Allotransplanted neurons used to repair peripheral nerve injury do not elicit overt immunogenicity

A major problem hindering the development of autograft alternatives for repairing peripheral nerve injuries is immunogenicity. We have previously shown successful regeneration in transected rat sciatic nerves using conduits filled with allogeneic dorsal root ganglion (DRG) cells without any immunosuppression. In this study, we re-examined the immunogenicity of our DRG neuron implanted conduits as a potential strategy to overcome transplant rejection. A biodegradable NeuraGen® tube was infused with pure DRG neurons or Schwann cells cultured from a rat strain differing from the host rats and used to repair 8 mm gaps in the sciatic nerve. We observed enhanced regeneration with allogeneic cells compared to empty conduits 16 weeks post-surgery, but morphological analyses suggest recovery comparable to the healthy nerves was not achieved. The degree of regeneration was indistinguishable between DRG and Schwann cell allografts although immunogenicity assessments revealed substantially increased presence of Interferon gamma (IFN-γ) in Schwann cell allografts compared to the DRG allografts by two weeks post-surgery. Macrophage infiltration of the regenerated nerve graft in the DRG group 16 weeks post-surgery was below the level of the empty conduit (0.56 fold change from NG; p<0.05) while the Schwann cell group revealed significantly higher counts (1.29 fold change from NG; p<0.001). Major histocompatibility complex I (MHC I) molecules were present in significantly increased levels in the DRG and Schwann cell allograft groups compared to the hollow NG conduit and the Sham healthy nerve. Our results confirmed previous studies that have reported Schwann cells as being immunogenic, likely due to MHC I expression. Nerve gap injuries are difficult to repair; our data suggest that DRG neurons are superior medium to implant inside conduit tubes due to reduced immunogenicity and represent a potential treatment strategy that could be preferable to the current gold standard of autologous nerve transplant.     

Sciatic nerve regeneration by transplantation of menstrual blood-derived stem cells

This is the first study demonstrating the efficacy of menstrual blood-derived stem cell (MenSC) transplantation via a neural guidance conduit, for peripheral nerve regeneration. The synthesized poly (?-caprolactone)/Gelatin conduit, filled with collagen type I and seeded with 3?×?10⁴ MenSCs, was implanted into a rat’s 10 mm sciatic nerve defect. The results of hot plate latency, sciatic functional index and weight-loss percentage of wet gastrocnemius muscle demonstrated that the MenSC transplantation had comparable nerve regeneration outcome to autograft, as the gold standard of nerve bridging. The transplantation of MenSCs via a synthetic conduit could ameliorate the functional recovery of sciatic nerve-injured rats which make them a potential candidate for cell therapy of peripheral nervous system disorders.     

Neuroregenerative effects of olfactory ensheathing cells transplanted in a multi-layered conductive nanofibrous conduit in peripheral nerve repair in rats

Background

The purpose of this study was to evaluate the efficacy of a multi-layered conductive nanofibrous hollow conduit in combination with olfactory ensheathing cells (OEC) to promote peripheral nerve regeneration. We aimed to harness both the topographical and electrical cues of the aligned conductive nanofibrous single-walled carbon nanotube/ poly (_L-lactic acid) (SWCNT/PLLA) scaffolds along with the neurotrophic features of OEC in a nerve tissue engineered approach.

Results

We demonstrated that SWCNT/PLLA composite scaffolds support the adhesion, growth, survival and proliferation of OEC. Using microsurgical techniques, the tissue engineered nerve conduits were interposed into an 8 mm gap in sciatic nerve defects in rats. Functional recovery was evaluated using sciatic functional index (SFI) fortnightly after the surgery. Histological analyses including immunohistochemistry for S100 and NF markers along with toluidine blue staining (nerve thickness) and TEM imaging (myelin sheath thickness) of the sections from middle and distal parts of nerve grafts showed an increased regeneration in cell/scaffold group compared with cell-free scaffold and silicone groups. Neural regeneration in cell/scaffold group was very closely similar to autograft group, as deduced from SFI scores and histological assessments.

