植入式电刺激治疗兔坐骨神经损伤的实验研究

[目的]通过建立新西兰大自兔急性坐骨神经损伤模型,验证植入式电刺激系统对于神经生长、防止肌肉萎缩的作用.[方法]选取20只健康成年新西兰大白兔,雌雄不限,建立急性坐骨神经损伤模型.随机分为两组,实验组10只,对照组10只.左侧为实验侧,同时行自身对侧对照.模型成功建立2周后,于原神经外膜标记损伤缝线的近端与远端肌肉组织埋入电极,以磁力棒控制电刺激器的启动,对动物实施每日8h的间歇电刺激.术后2、4周,观察各组动物行为学改变情况,检测坐骨神经的传导速度,测定腓肠肌湿重恢复率.分别于术后2、4周电生理检测后,处死动物并取材行组织学检测.取脊髓组织,行分子生物学检测.[结果]术后2、4周实验组手术侧胫神经、腓浅神经传导速度均明显快于对照组(P<0.05),实验组肌湿重恢复率明显高于对照组(P<0.05).4周后实验组HE染色肌纤维退行性改变较轻,对照组肌纤维细胞萎缩明显,间质可见炎性渗出,局部可见纤维化.实验组、对照组肌细胞均表达神经生长因子(nerve growth factor,NGF),2、4周间两两比较均有差异(P<0.05).电镜观察实验组神经轴突髓鞘结构正常;对照组神经均有髓鞘松弛、弯曲,髓鞘中间可见变性.RT-PCR各实验组与对照组均可见β-actin,nNOS mRNA,NGF tnRNA的表达,实验组与对照组2、4周间两两比较均有差异(P<0.05).[结论]在合适的刺激参数下,植入式电刺激系统有利于促进兔急性坐骨神经损伤的再生.     

补阳还五汤对骨髓间质干细胞移植治疗大鼠脊髓损伤的影响

目的:观察补阳还五汤联合骨髓间质干细胞(MSCs)移植对大鼠脊髓损伤后神经功能恢复以及移植的MSCs迁移情况的影响,并探讨其作用机制。方法:SD大鼠80只,其中70只用改良Allen法制备大鼠T10脊髓损伤模型,并随机分为中药 MSCs组20只、MSCs组20只、假手术组(无脊髓损伤)10只、中药组20只、空白对照组(无治疗)10只。中药 MSCs组、MSCs组、假手术组行大鼠尾静脉移植带Brdu标记的MSCs。各组大鼠于术后1、3、5周观察神经功能恢复、免疫组化检测带标记的MSCs迁移情况。结果:与空白对照组相比,治疗组神经功能测定在术后1、3、5周时均明显高于对照组(P<0·01)。术后1周移植组的脊髓组织内即可见Brdu标记阳性细胞(假手术组除外),术后5周中药 MSCs组Brdu阳性细胞计数较MSCs组有显著性差异(P<1.05)。结论:静脉注射移植的MSCs能够迁移到脊髓损伤组织,并促进神经功能的恢复。补阳还五汤能促进移植的MSCs迁移,同时有利于脊髓功能的恢复。     

成年大鼠许旺细胞复合去细胞神经支架修复坐骨神经缺损

目的探讨成年许旺细胞(SC)复合去细胞神经支架构建的组织工程化人工神经移植治疗外周神经损伤的效果。方法从Wallerian变性1周的成年SD大鼠远侧端神经中分离培养得到SC,复合去细胞神经支架构建组织工程化人工神经。移植分为SC+去细胞SD大鼠神经支架组(SC治疗组)和空细胞SD大鼠神经支架组(阴性对照组),每组各5只。比较两组大鼠术后2、4、8周损伤侧坐骨神经功能指数(SFI),术后8周损伤侧坐骨神经传导功能和小腿三头肌湿重恢复率等修复效果的指标。结果 SC治疗组术后2、4、8周损伤侧SFI,术后8周损伤侧坐骨神经传导功能和小腿三头肌湿重恢复率均优于或高于阴性对照组(均为P0.05)。结论采用成年SC复合去细胞神经支架的人工神经移植治疗外周神经缺损,可有效促进损伤神经的功能恢复。     

