Effect of folic acid coated-crosslinked urethane-doped polyester elastomer nerve conduit on promoting the repair of long distance peripheral nerve injury in rats / 中国修复重建外科杂志

OBJECTIVE@#To investigate the effect of folic acid coated-crosslinked urethane-doped polyester elastomer (fCUPE) nerve conduit in repairing long distance peripheral nerve injury.@*METHODS@#Thirty-six 3-month-old male Sprague Dawley rats weighing 180-220 g were randomly assigned to 3 groups, each consisting of 12 rats: CUPE nerve conduit transplantation group (group A), fCUPE nerve conduit transplantation group (group B), and autologous nerve transplantation group (group C), the contralateral healthy limb of group C served as the control group (group D). A 20-mm-long sciatic nerve defect model was established in rats, and corresponding materials were used to repair the nerve defect according to the group. The sciatic function index (SFI) of groups A-C was calculated using the Bain formula at 1, 2, and 3 months after operation. The nerve conduction velocity (NCV) of the affected side in groups A-D was assessed using neuroelectrophysiological techniques. At 3 months after operation, the regenerated nerve tissue was collected from groups A-C for S-100 immunohistochemical staining and Schwann cell count in groups A and B to compare the level of nerve repair and regeneration in each group.@*RESULTS@#At 3 months after operation, the nerve conduits in all groups partially degraded. There was no significant adhesion between the nerve and the conduit and the surrounding tissues, the conduit was well connected with the distal and proximal nerves, and the nerve-like tissues in the conduit could be observed when the nerve conduit stents were cut off. SFI in group A was significantly higher than that in group C at each time point after operation and was significantly higher than that in group B at 2 and 3 months after operation ( P<0.05). There was no significant difference in SFI between groups B and C at each time point after operation ( P>0.05). NCV in group A was significantly slower than that in the other 3 groups at each time point after operation ( P<0.05). The NCV of groups B and C were slower than that of group D, but the difference was significant only at 1 month after operation ( P<0.05). There was no significant difference between groups B and C at each time point after operation ( P>0.05). Immunohistochemical staining showed that the nerve tissue of group A had an abnormal cavo-like structure, light tissue staining, and many non-Schwann cells. In group B, a large quantity of normal neural structures was observed, the staining was deeper than that in group A, and the distribution of dedifferentiated Schwann cells was obvious. In group C, the nerve bundles were arranged neatly, and the tissue staining was the deepest. The number of Schwann cells in group B was (727.50±57.60) cells/mm 2, which was significantly more than that in group A [(298.33±153.12) cells/mm 2] ( t=6.139, P<0.001).@*CONCLUSION@#The fCUPE nerve conduit is effective in repairing long-distance sciatic nerve defects and is comparable to autologous nerve grafts. It has the potential to be used as a substitute material for peripheral nerve defect transplantation.

Nerve conduits of chitosan/polyvinyl alcohol with brain-derived neurotrophic factor microspheres for peripheral nerve defects in rats / 中国组织工程研究

BACKGROUND: The repair of peripheral nerve defects by nerve conduit bridging can provide a suitable microenvironment for nerve regeneration. On one hand, it can provide a unique channel for nerve regeneration, prevent the invasion of peripheral connective tissue and the formation of scars. On the other hand, it can maintain endogenous and exogenous neurotrophic factors, growth factors and other stimulants to promote axon growth. OBJECTIVE: To observe the therapeutic effect of chitosan/polyvinyl alcohol catheter injected with brain-derived neurotrophic factor sustained-release microspheres to bridge peripheral nerve defects. METHODS: Chitosan/polyvinyl alcohol nerve conduit was prepared by repeated freeze-thaw technique. The brain-derived neurotrophic factor microspheres were obtained by polymer-alloys combined with oil-oil emulsion/solvent evaporation method. A 15 mm sciatic nerve defect model was made in the right hindlimb of 60 adult male Sprague-Dawley rats. They were selected and randomly divided into four groups (n=15 per group): group A implanted with autogenous sciatic nerve; group B implanted with chitosan/polyvinyl alcohol nerve catheter, injected with normal saline; group C implanted with chitosan/ polyvinyl alcohol nerve catheter, injected with brain-derived neurotrophic factor solution; group D implanted with chitosan/polyvinyl alcohol nerve catheter, injected with brain-derived neurotrophic factor sustained-release microspheres. General observation, histological inspection, and electrophysiological determination were performed at 4 months after the surgery. This study was approved by the Research Ethics Committee of the Second Hospital of Hebei Medical University. RESULTS AND CONCLUSION: (1) Gross anatomy showed that muscle atrophy in group A and group D was lighter than that in the other two groups. The grafts in four groups were all adhered to the peripheral tissues, and the nerve in the autotransplantation segment was strongly adhered to the peripheral tissues. In group D, the regenerated nerve had connected the distal and proximal nerves, and the regenerated nerve filled the conduit. (2) Electrophysiological examination showed that the latency of group D was shorter than that of groups B and C (P 0.05). (3) Histological observation showed that there were regenerated nerve fibers in groups B, C, and D. The diameter, number and thickness of myelin sheath of group D were larger than those of group B and group C (P 0.05). (4) The results showed that the injection of brain-derived neurotrophic factor microspheres into chitosan/PVA catheter had a long-term promoting effect on peripheral nerve regeneration.

