of highest potential of compound muscle action potential (CMAP). The levels of inflammatory factors [interleukin 17 (IL-17), IL-6, IL-10 and oxidative stress factors catalase (CAT), superoxide dismutase (SOD), and malondialdehyde (MDA)] were detected by ELISA assay. The luxol fast blue (LFB) staining was used to observe the myelin density, fluorescence intensity of glial fibrillary acidic protein (GFAP) and myelin basic protein (MBP) was observed by immunofluorescence staining, and myelin morphology was observed by transmission electron microscopy with measurement of axon diameter. Western blot was used to detect the protein expressions of p38 mitogen-activated protein kinases (p38MAPK), phosphorylated p38MAPK (p-p38MAPK), heme oxygenase 1 (HO-1), and nuclear factor erythroid 2-related factor 2 (Nrf2). ResultsMTT assay showed that the addition of SMC significantly promoted the proliferation of RSC96 cells, and the low concentration could achieve an effective effect, so the treatment method of group C was selected for the subsequent experiments; ROS immunofluorescence test showed that group B showed a significant increase in the intensity of ROS fluorescence compared with that of group A, and group C showed a significant decrease in the intensity of ROS fluorescence compared with that of group B (P<0.05). Neuroelectrophysiological tests showed that the highest potential of CMAP in SMC group was significantly higher than that in PNI and Sham groups (P<0.05). ELISA assay showed that the levels of IL-6, IL-17, and MDA in PNI group were significantly higher than those in Sham group, and the levels of IL-10, SOD, and CAT were significantly lower; the levels of IL-6, IL-17, and MDA in SMC group were significantly lower than those in PNI group, and the levels of IL-10, SOD, and CAT were significantly higher (P<0.05). LFB staining and transmission electron microscopy showed that the myelin density and the diameter of axons in the SMC group were significantly higher than those of the PNI group and the Sham group (P<0.05). Immunofluorescence staining showed that the fluorescence intensity of GFAP and MBP in the SMC group were significantly stronger than those in the PNI group and Sham group (P<0.05). Western blot showed that the relative expressions of Nrf2 and HO-1 proteins in the SMC group were significantly higher than those in the PNI group and Sham group, and the ratio of p-p38MAPK/p38MAPK proteins was significantly higher in the PNI group than that in the SMC group and Sham group (P<0.05). Conclusion SMC may inhibit oxidative stress and inflammation after nerve injury by up-regulating the Nrf2/HO-1 pathway, and then inhibit the phosphorylation of p38MAPK pathway to promote the proliferation of Schwann cells, which ultimately promotes the formation of myelin sheaths and accelerates the regeneration of peripheral nerves.
Objective To explore the changes of calcitonin gene-related peptide (CGRP) and substance P (SP) levels after end-toend and end-to-side neurorrhaphy. Methods Twenty female Wistar rats were divided into 4 experimental groups and control group. In the experimental groups, common peroneal nerves were transected on both sides. End-to-side coaptation was performed on the left, while end-to-end coaptation on the right. After 1, 2, 4 and 27 weeks, the rats were sacrificed, and immunoreactivities of CGRP and SP in suture sites, lumbar spine and dorsal root ganglia(DRGs) were evaluated respectively. Results The expression ofCGRP and SP decreased in dorsal horn and DRGs within 1 week postoperatively. After 4 -27 weeks, CGRP and SP in dorsal horn could return to almost normal level, but they had little recovery in DRGs. Although the trend of change between end-to-end and end-to-side was coincident, in most experimental groups, thereexisted differences in the dorsal horn between end-to-end and end-to-side. The sciatic nerve stained by acetylcholinesterase, SP, CGRP and PGP 9.5 showed that the fibers could pass through the suture site of either end-to-end or end-to-side. Conclusion Nerve regeneration can be achieved by end-to-side neurorrhaphy, andthe mechanism of sensory nerve recovery of these two methods is similar. But the recovery in end-to-side coaptation is insufficient to some degree.