骨髓间充质干细胞移植促进脊髓损伤大鼠神经功能的恢复

目的 探讨骨髓间充质干细胞(MSCs)移植对脊髓损伤大鼠神经功能恢复的影响极其可能机制.方法 采集SD大鼠骨髓,分离出MSCs,并培养、纯化,取传至第6代的MSCs,移植前1 d以5溴2脱氧尿嘧啶核苷(Brdu)标记,然后用显微注射器将其缓慢注射到脊髓损伤大鼠模型(成年SD大鼠)的受损脊髓中心,以不进行移植的脊髓损伤大鼠模型(损伤组)和假手术组为对照.术后进行运动功能评分以判断神经功能恢复情况,并切取移植区域脊髓组织,应用荧光免疫化学染色检测脊髓损伤灶边周的神经元凋亡情况(TUNEI.与NeuN双标记)以及移植的MSCs的分布和向神经元分化情况(Brdu与NeuN双标记),应用逆转录聚合酶链反应检测损伤灶区域脑源性神经营养因子(BDNF)mRNA的表达.结果 移植后第3天开始,损伤组和移植组大鼠的BBB运动功能评分明显上升,第14天后进入平台期,但移植组各时间点的.BBB运动功能评分均明显高于损伤组(P<0.05).移植后3 d,在移植组的脊髓组织中可见Brdu标记阳性细胞,少部分Brdu标记阳性细胞同时表达神经元特异性标志物NeuN;移植后第3、7天,损伤组和移植组脊髓组织中的凋亡神经元均显著多于假手术组(P<0.01),凋亡的神经元多存在于损伤周边区,但移植组的凋亡神经元显著少于损伤组(P<0.01);移植后第3、7天,假手术组未检测到BDNF mRNA的表达,损伤组和移植组均可检测到BDNF mRNA的表达,移植组的表达量分别较损伤组高28.6%和39.2%(P<0.05).结论 移植的骨髓MSCs可促进脊髓损伤大鼠神经功能的恢复,其机制除MSCs直接分化为神经元进行修复外,还可能与其改善脊髓损伤区的微环境、上调BDNF、mRNA表达、减少神经细胞凋亡有关.     

静脉移植骨髓间充质干细胞促进脊髓损伤后GDNF基因的表达

[目的]探讨经静脉移植骨髓间充质干细胞(MSCs)对大鼠脊髓损伤(SCI)后胶质细胞源性神经营养因子(GDNF)表达的影响。[方法]MSCs提取自成年Wistar大鼠的股骨干骨髓,经原代培养、鉴定及5-溴脱氧尿嘧啶核苷(Brdu)核标记。以改良Allen′s打击装置制作大鼠T10节段脊髓损伤(SCl)模型,于伤后立即缝合切口并经尾静脉注射移植MSCs。实验共分为3组MSCs尾静脉移植组(A组)、生理盐水注射组(B组)、正常对照组(C组)。术后不同时间点应用免疫组化法和逆转录聚合酶链反应法(RT-PCR)观察MSCs移植后的存活状态以及不同时间点GDNF基因的表达变化。[结果]免疫组化结果MSCs经尾静脉移植后在损伤脊髓平面可以检测到迁移及存活,标记细胞呈棕黄色的核标记,以损伤区域为多并向周围迁移,移植术后第14d,A组Brdu阳性细胞较多,最远于距离损伤区2.5cm处可检测到。B及C组则均未检测到阳性细胞。RT-PCR结果移植术后第1、3、5d,A组表达量呈逐渐升高趋势,B组呈一过性表达升高,C组无变化。各时间点A组GDNFmRNA的表达量明显高于B组,P<0.05。免疫组化结果移植术后第7、14、28dGDNF的表达量明显高于B组,P<0.05。[结论]骨髓间充质干细胞经尾静脉移植后可迁移至脊髓损伤区域,并上调胶质细胞源性神经营养因子基因的表达,是该移植方式修复大鼠脊髓损伤的机制之一。     

神经断端肌同埋入防治残端神经瘤的实验研究

周围神经切断端发生神经瘤是周围神经损伤的常见并发症，经１０％患者有顽固性疼痛。为研究神经断端肌内埋入防治残端神经瘤的机理，选用ＳＤ大白鼠１６只，将双侧坐骨神经切断后，左则神经经断端肌内埋入为实验侧，右侧神经断端自然回缩不作处理为对照侧，运用组织学和电生理学检测。结果表明，对照侧的神经近端在术后１个月就有神经瘤形成，而实验侧其神经断端的神经纤维分散长入肌纤维间，无明确的神经瘤形成。说明，神经断端肌内     