A green route for the fabrication of thermo-sensitive chitosan nerve conduits and their property evaluation / 中国修复重建外科杂志

Objective: To explore a green route for the fabrication of thermo-sensitive chitosan nerve conduits, improve the mechanical properties and decrease the degradation rate of the chitosan nerve conduits. Methods: Taking advantage of the ionic specific effect of the thermo-sensitive chitosan, the strengthened chitosan nerve conduits were obtained by immersing the gel-casted conduits in salt solution for ion-induced phase transition, and rinsing, lyophilization, and 60Co sterilization afterwards. The nerve conduits after immersing in NaCl solutions for 0, 4, 12, 24, 36, 48, and 72 hours were obtained and characterized the general observation, diameters and mechanical properties. According to the above results, the optimal sample was chosen and characterized the microstructure, degradation properties, and cytocompatibility. The left sciatic nerve defect 15 mm in length was made in 20 male Sprague Dawley rats. The autologous nerves (control group, n=10) and the nerve conduits (experimental group, n=10) were used to repair the defects. At 8 weeks after operation, the compound muscle action potential (CMAP) was measured. The regenerated nerves were investigated by gross observation and toluidine blue staining. The gastrocnemius muscle was observed by HE staining. Results: With the increased ionic phase transition time, the color of the conduit was gradually deepened and the diameter was gradually decreased, which showed no difference during 12 hours. The tensile strength of the nerve conduit was increased gradually. The ultimate tensile strength showed significant difference between the 48 hours and 12, 24, and 36 hours groups ( P0.05). As a result, the nerve conduit after ion-induced phase transition for 48 hours was chosen for further study. The scanning electron microscope (SEM) images showed that the nerve conduit had a uniform porous structure. The degradation rate of the the nerve conduit after ion-induced phase transition for 48 hours was significantly decreased as compared with that of the conduit without ion-induced phase transition. The nerve conduit could support the attachment and proliferation of rat Schwann cells on the inner surface. The animal experiments showed that at 8 weeks after operation, the CMAPs of the experimental and control groups were (3.5±0.9) and (4.3±1.1) m/V, respectively, which showed no significant difference between the two groups ( P0.05］. The nerve conduit of the experimental group could repair the nerve defect. There was no significant difference between the experimental and control groups in terms of the histomorphology of the regenerated nerve fibers and the gastrocnemius muscle. Conclusion: The green route for the fabrication of thermo-sensitive chitosan nerve conduits is free of any toxic reagents, and has simple steps, which is beneficial to the industrial transformation of the chitosan nerve conduit products. The prepared chitosan nerve conduit can be applied to rat peripheral nerve defect repair and nerve tissue engineering.

Compound nerve conduits in peripheral nerve repair / 中华整形外科杂志

Peripheral nerve injury is a relatively common sequela of trauma. Nerve regeneration and target re-innervation are complex processes, and remain clinical challenges. For a long period of time, the placement of an autologous nerve graft is the gold standard. However, autologous nerve grafts are limited in supply due to its potentially associated with donor site morbidities. Therefore, researchers keep seeking the ideal alternatives for neural repair. The clinical application of artificial nerve regeneration conduit has made incremental advancement. Specifically, compound conduits are gaining more and more attention, which are reported to have comparable effect to autologous nerve graft. The purpose of this paper is to review the progress of construction and application of compound nerve conduits in peripheral nerve repair.