Conclusions

Our results indicated that the tissue engineered construct made of rolled sheet of SWCNT/PLLA nanofibrous scaffolds and OEC could promote axonal outgrowth and peripheral nerve regeneration suggesting them as a promising alternative in nerve tissue engineering.     

三维取向PLGA神经导管促进坐骨神经再生的实验研究

目的:探讨应用改进静电纺丝技术一次成型制备三维(3D)取向聚乳酸与聚羟基乙酸共聚物(PLGA)纳米神经导管的可行性,检测其对坐骨神经再生的促进作用。方法:应用改进的静电纺丝技术制备无缝取向PLGA纳米神经导管,通过扫描电镜和透射电镜检测支架的纳米结构;分别制备取向和非取向纳米纤维支架修复13mm坐骨神经缺损模型。36只成年SD大鼠随机分为3组(每组12只),A组:非取向PLGA神经导管组(阴性对照);B组:取向PLGA神经导管组,C组:自体神经移植组(阳性对照),于术后3月通过大体观察、行走足印分析、腓肠肌萎缩率、电生理检测、组织形态学检测、透射电镜检测及图像分析,评价无缝取向PLGA纳米神经导管修复坐骨神经缺损的效果。结果:神经导管修复神经缺损三月后,大体观察显示神经导管结构完整,无坍塌和断裂;各组再生神经均有通过神经导管长入远端。B组与C组的腓肠肌萎缩率和神经电传导速度无统计学差异(P0.05),均优于A组。B组与C组再生神经纤维数量及成熟程度均要明显优于A组。结论:无缝取向PLGA纳米神经导管能够诱导并促进神经再生,提高坐骨神经再生的质量,有望成为自体神经移植的替代物。     

Use new PLGL-RGD-NGF nerve conduits for promoting peripheral nerve regeneration

ABSTRACT: BACKGROUND: Nerve conduits provide a promising strategy for peripheral nerve injury repair. However, the efficiency of nerve conduits to enhance nerve regeneration and functional recovery is often inferior to that of autografts. Nerve conduits require additional factors such as cell adhesion molecules and neurotrophic factors to provide a more conducive microenvironment for nerve regeneration. METHODS: In the present study, poly{(lactic acid)-co-[(glycolic acid)-alt-(L-lysine)]} (PLGL) was modified by grafting Gly-Arg-Gly-Asp-Gly (RGD peptide) and nerve growth factor (NGF) for fabricating new PLGL-RGD-NGF nerve conduits to promote nerve regeneration and functional recovery. PLGL-RGD-NGF nerve conduits were tested in the rat sciatic nerve transection model. Rat sciatic nerves were cut off to form a 10 mm defect and repaired with the nerve conduits. All of the 32 Wistar rats were randomly divided into 4 groups: group PLGL-RGD-NGF, group PLGL-RGD, group PLGL and group autograft. At 3 months after surgery, the regenerated rat sciatic nerve was evaluated by footprint analysis, electrophysiology, and histologic assessment. Experimental data were processed using the statistical software SPSS 10.0. RESULTS: The sciatic function index value of groups PLGL-RGD-NGF and autograft was significantly higher than those of groups PLGL-RGD and PLGL. The nerve conduction velocities of groups PLGL-RGD-NGF and autograft were significantly faster than those of groups PLGL-RGD and PLGL. The regenerated nerves of groups PLGL-RGD-NGF and autograft were more mature than those of groups PLGL-RGD and PLGL. There was no significant difference between groups PLGL-RGD-NGF and autograft. CONCLUSIONS: PLGL-RGD-NGF nerve conduits are more effective in regenerating nerves than both PLGL-RGD nerve conduits and PLGL nerve conduits. The effect is as good as that of an autograft. This work established the platform for further development of the use of PLGL-RGD-NGF nerve conduits for clinical nerve repair.     