神经断端肌内埋入防治残端神经瘤的实验研究

周围神经切断端发生神经瘤是周围神经损伤的常见并发症，约１０％患者有顽固性疼痛。为研究神经断端肌内埋入防治残端神经瘤的机理，选用ＳＤ大白鼠１６只，将双侧坐骨神经切断后，左侧神经断端肌内埋入为实验侧，右侧神经断端自然回缩不作处理为对照侧，运用组织学和电生理学检测。结果表明，对照侧的神经近端在术后１个月就有神经瘤形成，而实验侧其神经断端的神经纤维分散长入肌纤维间，无明确的神经瘤形成。说明，神经断端肌内埋入可以防治残端神经瘤形成。     

丙戊酸钠充填导管促进大鼠外周神经再生的实验研究

目的 观察丙戊酸钠(VPA)充填导管对缺损外周神经再生的促进作用.方法 通过建立大鼠坐骨神经缺损模型.用硅胶管(1 cm)进行缺损神经段(0.8 cm)的桥接,局部应用8%VPA注射液10ul,观察VPA对神经再生和运动神经功能恢复的影响.30只大鼠随机分成2组,实验组在硅胶管局部注射VPA注射液;对照组在硅胶管局部注入生理盐水. 结果 术后每只大鼠每2周进行坐骨神经功能指数(SFI)检测,每4周做电生理检查,最后术后12周处死所有大鼠,对坐骨神经进行组织形态学分析.用数字图像分析软件检测有髓神经纤维髓鞘厚度.并对再生神经纤维轴突计数.通过统计软件分析发现SFI,电生理检查,神经组织性形态上两组差异均有统计学意义(P＜0.05). 结论 局部使用VPA于神经缺损的大鼠,可以促进损伤神经的轴突再生和运动功能恢复.因此VPA有望在临床上应用于外周神经损伤病例的治疗.     

无细胞的异体神经修复鼠坐骨神经缺损

目的 通过化学萃取同种异体神经,去除髓鞘和雪旺细胞,形成无细胞基膜管后桥接鼠坐骨神经缺损,研究神经再生效果。方法 正常鼠坐骨神经用非变性生物剂处理后得到无细胞的基膜管,桥接鼠坐骨神经20mm缺损。实验分3组:无细胞基膜管移植组(A组),自体神经移植组(B组)和异体神经移植组(C组)。术后进行肌电图、光镜、电镜及图象分析仪检查。结果 A组再生神经有大量轴突通过移植体,术后2个月电生理检测再生神经的潜伏期及波幅低于B组(P<0.05),术后3个月2组差异无显著意义。髓鞘厚度在术后3个月时亦低于B组,差异有显著意义(P<0.05)。轴突直径及数目两组无差异。C组因无神经再生,结果无法测量。结论 这种无细胞基膜管移植体能支持轴突的生长和雪旺细胞的迁移,是一种良好的神经移植替代材料。     

断端间距对周围神经再生影响的实验研究

目的 探索较佳修复周围神经损伤的方法。方法 以SD大鼠坐骨神经为实验模型,观察神经切断后保留不同间隙修复后的神经再生情况。术后6周、8周处死大鼠,分别行电生理检测、腓肠肌湿重观察、组织学观察及神经轴突计数。结果 保留3mm神经断端间隙时,各项检查结果均优于其他各组。结论 保留3mm神经断端间隙,术后神经功能恢复最好,可有效地发挥神经再生的趋化作用,且对轴突再生速度无影响。     

Functional recovery of sciatic nerve through inside-out vein graft in rats

Objective: Present study aimed at further comprehensive functional, histomorphometrical and immunohistochemical assessment of peripheral nerve regeneration using rat sciatic nerve transection model.Methods: The 10-mm rat sciatic nerve gap was created in rats. In control group nerve stumps were sutured to adjacent muscle and in treatment group the gap was bridged using an inside-out vein graft. In sham-operated group the nerve was manipulated and left intact. All animals underwent walking track analysis test 4, 8, and 12 weeks after surgery.Subsequently, muscle mass measurement was performed to assess reenervation, histological examination to observe the sciatic nerve regeneration morphologically and immunohistochemistry to detect Schwann cells using anti S-100. Results were analyzed using a factorial ANOVA with two between-subjects factors. Bonferroni test for pairwise comparisons was used to examine the effect of treatments.Results: Functional analysis ofmyelinated nerve fibers showed that nerve function improved significantly in the time course in treatment group. However, quantitative morphometrical analysis of myelinated nerve fibers showed that there was no significant difference between 8 and 12 weeks in treatment group. Muscle weight ratio was bigger and weight loss of the gastrocnemius muscle was ameliorated by inside-out vein grafting. The position of positive immunohistochemical reactions further implied that regenerated axons and Schwann cell-like cells existed after vein grafting was performed, and was accompanied by the process of myelination and structural recovery of regenerated nerves.Conclusion: Functional analysis of peripheral nerve repair is far more reliable than quantitative morphometrical analysis     