Application of the bionic multi-channel nerve conduit in the rabbit sciatic nerve defect by reducing mismatch of regenerated nerve fibers / 中华显微外科杂志

To investigate the role of the bionic multi-channel nerve conduit by reducing mis-match of regenerated nerve fibers in the rabbit sciatic nerve defect. Methods The experiment was conducted from July, 2017 to February, 2019. A total of 55 New Zealand white rabbits were randomly divided into two groups (First group, n=30 and Second group, n=25).There were 5 subgroups (n=6) in the first group, which were autograft and cus-tom-anatomic nerve conduits (CANC) with different channel (1-CANC, 2-CANC, 3-CANC, 4-CANC) that implanted to repair the rabbit sciatic nerve defect (10 mm). The electrophysiological, triceps muscle wet weight recovery rate, histological study and ankle index analysis were used to evaluate the treatment of each group at 12 and 24 weeks postoperatively. There were 5 subgroups (n=5) in the second group. The simultaneous retrograde tracing method was applied to compare with the number of mismatched nerve fibers at 24 weeks postoperatively. All data were recorded and analyzed by One-way ANOVA method, the Turkey’s method was used to compare the differences between each subgroup. The difference was considered to be statistically significant if P＜0.05. Results The autograft group showed the best recovery in the electrophysiology, histology study and ankle index at 12 and 24 weeks postoperatively (P＜0.05).Histology results showed that the same number of myelinated nerve fibers in all CANC group (P＞0.05), but di-ameters of nerve fiber and myelin thickness were higher in 2-CANC and 3-CANC [(10.67±0.56) μm,(10.65±0.53) μm, respectively] compared with 1-CANC and 4-CANC groups [(8.43±0.63) μm, (9.03±0.55) μm, respectively].The differ-ences were similar in electrophysiological, wet weight recovery rate of triceps muscle, histological study and ankle index analysis.Simultaneous retrograde tracing showed that the autograft group had highest total number of labeled profiles, but no significant difference of the total number of labeled profile was showed among the CANC groups. However, the 1-CANC group[(7.1±2.4) %] showed highest percentage of the FB-NY-neurons than other CANC groups[(2.7±1.9)% in 2-CANC, (2.5±2.3) % in 3-CANC, and (2.2±1.2)% in 4-CANC](P＜0.05). Conclusion The autograft group showed the best results among all groups.Compared with the 1-CANC group, the 2-CANC and 3-CANC group obtained more mature regenerated nerve fibers and with a fewer mismatch rate.Moreover, that did not affect the number of regenerated fibers.

Construction strategy of nerve conduits bridging long peripheral nerve defects / 中国医师进修杂志

The repair, regeneration and functional recovery of peripheral nerve defects are the clinical problems needed to be solved urgently.Researches showed that the hollow conduits of bridging nerve defects only had the functions of support and isolation, but the composite conduits could simulate nerve regeneration microenvironment and possess biological activity, which could effectively guide the migration of Schwann cells and the regeneration of axons to achieve satisfactory effects of restoration and functional recovery.With the development of tissue engineering, regenerative medicine, biomaterial and the techniques of cells and molecular biology, the construction of biomimetic and functional nerve conduits to bridge long nerve defects has attracted more and more attention. This review will provide a brief overview of the construction strategy of nerve conduits repairing long nerve defects of peripheral nerves.

The effect of new PRGD/PDLLA/VPA composite nerve conduits on peripheral nerve regeneration in a rat model / 中华显微外科杂志