嗅鞘细胞复合PLGA导管修复周围神经缺损的研究

探讨嗅鞘细胞(OECs)复合聚乳酸-聚羟基乙酸共聚物(PLGA)导管对大鼠坐骨神经缺损的修复作用。方法:SD大鼠80只,随机分成4组,切除右侧部分神经干造成10mm的神经缺损。OECs PLGA组用充满细胞外基质凝胶和OECs悬液(CM-DiI预标记)的PLGA导管桥接坐骨神经缺损;OECs 硅胶管组用含相同内容物的硅胶管桥接;PLGA组和硅胶管组则分别用充满细胞外基质凝胶和DMEM/F12培养基的PLGA导管和硅胶管桥接。术后每周进行感觉运动功能检测,8周时行腓肠肌湿重恢复率、乙酰胆碱脂酶(AChE)染色、电生理和组织形态学分析等检测,同时移植细胞的两组每周进行细胞示踪观察。结果:移植细胞沿神经纵轴分布;除坐骨神经功能指数(SFI)指标外,OECs PLGA组的各项再生功能指标均优于其它三组。结论:OECs复合PLGA导管能够促进再生神经的成熟和靶组织功能的恢复,二者联合移植是一种有效的周围神经缺损修复方法。     

A randomized prospective study of polyglycolic acid conduits for digital nerve reconstruction in humans

This article reports the first randomized prospective multicenter evaluation of a bioabsorbable conduit for nerve repair. The study enrolled 98 subjects with 136 nerve transections in the hand and prospectively randomized the repair to two groups: standard repair, either end-to-end or with a nerve graft, or repair using a polyglycolic acid conduit. Two-point discrimination was measured by a blinded observer at 3, 6, 9, and 12 months after repair. There were 56 nerves repaired in the control group and 46 nerves repaired with a conduit available for follow-up. Three patients had a partial conduit extrusion as a result of loss of the initially crushed skin flap. The overall results showed no significant difference between the two groups as a whole. In the control group, excellent results were obtained in 43 percent of repairs, good results in 43 percent, and poor results in 14 percent. In those nerves repaired with a conduit, excellent results were obtained in 44 percent, good results in 30 percent, and poor results in 26 percent (p = 0.46). When the sensory recovery was examined with regard to length of nerve gap, however, nerves with gaps of 4 mm or less had better sensation when repaired with a conduit; the mean moving two-point discrimination was 3.7 +/- 1.4 mm for polyglycolic acid tube repair and 6.1 +/- 3.3 mm for end-to-end repairs (p = 0.03). All injured nerves with deficits of 8 mm or greater were reconstructed with either a nerve graft or a conduit. This subgroup also demonstrated a significant difference in favor of the polyglycolic acid tube. The mean moving two-point discrimination for the conduit was 6.8 +/- 3.8 mm, with excellent results obtained in 7 of 17 nerves, whereas the mean moving two-point discrimination for the graft repair was 12.9 +/- 2.4 mm, with excellent results obtained in none of the eight nerves (p < 0.001 and p = 0.06, respectively). This investigation demonstrates improved sensation when a conduit repair is used for nerve gaps of 4 mm or less, compared with end-to-end repair of digital nerves. Polyglycolic acid conduit repair also produces results superior to those of a nerve graft for larger nerve gaps and eliminates the donor-site morbidity associated with nerve-graft harvesting.     

A Silk Fibroin/Collagen Nerve Scaffold Seeded with a Co-Culture of Schwann Cells and Adipose-Derived Stem Cells for Sciatic Nerve Regeneration

As a promising alternative to autologous nerve grafts, tissue-engineered nerve grafts have been extensively studied as a way to bridge peripheral nerve defects and guide nerve regeneration. The main difference between autogenous nerve grafts and tissue-engineered nerve grafts is the regenerative microenvironment formed by the grafts. If an appropriate regenerative microenvironment is provided, the repair of a peripheral nerve is feasible. In this study, to mimic the body’s natural regenerative microenvironment closely, we co-cultured Schwann cells (SCs) and adipose-derived stem cells (ADSCs) as seed cells and introduced them into a silk fibroin (SF)/collagen scaffold to construct a tissue-engineered nerve conduit (TENC). Twelve weeks after the three different grafts (plain SF/collagen scaffold, TENC, and autograft) were transplanted to bridge 1-cm long sciatic nerve defects in rats, a series of electrophysiological examinations and morphological analyses were performed to evaluate the effect of the tissue-engineered nerve grafts on peripheral nerve regeneration. The regenerative outcomes showed that the effect of treatment with TENCs was similar to that with autologous nerve grafts but superior to that with plain SF/collagen scaffolds. Meanwhile, no experimental animals had inflammation around the grafts. Based on this evidence, our findings suggest that the TENC we developed could improve the regenerative microenvironment and accelerate nerve regeneration compared to plain SF/collagen and may serve as a promising strategy for peripheral nerve repair.     

