神经干细胞移植联合促红细胞生成素对大鼠脊髓损伤的修复作用

目的:观察神经干细胞(NSCs)联合促红细胞生成素(EPO)对横断性大鼠脊髓损伤的修复作用,为临床治疗脊髓损伤提供理论依据。方法:40只成年雌性Wistar大鼠建立大鼠T10全横断脊髓损伤模型,并随机分为对照组、NSCs组、EPO组和NSCs+EPO组,每组10只。术后8周采用NF-200免疫组织化学染色和免疫荧光染色对各组大鼠损伤区脊髓神经纤维再生情况进行形态学观察;应用BBB评分评估各组大鼠后肢运动功能恢复情况。结果:组织形态学观察,对照组大鼠横断处脊髓组织残端萎缩,脊髓白质和灰质间可见大量空洞形成,未见有明显的神经纤维再生;NSCs+EPO组大鼠横断处脊髓组织残端轻度萎缩,横断区可见大量呈杂乱、无序生长的神经纤维,可见有连续性神经纤维通过脊髓横断区,脊髓白质和灰质间可见少量空洞形成。NSCs+EPO组大鼠中,FITC共轭抗神经丝蛋白抗体NF-200标记的再生神经纤维在横断区头侧大量再生,呈无序状生长,并通过损伤区到达尾侧;NSCs组大鼠可见少量神经纤维再生,未通过损伤区到达尾侧。NSCs+EPO组大鼠后肢运动功能BBB评分,术后7 d内均高于其他各组(P<0.05);NSCs组大鼠后肢运动功能BBB评分术后7 d内均高于对照组和EPO组(P<0.05)。结论:NSCs移植联合腹腔注射EPO可有效促进大鼠损伤脊髓功能的恢复、轴突的存活和再生。     

神经干细胞移植治疗脊髓横断损伤的研究

目的研究神经干细胞移植治疗对脊髓横断损伤的修复作用.方法采用成年S-D大鼠,随机分为脊髓(胸10)横断损伤对照组(n=7)和神经干细胞移植治疗组(n=7).术后10周,感觉运动区皮质注射10?A示踪剂.动物再存活两周后取出相应的脑和脊髓组织,冰冻切片后,采用自由漂浮法行BDA染色显影,光镜观察.两组实验动物均于脊髓横断术后采用BBB评分法定期评估运动功能.结果神经干细胞移植治疗组所有大鼠BBB运动功能评分在横断损伤4周以后明显高于对照组(P＜O.05).BDA神经束路示踪技术研究显示神经干细胞治疗组7只中的6只大鼠在术后12周有部分BDA阳性标记的皮质脊髓束再生通过脊髓横断损伤部位,而对照组大鼠均未见再生的皮质脊髓束通过.结论在脊髓横断损伤后应用神经干细胞移植治疗,可明显改善损伤的运动功能.神经干细胞移植治疗促进了皮质脊髓束的再生并重新建立了部分神经纤维联系.BDA神经束路示踪技术为脊髓损伤后神经功能的恢复提供了解剖形态学依据.     

神经干细胞与嗅鞘细胞移植对脊髓全横断大鼠后肢运动功能的影响

目的探讨神经干细胞（NSCs）与嗅鞘细胞（OECs）移植对脊髓全横断大鼠后肢运动功能的影响．方法SD大鼠行T11脊髓全横断术后，随机分为NSCs移植组、OECs移植组、联合移植组和对照组．术后将取自绿色荧光蛋白（GFP）转基因胚胎小鼠海马和新生小鼠嗅球的、经培养鉴定后的NSCs和OECs，以明胶海绵做载体分别或联合移植到各移植组动物脊髓损伤处，对照组不移植细胞，用BBB评分法评价1～8周大鼠后肢运动功能恢复的程度．8周后移植各组动物行左心室主动脉内4％多聚甲醛灌注，取横断处上下各约1mL脊髓，连续冰冻切片（片厚20tam），置于荧光显微镜下观察切片中有无绿色荧光细胞，确定NSCs及OECs的存活情况．结果NSCs及OECs细胞在移植脊髓损伤后能存活，并向周围迁移；术后3和4周末，NSCs组、OECs组和联合移植组大鼠运动功能均较对照组有明显改善（P〈0．05）；且3周时，联合移植组和OECs组比NSCs组后肢运动功能变化显著（P〈0．05）；而联合移植组和OECs组比较后肢运动功能未见显著差别（P〉0．05）．绪论NSCs、OECs和联合移植在早期均可促进脊髓全横断大鼠运动功能恢复；OECs组、联合移植组效果均优于NSCs组；联合移植对大鼠运动功能恢复在早期没有显现最佳作用．     