Repair of nerve defect with acellular nerve graft supplemented by bone marrow stromal cells in mice

The acellular nerve graft that can provide internal structure and extracellular matrix components of the nerve is an alternative for repair of peripheral nerve defects. However, results of the acellular nerve grafting for nerve repair still remain inconsistent. This study aimed to investigate if supplementing bone marrow mesenchymal stromal cells (MSCs) could improve the results of nerve repair with the acellular nerve graft in a 10-mm sciatic nerve defect model in mice. Eighteen mice were divided into three groups (n = 6 for each group) for nerve repairs with the nerve autograft, the acellular nerve graft, and the acellular nerve graft by supplemented with MSCs (5 × 10(5)) fibrin glue around the graft. The mouse static sciatic index was evaluated by walking-track testing every 2 weeks. The weight preservation of the triceps surae muscles and histomorphometric assessment of triceps surae muscles and repaired nerves were examined at week 8. The results showed that the nerve repair by the nerve autografting obtained the best functional recovery of limb. The nerve repair with the acellular nerve graft supplemented with MSCs achieved better functional recovery and higher axon number than that with the acellular nerve graft alone at week 8 postoperatively. The results indicated that supplementing MSCs might help to improve nerve regeneration and functional recovery in repair of the nerve defect with the acellular nerve graft.     

周围神经损伤修复后早期骨骼肌细胞凋亡的初步研究

目的 探讨神经修复后早期大鼠骨骼肌的细胞凋亡情况.方法 取SD大鼠54只,随机分为3组:失神经组(A组)18只,神经缝合组(B组)18只,健康对照组(C组)18只.A组大鼠切除左侧1 cm长坐骨神经,B组大鼠横断左侧坐骨神经后立即用10-0医用尼龙线行外膜缝合,C组大鼠不做任何处理.以腓肠肌肌湿重作为骨骼肌萎缩指标.分别应用脱氧核糖核苷酸末端转移酶介导的缺口末端标记法(TUNEL)和分光光度法检测术后2 d、14 d、28 d时骨骼肌凋亡细胞核和天冬氨酸特异性半胱氨酸蛋白酶(Caspase)-3与Caspase8活性.结果 神经缝合后早期骨骼肌与正常骨骼肌比较,细胞凋亡现象增加,凋亡相关蛋白Caspase-3和Caspase-8活性上升,但程度弱于失神经骨骼肌.结论 细胞凋亡可能是神经修复后早期骨骼肌萎缩原因之一,死亡受体信号通路参与神经修复后早期骨骼肌凋亡过程中.

Abstract：

Objective To explore skeletal muscle apoptosis at the early stage of peripheral never regeneration in rats.Methods A total of 54 male SD rats were randomly assigned to 3 groups ( n = 18/group): denervation group (A), neurorrhaphy group (B), normal control group (C).In group A, a 1 cm segment of the left sciatic nerve was removed.In group B, the left sciatic nerve was transected above the bifurcation and immediately repaired with 10-0 sutures.No surgery was done in group C.The gastrocnemius muscle wet weight served as an indicator of the degree of muscle atrophy.Marker of apoptosis, nuclear DNA fragmentation, was detected using terminal deoxyribnudeotidyl transferase mediated dUTP nick end labeling (the TUNEL method) and observed under confocal microscopy at 2, 14 and 28 days postoperatively.Another portion of the gastrocnemius muscle was homogenized to analyze the activity of caspase-3 and -8 by spectrophotometry.Results TUNEL labeling of fragmented DNA on histological sections in the neuorrhaphy group revealed levels of apoptotic nuclei higher than the control group and lower than the denervation group at the early stage ( ＜ 28days ).The activity of caspase-3 and -8 in the neuorrhaphy group was also higher than the control group but lower than the denervation group.Conclusion At the initial stage of peripheral never regeneration, apoptosis may contribute to muscle atrophy and extrinsic apoptotic pathways may take part in it.     