三维取向PLGA神经导管促进坐骨神经再生的实验研究

目的：探讨应用改进静电纺丝技术一次成型制备三维（3D）取向聚乳酸与聚羟基乙酸共聚物（PLGA）纳米神经导管的可行性,检测其对坐骨神经再生的促进作用。方法：应用改进的静电纺丝技术制备无缝取向PLGA纳米神经导管,通过扫描电镜和透射电镜检测支架的纳米结构;分别制备取向和非取向纳米纤维支架修复13mm坐骨神经缺损模型。36只成年SD大鼠随机分为3组（每组12只）,A组：非取向PLGA神经导管组（阴性对照）;B组：取向PLGA神经导管组,C组：自体神经移植组（阳性对照）,于术后3月通过大体观察、行走足印分析、腓肠肌萎缩率、电生理检测、组织形态学检测、透射电镜检测及图像分析,评价无缝取向PLGA纳米神经导管修复坐骨神经缺损的效果。结果：神经导管修复神经缺损三月后,大体观察显示神经导管结构完整,无坍塌和断裂;各组再生神经均有通过神经导管长入远端。B组与C组的腓肠肌萎缩率和神经电传导速度无统计学差异（P〈0.05）,均优于A组。B组与C组再生神经纤维数量及成熟程度均要明显优于A组。结论：无缝取向PLGA纳米神经导管能够诱导并促进神经再生,提高坐骨神经再生的质量,有望成为自体神经移植的替代物。     

Engineering a prokaryotic apocytochrome c as an efficient substrate for Saccharomyces cerevisiae cytochrome c heme lyase

In spite of the extensive research using induced pluripotent stem (iPS) cells, the therapeutic potential of iPS cells in the treatment of peripheral nerve injury is largely unknown. In this study, we repaired peripheral nerve gaps in mice using tissue-engineered bioabsorbable nerve conduits coated with iPS cell-derived neurospheres. The secondary neurospheres derived from mouse iPS cells were suspended in each conduit (4000,000 cells per conduit) and cultured in the conduit in three-dimensional (3D) culture for 14 days. We then implanted them in the mouse sciatic nerve gaps (5 mm) (iPS group; n=10). The nerve conduit alone was implanted in the control group (n=10). After 4, 8 and 12 weeks, motor and sensory functional recovery in mice were significantly better in the iPS group. At 12 weeks, all the nerve conduits remained structurally stable without any collapse and histological analysis indicated axonal regeneration in the nerve conduits of both groups. However, the iPS group showed significantly more vigorous axonal regeneration. The bioabsorbable nerve conduits created by 3D-culture of iPS cell-derived neurospheres promoted regeneration of peripheral nerves and functional recovery in vivo. The combination of iPS cell technology and bioabsorbable nerve conduits shows potential as a future tool for the treatment of peripheral nerve defects.     

神经生长因子与冻干异体神经桥接大鼠神经缺损的研究

实验采用冻干处理的异体神经与外源性神经生长因子（ＮＧＦ）结合来桥接大鼠的坐骨神经１．０ｃｍ的缺损。用雄性Ｗｉｓｔａｒ大鼠进行的四组实验结果表明：冻干处理的异体神经可降低其抗原性,但处理后并不损害雪旺氏细胞（ＳＣ）基底膜的完整性,在移植后可能成为轴突再生的通道和支架;外源性ＮＧＦ与冻干神经结合形成的复合体,可为神经的再生提供一个较好的微环境,具有成为理想桥接材料的可能性     

An alternative to the classical nerve graft for the management of the short nerve gap