Objective To observe enhancing effect of nerve regeneration on peripheral nerve defect models bridged by a new PRGD/PDLLA/VPA composite conduit.Methods In this study from February,2012 to March,2014,PRGD/PDLLA/VPA nerve conduits were tested in the rat sciatic nerve transection model.At different periods after operation,its ability to promote nerve regeneration was evaluated by sciatic functional index(SFI),electrophysiology (CMAPs,NCVs) and histologic assessment.Forty rats were randomly divided into 4 groups (n =10),group A:PRGD/PDLLA/VPA,group B:PDLLA/VPA,group C:PRGD/VPA and group D:autograft.Results At 12 weeks after surgery,the SFI value of group A (-45 ± 3.19)and group D (-42 ± 3.01)were significantly higher than those of group B(-79 ± 3.06) and group C(-72 ± 2.07)(P ＜ 0.05);The CMAPs of group A (24.89 ± 5.01) and group D (25.39 ± 5.63) were significantly higher than those of group B(14.88 ± 3.11) and C(15.00 ± 5.54);the NCVs of group A (31.42 ± 2.43) and group D (31.50 ± 2.16) were significantly higher than those of group B (20.11 ± 2.39) and group C(21.00 ± 2.13)(P ＜ 0.05).At 12 weeks after surgery,the numbers of regenerated nerve in the tube of group A (258 ± 6.18) and D(259 ± 5.59) were significantly higher than those of group B (231 ± 5.00) and group C(230 ± 5.07)(P ＜ 0.05).There was no significant difference between groups A and D(P ＞ 0.05).Conclusion These results illustrated that this new PRGD/PDLLA/VPA conduit could significantly facilitate the regeneration of short nerve defect and recovery of motor nerve,which provides a new thought for treatment of peripheral nerve injury.

PRGD composite nerve conduit in repair of human peripheral nerve defect in upper extremity / 中华显微外科杂志

Objective To explore the clinical efficacy of PRGD composite nerve conduit in the treatment of human large-diameter,critical peripheral nerve defect in upper extremity.Methods From December,2011 to August,2014,19 patients with large-diameter,critical peripheral nerve defect in upper extremity were treated with PRGD composite nerve conduit.The patients were followeded-up periodically.The sensory and motor function recovery,high frequency ultrasound,and EMG were employed to assess the efficacy.Results The patients were followed up for an average time of 12-32 months(mean 21.75 ± 6.86 months),sensory and motor function recovered excellent in 7 patients,satisfactory in 7 patients,tolerable in 3 patients and no improvement in 2 patients were obtained according to the peripheral nerve function assessment standard built by British medical research council,the rate excellent and satisfactory results was 73.7％.Conclusion It is clinically promising to use PRGD composite nerve conduit to repair large-diameter,critical peripheral nerve defect in upper extremity,thus laying a foundation for its further application in clinical practice.

Comparison of Processed Nerve Allograft and Laminin Derived Peptide Incorporated Nerve Conduit for Peripheral Nerve Regeneration / 대한정형외과학회잡지

PURPOSE: To compare a processed nerve allograft, laminin derived peptide incorporated nerve conduit, and autograft in terms of electrodiagnostic testing and nerve histomorphometry for peripheral nerve regeneration in a rabbit sciatic nerve defect model. MATERIALS AND METHODS: Thirty New Zealand white rabbits were divided into three groups, and a unilateral 15 mm sciatic nerve defect was made. Group I, II and III was repaired with a reversed autograft, a processed acellular nerve allograft, and a laminin derived peptide incorporated nerve conduit, respectively. At twelve weeks, the animals were evaluated with the compound muscle action potential, wet muscle weight, and nerve histomorphometric parameters such as nerve area, number of axons, and myelin thickness. RESULTS: At twelve weeks, the compound muscle action potential for group I, II and III was 54.1%, 38.2% and 26.4%, respectively. Significant differences were found between the three groups (p<0.001, group I vs II; p<0.001, group I vs III; p<0.001, group II vs III). The wet muscle weight for group I, II and III was 57.8%, 54.4% and 43.9%, respectively. Group I had significantly more muscle weight than group III (p<0.001), but the difference was not significant with group II (p=0.256). Group II and III showed a significant difference (p=0.002). The number of axons in group III decreased and the shape of the axon was irregular, even though the nerve area and myelin thickness were similar in the three groups. CONCLUSION: An autograft remains the gold standard to repair a segmental nerve defect. Processed allograft demonstrated superior nerve recovery compared to the laminin derived peptide incorporated nerve conduit.