OBJECTIVE: To explore the possibility to bridge peripheral nerve defects by xenogeneic acellular nerve basal lamina scaffolds. METHODS: Thirty SD rats were randomly divided into 5 groups; in each group, the left sciatic nerves were bridged respectively by predegenerated or fresh xenogeneic acellular nerve basal lamina scaffolds, autogenous nerve grafting, fresh xenogeneic nerve grafting or without bridging. Two kinds of acellular nerve basal lamina scaffolds, extracted by 3% Triton X-100 and 4% deoxycholate sodium from either fresh rabbit tibial nerves or predegenerated ones for 2 weeks, were transplanted to bridge 15 mm rat sciatic nerve gaps. Six months after the grafting, the recovery of function was evaluated by gait analysis, pinch test, morphological and morphometric analysis. RESULTS: The sciatic nerve function indexes (SFI) were -30.7% +/- 6.8% in rats treated with xenogeneic acellular nerve, -36.2% +/- 9.7% with xenogeneic predegenerated acellular nerve, and -33.9% +/- 11.3% with autograft respectively (P gt; 0.05). The number of regenerative myelinated axons, diameter of myelinated fibers and thickness of myelin sheath in acellular xenograft were satisfactory when compared with that in autograft. Regenerated microfascicles distributed in the center of degenerated and acellular nerve group. The regenerated nerve fibers had normal morphological and structural characters under transmission electron microscope. The number and diameter of myelinated fibers in degenerated accellular nerve group was similar to that of autograft group (P gt; 0.05). Whereas the thickness of myelin sheath in degenerated accellular nerve group was significantly less than that of autograft group (P lt; 0.05). CONCLUSION: The above results indicate that xenogeneic acellular nerve basal lamina scaffolds extracted by chemical procedure can be successfully used to repair nerve defects without any immunosuppressants.
ObjectiveTo investigate the expression pattern and significance of Sonic Hedgehog (Shh) signaling pathway by observing whether the Shh signaling pathway components express in the adult rat after spinal cord injury (SCI). MethodsSixty-four healthy male Sprague-Dawley rats were randomly divided into normal group (group A, 8 rats), sham group (group B, 8 rats), and SCI group (group C, 48 rats). In group A, the rats served as controls without any treatment; a decompressive laminectomy was performed on T7-9 levels without SCI in group B; and modified Allen's method was used to make SCI model in group C. Basso Beattie Bresnahan (BBB) scale was used to assess the hind limb motor function at 12 hours, 1 day, 3 days, 7 days, 14 days, and 21 days after SCI; the immunofluorescence staining, real-time PCR, and Western blot were performed to detect the mRNA and protein expression levels of Shh and Glioma-associated oncogene homolog-1 (Gli-1) in SCI zone. ResultsThe BBB score slowly increased with time in group C, but the scores at each time point in group C were significantly lower than those in group A and group B (P<0.05). The results of immunofluorescence staining showed that Shh and Gli-1 rapidly increased after SCI in astrocytes. Real-time PCR and Western blot showed that the relative expression levels of Shh and Gli-1 mRNA and protein were gradually increased in group C and reached a maximum at 7 days. In addition, the relative expression levels of Shh and Gli-1 mRNA and protein in group C were significantly higher than those in group A and group B (P<0.05). On the other hand, compared with group A, the expression of Gli-1 protein was reduced in the cytoplasm but increased in nucleus in group C. ConclusionAstrocytes synthesize and secrete Shh and Gli-1 signaling molecules after SCI, both Shh and Gli-1 significantly up-regulate and exhibit dynamic changes, which suggests Shh signaling pathway may be involved in nerve cell regeneration after SCI.
OBJECTIVE: To evaluate the nerve regeneration after implantation of chitin tubes containing nerve growth factor(NGF) in the rabbit facial nerve. METHODS: Bilateral 8 mm defect of superior buccal divisions of the facial nerves were made in 16 New Zealand rabbits. Chitin tubes containing NGF were implanted into the gaps, and autologous nerves were implanted into the right gaps as control. The nerve regeneration was evaluated with electrophysiological and ultrastructural examination after 8 and 16 weeks of operation. RESULTS: Chitin tubes containing NGF successfully induced the nerve regeneration, regularly arranged myelinated and unmyelinated axons could be observed across the 8 mm gaps, and the myelin sheath was thick with clear lamellar structure at 8 weeks after operation, The regenerated nerve fibers increased and were more mature at 16 weeks after operation. There were no significant difference in electrical impulse conduction velocity through the neural regeneration between the experimental and control sides (P gt; 0.05). CONCLUSION: Chitin tubes containing NGF can provide optimal conditions for regeneration of rabbit facial nerve.
A comparative study of four methods of laryngeal muscle reinnervation in dogs is presented. Twenty-eight cases were divided into four groups to undergo main branch and branch of ansa cervicalis nerve anastomosis, and nerves implantation an neuromuscular pedicles transfer respectively for restoration of vocal cord adduction on left sides. The results showed that the four procedures seemed to induce effective reinnervation of adductor muscles. But the main branch of ansa cervicalis nerve suture was superior to the other methods among which little difference was noted in the functional recovery, electrophysiological activity and muscle strength. It demonstrated that main branch of ansa nerve suture was the best procedure for treatment of unilateral vocal cord paralysis among the four methods.