Effect of prolonged ischaemic time on muscular atrophy and regenerating nerve fibres in transplantation of the rat hind limb

Abstract Our aim was to test the influence of cold ischaemia on replanted limbs, focusing on muscular atrophy and neurological recovery. Inbred wild-type and green fluorescent protein (GFP) transgenic (Tg) Lewis rats aged 8-10 weeks were used. The amputated limbs were transplanted at several cold ischaemic times (0, 1, 8, 12, 24, 48, and 72 hours). An arterial ischaemic model and a denervation model were used as controls. To study nerve regeneration, a GFP limb was transplanted on to the syngenic wild Lewis rat. These animals were evaluated histologically, electrophysiologically, and immunohistochemically. The longer the ischaemic time, the more evident was atrophy of the muscles. Electrophysiological investigation showed that the latency at 3 weeks was longer in the transplantation models than in the normal controls, particularly in the longer ischaemia group. Larger numbers of migrating Schwann cells were seen in the group with no delay than in the group that had been preserved for 12 hours. Ischaemia after amputation of a limb causes muscle cells to necrose and atrophy, and these changes worsen in proportion to the ischaemic preservation time. A delay in nerve regeneration and incomplete paralysis caused by malregeneration also affect muscular atrophy.     

Regeneration capacity of the peripheral nerve: experimental study--final results

Any injury affecting peripheral nerve continuity is followed by a degenerative reaction from nerve itself and its muscle. Nerve regeneration is possible when the two ends are put in direct contact (direct suture), proximity (autologous or artificial grafts), or through muscle neurotization. The aim of this experimental study was to evaluate sciatic nerve degeneration and regeneration in rats after no suture (group 1), direct suture (group 2), nerve grafting (group 3) and muscle neurotization (group 4). Nerve samples were analyzed by optical microscopy at 8 and 16 weeks after surgery, and nerve degeneration (group 1), axonal growth (group 2) and neuromuscular regeneration (group 4) were assessed. Optical microscopy and different staining techniques confirmed nerve degeneration, regeneration, and axonal growth in different moments. Cholinesterase test after direct muscle neurotization showed enzymatic activity at the level of muscle implantation of nerve fibers, indirectly confirming the presence of neuromuscular synapse.     

神经营养素3基因修饰的神经干细胞对周围神经再生修复的影响

目的 探讨神经营养素3基因修饰的神经干细胞对周围神经再生的影响。方法 54只SD大鼠随机分为3组，造成坐骨神经切断损伤模型，神经外膜端端缝合，于小腿三头肌每周分别注射生理盐水、未被神经营养素3基因修饰的神经干细胞、及神经营养素3基因修饰的神经干细胞。术后3、6、9周动态观察坐骨神经功能指数(SFI)以了解后肢功能恢复情况、组织学切片观察、肌湿重恢复率测定、9周后吻合口神经干的电镜观察。结果 神经营养素3基因修饰的神经干细胞组动物的有髓神经纤维密度、神经组织面积、髓鞘厚度、肌湿重以及坐骨神经功能指数均显著优于未被神经营养素3基因修饰的神经干细胞组和生理盐水组，9周时差异有统计学意义。单纯神经干细胞组各项指标优于生理盐水组。结论 将神经营养素3基因修饰的神经干细胞移植于修复的周围神经，使局部释放的NT-3加快轴突再生速度以促进周围神经再生，减缓失神经支配肌肉的萎缩。     

班布特罗与当归补血汤对失神经肌肉萎缩防治作用的实验研究

目的：研究班布特罗及当归补血汤对防治失神经骨骼肌萎缩的作用效果及有无协同作用。方法 SD大鼠40只，随机分组，每组10只，建立失神经骨骼肌萎缩的实验模型。术后4周处死大鼠，双侧腓肠肌及心脏称重，测定肌肉SOD含量，测肌纤维直径，检测细胞凋亡情况。结果 各用药组与对照组肌总蛋白含量、SOD含量、肌纤维直径均无显著性差异（P〉0．05）。在排除个体体重差异后肌肉湿重亦无显著性差异。两种药物之间无协同作用。各组均可见凋亡细胞，荧光显微镜下肉眼观测凋亡细胞的数量无显著性差异。结论 班布特罗与当归补血汤在4周时无明显预防肌萎缩作用及协同作用；细胞凋亡在失神经肌萎缩中发挥一定的作用；本实验为探索药物长期应用防治失神经骨骼肌萎缩的研究提供借鉴。     