Graft repair of peripheral nerve defects using small intestinal submucosa with crude Schwann cells / 上海第二医科大学学报

Objective To investigate the results of graft repair of peripheral nerve defects using small intestinal submucosa(SIS) with crude Schwann cells. Methods Thirty-six male SD rats were randomly divided into three groups,with 12 in each group.Firstly,12-mm gaps of sciatic nerve were made in rats and then treated respectively with porcine SIS(group A),SIS compounded with crude Schwann cells(group B) and auto-nerve grafting(group C) in the three groups.The grafts were harvested 16 weeks postoperatively to evaluate the results of nerve regeneration through histological examination, triceps surae weight,computerized imaging analysis,Trueblue retrograde tracing and transmission electron microscopy. Results Nerve regeneration was confirmed to have extended through the gaps according to the results of histological examination,Trueblue retrograde tracing and transmission electron microscopy.Furthermore,in the respect of triceps surae weight,quantity of axis cylinder per area and percentage of neural tissue,the results of group B and group C were superior to those of group A with a significantly statistical difference(P0.05). Conclusion SIS compounded crude Schwann cells was able to achieve the outcome of auto-nerve grafting and was a promising replacement graft.

Experimental study of peripheral nerve regeneration by using non-tubular natural cellulose membrane nerve conduit

Styela clava, called non-native tunicate or sea squirt, is habitat which include bays and harbors in Korea and several sites in the sea faced world. We fabricate cellulose membrane nerve conduit (CMNC) from this native sea squirt skin, and evaluate the capacity of promoting peripheral nerve regeneration in the rat sciatic nerve defect model. After processing the pure cellulose membrane from the sea squirt skin as we already published before, CMNC was designed as a non-tubular sheet with 14 mm length and 4 mm width. Total eleven male Spraque-Dawley rats (12 weeks, weighing 250 to 300g) were divided into sham group (n=2), silicone tube grafted control group (n=3) and experimental group (n=6). Each CMNC grafted nerve was evaluated after 4, 8 and 12 weeks in the experimental group, and after 12 weeks, sciatic function was evaluated with sciatic function index (SFI) and gait analysis, and histomorphology of nerve conduit and the innervated tissues of sciatic nerve were all examined using image analyzer and electromicroscopic methods in the all groups. The regenerated axon and nerve sheath were found only in the inner surface of the CMNC after 4 weeks and became more thicker after 8 and 12 weeks. In the TEM study, CMNC grafted group showed more abundant organized myelinated nerve fibers with thickened extracellular matrix than silicone conduit grafted group after 12 weeks. The sciatic function index (SFI) and ankle stance angle (ASA) in the functional evaluation were -47.2+/-3.9, 35.5.+/-4.9.in CMNC grafted group (n=2) and -80.4+/-7.4, 29.2.+/-5.3.in silicone conduit grafted group (n=3), respectively. And the myelinated axon was 41.59% in CMNC group and 9.51% in silicone conduit group to the sham group. The development of a bioactive CMNC to replace autogenous nerve grafts offers a potential and available approach to improved peripheral nerve regeneration. As we already published before, small peptide fragment derived from the basement membrane matrix proteins of squirt skin, which is a kind of anchoring protein composed of glycocalyx, induced the effective axonal regeneration with rapid growth of Schwann cells beneath the inner surface of CMNC. So the possibilities of clinical application as a peripheral nerve regeneration will be able to be suggested.

Morphological study on the angiogenesis of peripheral nerve affected by nerve growth factor / 中国矫形外科杂志

[Objective]To discuss the effects of angiogenesis about nerve growth factor(NGF) during the peripheral nerve regeneration.[Method]Thirty-six Sprague-Dawley rats with 10mm gap of sciatic nerve were randomly divided into two groups which had been bridged with the new double channel nerve conduit of fusiform shape.Each group contained eighteen animals,in the first group,200 ?l of chitin for medical use was injected into the conduit,in the second group,the two branches of the conduit contained 100 ?l of the chitin and 5 ?l NGF or ciliary neurotrophic factor(CNTF).At four,eight or sixteen week after operation,the angiogenesis of NGF was evaluated with Hematoxylin and Eosin(HE) staining and electron microscope.[Result]There were not significant differences of the regenerative nerve fibres between two channels in the first groups,but in the second group,the regenerative nerve of NGF branch channel was red,crisp and the nerve of CNTF branch channel was yellow and tenacious.HE staining showed that there were much more new vessel in the regernerative nerve tract of NGF the branch channel,and the regernerative nerve fibre was disorder,there were much more fibroblasts and vessels observed under eletron microscope.[Conclusion]NGF can significantly promote the angiogenesis during the peripheral nerve regeneration,the mechanism may be related to fibroblast.