Reconstruction of a short nerve gap by a nerve graft produces donor-site scarring, loss of donor nerve function, and neuroma formation. This study compared the regeneration achieved after 1 year in 16 monkeys across a 3-cm upper arm ulnar nerve gap with a bioabsorbable polyglycolic acid nerve conduit with the regeneration achieved with a classical interfascicular interpositional sural nerve graft. The results demonstrated electrophysiologic and histologic evidence of neural regeneration across the gaps in all experimental groups. The bioabsorbable nerve conduit groups and the sural nerve graft group had mean fiber diameters, amplitudes, and conduction velocities each significantly less than those of normal control ulnar nerves. There was, however, no significant difference between any of the experimental groups. Electromyography demonstrated recovery of 19 of the 28 (68 percent) intrinsic muscles studied. These results demonstrate that the primate peripheral nerve can regenerate across short nerve gaps when guided by an appropriate nerve conduit, suggesting that a single-stage biodegradable polyglycolic acid conduit may be used as an alternative to a short interfascicular nerve graft.     

Axonal Regeneration and Remyelination Evaluation of Chitosan/Gelatin-Based Nerve Guide Combined with Transforming Growth Factor-β1 and Schwann Cells

Despite efforts in peripheral nerve injury and regeneration, it is difficult to achieve a functional recovery following extended peripheral nerve lesions. Even if artificial nerve conduit, cell components and growth factors can enhance nerve regeneration, integration in peripheral nerve repair and regeneration remains yet to be explored. For this study, we used chitosan/gelatin nerve graft constructed with collagenous matrices as a vehicle for Schwann cells and transforming growth factor-β1 to bridge a 10-mm gap of the sciatic nerve and explored the feasibility of improving regeneration and reinnervation in rats. The nerve regeneration was assessed with functional recovery, electrophysiological test, retrograde labeling, and immunohistochemistry analysis during the post-operative period of 16 weeks. The results showed that the internal sides of the conduits were compact enough to prevent the connective tissues from ingrowth. Nerve conduction velocity, average regenerated myelin area, and myelinated axon count were similar to those treated with autograft (p > 0.05) but significantly higher than those bridged with chitosan/gelatin nerve graft alone (p < 0.05). Evidences from retrograde labeling and immunohistochemistry analysis are further provided in support of improving axonal regeneration and remyelination. A designed graft incorporating all of the tissue-engineering strategies for peripheral nerve regeneration may provide great progress in tissue engineering for nerve repair.     

Increased muscle regeneration after repair of divided motor nerve with neuronotrophic factors containing glue

Neuronotrophic factors (NTFs) directed to spinal cord motor neurons were collected in rats within silicone nerve regeneration chambers according to LONGO et al. (1983b). Unilateral addition of NTFs to the fibrin glue used for the repair of divided sciatic nerves improved locally nerve regeneration without affecting the controlateral side. Nerve regeneration was assessed by weight gain of the reinnervated muscles and by radioactive labelling of the acid-soluble phosphate fractions of both nerve Schwann cells and reinnervated muscle cells. Fast gastrocnemius and slow soleus muscles, the motor nerve of which had been repaired with added NTFs, were significantly heavier (21 and 28%) than their controlateral controls, and the metabolic dedifferentiation attendant on post-division nerve repair was less marked. It is suggested that this experimental nerve regeneration model is suitable to test potential nerve-active agents in vivo, under conditions close to the usual clinical setting, with, as ultimate goal, the improvement of the end-results of microsurgical repair of peripheral nerve in man.     

Effects of nerve growth factor on nerve regeneration through a vein graft across a gap.

