目的 探讨移植神经干细胞对脊髓全横断性大鼠部分结构与功能修复的影响。方法 在脊髓全横断处移植神经干细胞60d 后，在横断处下方3mm注射荧光金逆行标记轴突再生的上运动神经元，用免疫组织化学检测神经干细胞在宿主内的分化，同时用体视学方法观测脑干红核和大脑皮质感觉运动区内锥体细胞层的神经元密度变化。用BBB 评分法和爬网格法观测大鼠后肢运动功能的恢复等。结果 移植的神经干细胞能在宿主内存活并向前后方向迁移到脊髓内，部分神经干细胞分化为GFAP、NF2200和GAP243 阳性细胞。移植神经干细胞后在红核和感觉运动区内锥体细胞层可见有被荧光金标记的神经元胞体，脊髓横断处附近脊髓组织的溃变程度减轻，红核及躯体感觉运动区内神经元密度高于未移植组，大鼠后肢的自主运动功能明显好于未移植组。结论 神经干细胞移植入损伤脊髓后能分化为神经元及神经胶质细胞，能减轻脊髓的继发性损伤，保护受损伤的神经元，促进运动功能的恢复。

Spinal cord transection results in severe neurological sequelae, and to date, there is no effective treatment. Because of the limited capacity for axonal regeneration in the spinal cord, recovery is minimal. Recently, efforts have been made to establish, by grafting neural tissue, a functional relay-station between the severed stumps of the injured cord. Previously, we used co-transplantation of neural stem cells (NSCs) and Schwann cells (SCs) to improve functional recovery of transected spinal cord. However, this effort has been partially impeded by limited neuronal differentiation of transplanted NSCs. To circumvent this problem, we have pre-differentiated NSCs toward neurons in vitro with the application of retinoic acid (RA) prior to cell grafting. Further, we genetically modifiedSCs to overexpress human neurotrophin-3 (hNT-3). When these cells were co-transplanted into the transected spinal cord of rats, injured animals had partial improvement (both functionally and structurally), including improved Basso, Beattie, and Bresnahan (BBB) scores, increased axonal regeneration/remyelination, and reduced neuronal loss. However, this pre-differentiation of NSCs in vitro only mildly improved neuronal differentiation of NSCs in vivo.

目的 探讨胶原或明胶吸附施万细胞移植对全横断脊髓结构和功能修复的影响。方法 将胶原或明胶吸附施万细胞移植到成年大鼠全横断脊髓的损伤处，术后3个月用爬网格方式测试动物后肢自主运动功能恢复情况；用荧光金逆行标记法观察大脑感觉运动区和脑干红核的神经元神经纤维再生；用免疫组织化学法检测脊髓CGRP和52HT能神经纤维再生。结果 移植施万细胞的大鼠后肢自主运动功能有显著的恢复。大脑感觉运动区和脑干红核均有被荧光金标记的神经元胞体，提示两个区域的神经元轴突能在脊髓再生并穿越损伤区到达尾侧脊髓。脊髓损伤处有CGRP 和52HT阳性神经纤维，以及损伤处尾侧有52HT阳性神经纤维。对照组大鼠以上结果均为阴性。结论 施万细胞移植可促进大鼠全横断脊髓结构和功能的恢复。胶原或明胶可作为移植细胞的网架用于细胞治疗。

This study investigated whether electro-acupuncture (EA) would improvethe survival and migration of neural stem cells (NSCs) transplanted ininjured spinal cord as well as the potential mechanisms. TlO spinal cordsegments of 50 adult Sprague-Dawley (SD) rats were completely transected,and then NSCs were immediately transplanted into the transected site of theexperimental animals, while control animals were sham operated withouttransplantation. Five days post-operation, electro-acupuncture treatment onGV9 (Zhiyang), GV6 (Jizhong), GV2 (Yaoshu) and GVI (Changqiang)acupoints was applied for 14 days (EA+NSCs 14d) and 30 days (EA+NSCs30d). ELISA and immunohistochemical staining were used to assess thecontent of neurotrophine-3 (NT-3) and the characteristics of transplantedNSCs. We found that the number of transplanted NSCs the survived inEA+NSCs14d group was significantly increased as compared to that of theNSCs30d group (5825.20+819.01vs 4781.40+500.49, P<0.05).19Immunostaining indicated that some transplanted NSCs developed intomicrotubule association protein 2 (MAP2) positive cells and many of themdeveloped into glial fibrillary acidic protein (GFAP) positive cells in theNSCs30d group. Further, the migration length of transplanted NSCs towardcaudal tissue in the injured site was longer in the EA+NSCs30d group thanthat in NSCs30d group (5.98+0.79mm vs 3.96+1.72mm; P<0.05). AlsoNT-3 in injured spinal cord tissue was 23% increased in the EA+NSCs14dgroup. These results suggest that the combination of EA and NSCs improvesthe survival and migration of NSCs in injured spinal cord in rats.

AbstractWe have constructed a recombinant adenovirus expression vector carrying the human neurotrophin-3 (NT-3) receptor TrkC (tyrosine proteinkinase C) gene (rAd-TrkC; 2478 bp) and confirmed the expression of the encoded TrkC in green fluorescent protein (GFP)-murine neural stem cells (NSCs) by reverse transcription polymerase chain reaction (RT-PCR),Western blot analysis, andimmunocytochemistry. The activity of the expressedrAd-TrkC was verified in vitro by evaluating dose-related responses of NSCs to NT-3, a TrkC specific ligand. TrkC-GFP-NSCs had a significantly higher percentage of neuronal differentiation when treated with NT-3 relative to the rAd-LacZ control cells (55.2% vs. 29.8%; P<0.05, χ2 test).Thus, our rAd-TrkC vector can transfect NSCs and produce functional TrkC receptors to promote neuronal differentiation of NSCs.