Ciliary neurotrophic factor-coated polyglecolic-polylactic acid nerve conduits to repair canine tibial nerve defects / 中华显微外科杂志

Objective To explore the effect of ciliary,neurotrophic factor (CNTF)-coated polyglycolic and polylactic acid (PGLA) nerve conduits treated by pulsed plasma to repair canine tibial nerve defects. Methods A 2.5 cm long tibial nerve defect was made in eighteen cross-bred dogs.The nerve defects were re- constructed by three different methods:group A:pulsed plasma treated and CNTF coated PGLA nerve conduits (n=6);group B:PGLA nerve conduits alone (n=6);group C:nerve autografts (n=6).HE staining, Massons' trichrome staining,S-100 immunostaining,electrophysiological test and axon counting were used to evaluate the results of nerve regeneration in three groups.In addition,the dynamic walking pattern was recor- ded individually.The observation period lasted for three months.Results All nerve conduits were well vas- cularized and mostly degraded as well as absorbed.It was found that the regenerating axons could traverse all nerve conduits.In regard to nerve conduction velocity and axon counting there was no significant difference be- tween group A and group C (P＞0.05),while the data of group A and group C were significantly better than those of group B(P＜0.05).The dogs in group A and C recovered nearly normal walk pattern while those in group B were still crippled.Conclusion Pulsed plasm-treated and then CNTF-coated PGLA nerve conduits could effectively repair 2.5-cm-long canine tibial nerve defects,and the effect is similar to that of autografts.

Sciatic nerve regeneration using calcium phosphate coated conduit and brain-derived neurotrophic factor gene-transfected schwann cell in rat

PURPOSE OF STUDY: Peripheral nerve regeneration depends on neurotrophism of distal nerve stump, recovery potential of neuron, supporting cell like Schwann cell and neurotrophic factors such as BDNF. Peripheral nerve regeneration can be enhanced by the conduit which connects the both sides of transected nerve. The conduit maintains the effects of neurotrophism and BDNF produced by Schwann cells which can be made by gene therapy. In this study, we tried to enhance the peripheral nerve regeneration by using calcium phosphate coated porous conduit and BDNF-Adenovirus infected Schwann cells in sciatic nerve of rats. MATERIALS AND METHODS: Microporous filter which permits the tissue fluid essential for nerve regeneration and does not permit infiltration of fibroblasts, was made into 2mm diameter and 17mm length conduit. Then it was coated with calcium phosphate to improve the Schwann cell adhesion and survival. The coated filter was evaluated by SEM examination and MTT assay. For effective allogenic Schwann cell culture, dorsal root ganglia of 1-day old rat were extracted and treated with enzyme and antimitotic Ara-C. Human BDNF cDNA was obtained from cDNA library and amplified using PCR. BDNF gene was inserted into adenovirus shuttle vector pAACCMVpARS in which E1 was deleted. We infected the BDNF-Ad into 293 human mammary kidney cell-line and obtained the virus plaque 2 days later. RT-PCR was performed to evaluate the secretion of BDNF in infected Schwann cells. To determine the most optimal m.o.i of BDNF-Ad, we infected the Schwann cells with LacZ adenovirus in 1, 20, 50, 75, 100, 250 m.o.i for 2 hours and stained with beta-galactosidase. Rats(n=24) weighing around 300g were used. Total 14mm sciatic nerve defect was made and connected with calcium phosphate coated conduits. Schwann cells(1x10(6)) or BDNF-Ad infected Schwann cells(1x10(6)) were injected in conduit and only media(MEM) was injected in control group. Twelve weeks after surgery, degree of nerve regeneration was evaluated with gait analysis, electrophysiologic measurements and histomorphometric analysis. RESULTS: 1. Microporous Millipore filter was effective conduit which permitted the adhesion of Schwann cells and inhibited the adhesion of fibroblast. We could enhance the Schwann cell adhesion and survival by coating Millipore filter with calcium phosphate. 2. Schwann cell culture technique using repeated treatment of Ara-C and GDNF was established. The mean number of Schwann cells obtained 1 and 2 weeks after the culture were 1.54+/-4.0*10(6) and 9.66+/-9.6*10(6). 3. The mRNA of BDNF in BDNF-Ad infected Schwann cells was detected using RT-PCR. In Schwann cell 0.69 microgram/microliter of DNA was detected and in BDNF-Adenovirus transfected Schwann cell 0.795 microgram/microliter of DNA was detected. The most effective infection concentration was determined by LacZ Adenovirus and 75 m.o.i was found the most optimal. CONCLUSION: BDNF-Ad transfected Schwann cells successfully regenerated the 14mm nerve gap which was connected with calcium phosphate coated Millipore filter. The BDNF-Ad group showed better results compared with Schwann cells only group and control group in aspect to sciatic function index, electrophysiologic measurements and histomorphometric analysis.