The limited availability of donor sites for nerve grafts and their inherent associated morbidity continue to stimulate research toward finding suitable alternatives. In the following study, the effect of direct administration of nerve growth factor (NGF) into a nerve conduit across a gap was tested in a rat sciatic nerve model. A 1-cm segment of the right sciatic nerve in Sprague-Dawley rats was resected, and the gap was then bridged using one of three methods: group I (NGF-treated group, n = 12), a vein graft filled with NGF (100 ng in 0.3-ml phosphate buffered saline); group II (control group, n = 12), a vein graft filled with phosphate buffered saline only; group III (standard nerve graft, n = 11), a resected segment of the sciatic nerve. All animals were evaluated at 3 and 5 weeks by behavioral testing and at 5 weeks by electrophysiologic testing. At 3 weeks, sensory testing showed that the latency to a noxious stimulus in group I animals (8.0 +/- 5.4 sec, mean +/- SD) was significantly lower than that of group II animals (13.2 +/- 6.5 sec), indicating that sensory recovery was superior in the animals receiving NGF. The mean latency of animals in group III was 12.9 +/- 6.5 sec, but the difference between the latencies of group I and group III did not reach statistical significance. At 5 weeks, there was no difference in sensory testing between groups. Motor function in groups I and III as measured by walk pattern analysis was superior to that of group II at 5 weeks (toe spread ratios 0.66 +/- 0.09, 0.48 +/- 0.07, and 0.69 +/- 0.09 for groups I, II, and III, respectively). Mean motor conduction velocities across the 1-cm gap were 8.6 +/- 4.7 m/sec, 2.5 +/- 0.7 m/sec, and 6.9 +/- 2.9 m/sec in groups I, II, and III respectively. The difference between groups I and III was not statistically significant, but the motor conduction velocity of group II was significantly slower than that of either group I or III (p < 0.002). The positive effects of NGF on regeneration of nerves across a gap seen in this study suggest that it may be useful for treating peripheral nerve injuries in combination with autogenous vein grafts.     

Anti-CD40 ligand antibody permits regeneration through peripheral nerve allografts in a nonhuman primate model

Systemic immunosuppression is typically required to prevent allograft rejection. Antibody-based therapies that induce immune unresponsiveness represent an appealing alternative to nonspecific immunosuppression, which is often associated with significant morbidity. In mice, successful prevention of nerve allograft rejection has been demonstrated through interference with the CD40/CD40 ligand interaction. This study investigated the effectiveness of anti-CD40 ligand monoclonal antibody as single-agent therapy in preventing rejection and supporting nerve regeneration across long nerve allografts in nonhuman primates. Twelve outbred cynomolgus macaques were arranged into six genetically mismatched pairs, with each animal receiving a 5-cm ulnar nerve allograft in the right arm and a 5-cm autograft in the left arm. Mixed lymphocyte reaction assays were used to assess resulting immune unresponsiveness. Treated animals (n = 10) received anti-CD40 ligand monoclonal antibody 10 mg/kg one time, locally applied, and 20 mg/kg systemically on postoperative days 0, 1, 3, 10, 18, and 28, and then monthly. Untreated animals (n = 2) served as the untreated controls. At 4 or 6 months after transplantation, nerves were harvested for histological analysis. Four treated animals underwent an additional challenge after cessation of anti-CD40 ligand monoclonal antibody therapy and nerve graft harvests. Autogenous and allogeneic skin and nerve inlay grafting was performed to assess the permanence of immune unresponsiveness induced by anti-CD40 ligand monoclonal antibody. Animals that received anti-CD40 ligand monoclonal antibody demonstrated robust regeneration across nerve allografts, similar to that seen in the autograft control in the contralateral arm. The histomorphometric analysis of allografts in the untreated animals demonstrated significantly worse measurements compared with their matched autograft controls. Animals that received anti-CD40 ligand monoclonal antibody with concomitant skin allografts had virtually no evidence of nerve regeneration through allografts. Allogeneic skin and nerve allografts applied 2 to 12 months after withdrawal of anti-CD40 ligand monoclonal antibody therapy were consistently rejected. This study demonstrates that anti-CD40 ligand monoclonal antibody prevents rejection and allows regeneration of peripheral nerve allografts in nonhuman primates. The effect of anti-CD40 ligand monoclonal antibody appears to be transient, however, with restoration of immunocompetence shortly after withdrawal of therapy.     

The peripheral nerve allograft: an assessment of regeneration in the immunosuppressed host

Regeneration across the nerve allograft in the immunosuppressed host was assessed using electrical and histologic parameters. The Lewis rat (RTIl) served as the recipient animal, and ACI rats (RTIa) provided the donor nerve allografts. Hydrocortisone and azathioprine were used in various dose schedules as the immunosuppressive agents. Animals were immunosuppressed for either 30 or 100 days. Histologic and electrophysiologic measurements of nerve regeneration were assessed at 30, 100, and 180 days. The degree of nerve regeneration was similar in all experimental groups. Short-term, low-dose immunosuppression was as successful as longer-term, higher-dose immunosuppression therapy. The degree of nerve regeneration in all experimental groups was significantly better than that in the fresh, untreated nerve allograft control group (Lewis/ACI) but was not as good as that seen in the autograft control group (Lewis/Lewis).     