锂剂促进周围神经损伤修复的初步研究

目的 探讨锂剂对周围神经损伤后神经再生的影响.方法 取48只雌性SD大鼠,制作大鼠右侧坐骨神经损伤动物模型,通过腹腔注射氯化锂,在不同时间点观察动物下肢活动情况,检测小腿三头肌神经电生理及肌湿重,并对损伤远端神经纤维的神经丝蛋白(NF200)、单核巨噬细胞抗原(ED1)、P-75和运动终板进行免疫组织化学染色观察.结果 损伤后4周,实验组动物下肢已接近正常行走步态,对照组右侧肢体仍明显跛行;损伤后2周和4周,实验组的复合肌肉动作电位(CMAP)波幅较对照组明显增大,两组间的差异有统计学意义(P＜0.05);损伤后4周、8周,实验组的小腿三头肌重量较对照组明显增大,两组间差异有统计学意义(P＜0.05);损伤后3 d,在距离损伤远端5 mm处,实验组坐骨神经纤维内NF200呈连续丝状染色,而对照组仍然是颗粒状染色;损伤后4周,在神经肌肉接头处,可观察到实验组肌肉运动终板有新生神经纤维支配,而对照组运动终板上无神经纤维支配,两组ED1及P75染色未见明显差别.结论 锂剂可有效促进周围神经损伤后的再生,但其机制仍需进一步研究.     

Effect of prolonged ischaemic time on muscular atrophy and regenerating nerve fibres in transplantation of the rat hind limb

Abstract

Our aim was to test the influence of cold ischaemia on replanted limbs, focusing on muscular atrophy and neurological recovery. Inbred wild-type and green fluorescent protein (GFP) transgenic (Tg) Lewis rats aged 8–10 weeks were used. The amputated limbs were transplanted at several cold ischaemic times (0, 1, 8, 12, 24, 48, and 72 hours). An arterial ischaemic model and a denervation model were used as controls. To study nerve regeneration, a GFP limb was transplanted on to the syngenic wild Lewis rat. These animals were evaluated histologically, electrophysiologically, and immunohistochemically. The longer the ischaemic time, the more evident was atrophy of the muscles. Electrophysiological investigation showed that the latency at 3 weeks was longer in the transplantation models than in the normal controls, particularly in the longer ischaemia group. Larger numbers of migrating Schwann cells were seen in the group with no delay than in the group that had been preserved for 12 hours. Ischaemia after amputation of a limb causes muscle cells to necrose and atrophy, and these changes worsen in proportion to the ischaemic preservation time. A delay in nerve regeneration and incomplete paralysis caused by malregeneration also affect muscular atrophy.     

Muscle reinnervation with delayed or immediate transplant of embryonic ventral spinal cord cells into adult rat peripheral nerve

Muscle denervation is common in various neuromuscular diseases and after trauma. It induces skeletal muscle atrophy. Only muscle reinnervation leads to functional recovery. In previous studies, denervated adult rat muscles were rescued by transplantation of embryonic day 14-15 (E14-15) ventral spinal cord cells into a nearby peripheral nerve. In the present study, changes were made in the environment into which the cells were placed to test whether reinnervation was improved by: 1) prior nerve degeneration, induced by sciatic nerve transection 1 week before cell transplantation; 2) transplantation of 1 million versus 5 million cells; 3) addition of nerve growth factor (NGF) to the transplant. Ten weeks after cell transplantation, axons had grown from all of the transplants. The numbers of myelinated axons that regenerated into the tibial, medial (MG), and lateral gastrocnemius-soleus (LGS) nerves were similar across treatments. The mean diameters of large LGS axons (>6 microm) were significantly larger with nerve degeneration before transplantation. The mean diameters of MG and LGS axons were significantly larger with transplantation of 1 million versus 5 million cells. Silver-stained experimental and control lateral gastronemius (LG) muscles showed axons that terminated at motor end plates. Nodal and terminal sprouts were more common in reinnervated muscles (45-63% of all end plates) than in control muscles (10%). Electrical stimulation of the transplants induced weak contractions in 39 of 47 MG muscles (83%) and 33 of 46 LG muscles (72%) but at higher voltages than needed to excite control muscles. The threshold for MG contraction was lower with transplantation of 1 million cells, while LG thresholds were lower without NGF. The cross-sectional area of whole LG muscles was significantly larger with cell transplantation (immediate or delayed) than with media alone, but all of these muscle areas were reduced significantly compared with control muscle areas. These data suggest that delayed transplantation of fewer cells without NGF assists regeneration of larger diameter axons and prevents some muscle atrophy.