Peripheral nerve regeneration using a three-dimensionally cultured schwann cell conduit

The use of artificial nerve conduit containing viable Schwann cells is one of the most promising strategies to repair the peripheral nerve injury. To fabricate an effective nerve conduit whose microstructure and internal environment are more favorable in the nerve regeneration than existing ones, a new three-dimensional Schwann cell culture technique using Matrigel(R) and dorsal root ganglion (DRG) was developed. Nerve conduit of three-dimensionally arranged Schwann cells was fabricated using direct seeding of freshly harvested DRG into a Matrigel(R) filled silicone tube (I.D. 1.98 mm, 14 mm length) and in vitro rafting culture for 2 weeks. The nerve regeneration efficacy of three-dimensionally cultured Schwann cell conduit (3D conduit group, n=6) was assessed using SD rat sciatic nerve defect of 10 mm, and compared with that of silicone conduit filled with Matrigel(R) and Schwann cells prepared from the conventional plain culture method (2D conduit group, n=6). After 12 weeks, sciatic function was evaluated with sciatic function index (SFI) and gait analysis, and histomorphology of nerve conduit and the innervated tissues of sciatic nerve were examined using image analyzer and electromicroscopic methods. The SFI and ankle stance angle (ASA) in the functional evaluation were -60.1+/-13.9, 37.9 degrees +/-5.4 degrees in 3D conduit group (n=5) and -87.0 +/-12.9, 32.2 degrees +/-4.8 degrees in 2D conduit group (n=4), respectively. And the myelinated axon was 44.91%+/-0.13% in 3D conduit group and 13.05%+/-1.95% in 2D conduit group to the sham group. In the TEM study, 3D conduit group showed more abundant myelinated nerve fibers with well organized and thickened extracellular collagen than 2D conduit group, and gastrocnemius muscle and biceps femoris tendon in 3D conduit group were less atrophied and showed decreased fibrosis with less fatty infiltration than 2D conduit group. In conclusion, new three-dimensional Schwann cell culture technique was established, and nerve conduit fabricated using this technique showed much improved nerve regeneration capacity than the silicone tube filled with Matrigel(R) and Schwann cells prepared from the conventional plain culture method.

Effects of Fresh and Degenerated Autogenous Nerve Graft in Segmental Defect of Sciatic Nerve of Rabbit / 대한정형외과학회잡지

The potential for nerve regeneration and recovery of its function exists in the presence of a suitable pathway for regenerating axon and endoneurial tube can serve as nerve conduit for regenerating axon in fresh nerve graft. But value of degenerated nerve as donor nerve has not been established till now. This experiment assessed the chronologic influence of fresh and degenerated nerve graft on axonal growth for 10mm gap of sciatic nerve in rabbit and how long degenerated nerve was ahle to serve as a nerve conduit microscopically. Electromicroscopically, the regenerating axons which were ohserved in the degenerated nerve graft had more abundant unmyelinated fibers and revealed abundant collagen fibers in the endoneurium. And these regenerating axons became gradually surrounded with newly developed basal lamina and decreased the collagen fibers at l2 week of degenerated nerve graft. Histologically, myelinated axons which were observed in central area of the degenerated nerve graft at 8 week of degenerated nerve graft were relatively thinner, but a definite structural difference of regenerating axons was not found except reduction of number compared with those of tresh nerve graft. The numher of myelinated axons was 6,072+/-l42 in normal sciatic nerve, 4,479+/- 157 in fresh nerve graft group, and 2,968+/-168 in the degenerated nerve graft group. Difference of the number of myelinated axons between fresh and degenerated nerve graft group was significant stati stically(P<0.05). These results showed that the ability of a degenerated nerve graft as a passage for the regenerating axons and it can be employed as one of the favorable nerve conduits.