Improving Nerve Regeneration of Acellular Nerve Allografts Seeded with SCs Bridging the Sciatic Nerve Defects of Rat

The objective of the paper is to evaluate the effect of acellular nerve allografts (ANA) seeded with Schwann cells to promote nerve regeneration after bridging the sciatic nerve defects of rats and to discuss its acting mechanisms. Schwann cells were isolated from neonatal Wistar rats. In vitro Schwann cells were microinjected into acellular nerve allografts and co-cultured. Twenty-four Wistar rats weighing 180–220 g were randomly divided into three groups with eight rats in each group: ANA seeded with Schwann cells (ANA + SCs), ANA group and autografts group. All the grafts were, respectively, served for bridging a 10-mm long surgically created sciatic nerve gap. Examinations of regeneration nerve were performed after 12 weeks by transmission electron microscope (TEM), scanning electron microscope (SEM), and electrophysiological methods, and then analyzed statistically. The results obtained indicated that in vitro Schwann cells displayed the feature of bipolar morphology with oval nuclei. Compared with ANA group, the conduction velocity of ANA + SCs group and autograft group was faster after 12 weeks, latent period was shorter, and wave amplitude was higher (P < 0.05). The difference between ANA + SCs group and autograft group is not significant (P > 0.05). Regeneration nerve myelinated fiber number, myelin sheath thickness, and myelinated fibers/total nerves (%) in both ANA + SCs group and autograft group are higher than that in ANA group; the difference is significant (P < 0.05). The difference between the former two is not significant (P > 0.05). In conclusion, ANA seeded with SCs could improve nerve regeneration and functional recovery after bridging the sciatic nerve gap of rats, which offers a novel approach for the repair peripheral nerve defect.     

Enhancement of nerve regeneration and orientation across a gap with a nerve graft within a vein conduit graft: a functional, stereological, and electrophysiological study

In this study, the right sciatic nerves of 40 rats were used to determine whether a nerve graft within a vein graft might accelerate and facilitate axonal regeneration, compared with a nerve graft alone. The animals were separated into four groups, as follows: group 1, sham control; group 2 (control), segmental nerve resection and no repair; group 3, segmental nerve resection and nerve grafting; group 4, segmental nerve resection and reconstruction with a nerve graft within a vein conduit graft. For all groups, sciatic functional indices were calculated before the operation and on postoperative days 7 and 90. On postoperative day 90, the sciatic nerves were reexposed and nerve conduction velocities were recorded. The sciatic nerves were harvested from all groups for counting of the myelinated axons with a stereological method. No statistically significant differences with respect to return of gait function, axon count, or nerve conduction were noted between groups 3 and 4 (p > 0.05). However, functional recovery in group 4 on postoperative day 90 was significant, compared with group 2 (p < 0.05); the recovery difference between groups 2 and 3 was not significant (p > 0.05). This study was not able to demonstrate any functional benefits with the use of a nerve graft within a vein graft, compared with standard nerve grafting.     

Functional evaluation of complete sciatic, peroneal, and posterior tibial nerve lesions in the rat

Quantification of peripheral nerve regeneration in animal studies of nerve injury and repair by histologic, morphologic, and electrophysiologic parameters has been controversial because such studies may not necessarily correlate with actual nerve function. This study modifies the previously described sciatic functional index (SFI), tibial functional index (TFI), and peroneal functional index (PFI) based on multiple linear regression analysis of factors derived from measurements of walking tracks in rats with defined nerve injuries. The factors that contributed to these formulas were print-length factor (PLF), toe-spread factor (TSF), and intermediary toe-spread factor (ITF). It was shown that animals with selective nerve injuries gave walking tracks that were consistent, predictable, and based on known neuromuscular deficits. The new formula for sciatic functional index was compared with previously described indices. The sciatic functional index, tibial functional index, and peroneal functional index offer the peripheral nerve investigator a noninvasive quantitative assessment of hindlimb motor function in the rat with selective hindlimb nerve injury.