氯化锂联合脐血干细胞移植修复大鼠脊髓损伤

目的 观察氯化锂联合脐血干细胞移植对大鼠脊髓全横断损伤的修复效果.方法 成年雌性SD大鼠80只,建立胸9脊髓全横断损伤模型,随机分对照组(A组,n=20)、氯化锂组(B组,n=20)、细胞移植组(C组,n=20)和氯化锂+细胞移植组(D组,n=20).于术后1 d、3 d及每周的最后1天,应用BBB评分评价大鼠脊髓功能的恢复情况;8周后取材,通过Brdu细胞核标记观察移植细胞的存活、迁移;通过荧光金逆行追踪,观察脊髓神经纤维的再生与分布.结果 4只大鼠死亡.术后8周可观察到Brdu标记的脐血干细胞在体内存活并在脊髓内迁移;细胞移植组和氯化锂+细胞移植组可见少量连续性神经纤维通过损伤区.荧光金逆行脊髓追踪显示C组和D组可见被荧光金标记的神经锥体细胞穿越损伤区.术后8周A、B、C、D四组后肢功能运动BBB评分分别为4.11+0.14、4.50+0.15、8.31±0.11、11.15±0.18.结论 氯化锂能促进人脐血间充质干细胞在损伤区的存活并向神经细胞分化,氯化锂联合脐血干细胞与脊髓去细胞支架移植能够促进细胞移植修复大鼠脊髓损伤的效果.     

骨髓间充质干细胞和嗅黏膜神经干细胞共移植治疗大鼠脊髓损伤

【摘要】目的 探讨骨髓间充质干细胞(BMSCs)和嗅黏膜神经干细胞(OM NSCs)共移植对大鼠脊髓损伤后运动功能的影响。方法 分别提取培养并鉴定大鼠BMSCs和OM NSCs。将60只雄性SPF级SD大鼠随机分为假手术组（仅做椎板切除,不损伤脊髓）,对照组（在脊髓损伤区域注射5 μL生理盐水）,BMSCs移植组（在脊髓损伤区域注射5 μL 5×104个细胞BMSCs单细胞悬液）,OM NSCs移植组（在脊髓损伤区域注射5 μL 5×104个细胞 OM NSCs单细胞悬液,BMSCs和OM NSCs共移植组（在脊髓损伤区域注射5μL 1〖DK〗∶1 5×104个细胞BMSCs和OM NSCs细胞混悬液）。细胞移植后第1、3、7、14、21、28天分别对大鼠进行BBB评分,28天后处死大鼠,分离脊髓组织,进行免疫荧光染色观察细胞迁移情况。结果 造模后,大鼠双后肢瘫痪,BBB评分0分。细胞移植后14天起,与对照组相比,干细胞移植组运动功能有显著改善,BBB评分比较差异具有统计学意义（P＜005）。与BMSCs移植组和OM NSCs移植组相比,移植后21、28天,共移植组BBB评分显著增加（P＜005）。免疫荧光观察结果表明,脊髓损伤28天后,损伤区脊髓内可见空泡形成,两种干细胞聚集在空泡周围,BMSCs在损伤区分布较为集中,而OM NSCs相对散在分布,细胞较为伸展。结论 干细胞移植可以一定程度上恢复脊髓损伤大鼠的运动功能,BMSCs和OM NSCs两种干细胞共移植效果优于单细胞移植。     

胶质细胞源性神经营养因子对脊髓损伤后皮质脊髓束再生的保护作用

目的:观察外源性胶质细胞源性神经营养因子(GDNF)对大鼠损伤脊髓皮质脊髓束再生的影响.方法:利用Nystrm法制备大鼠胸髓压迫损伤模型.蛛网膜下腔置管至损伤局部,连续1周给予GDNF.应用辣根过氧化物酶 (HRP)顺行追踪技术和胶质纤维酸性蛋白(GFAP)、神经中丝(NF)免疫组化活性的变化来评价外源性GDNF对伤区皮质脊髓束和轴突再生的影响.结果:脊髓损伤后4周GDNF治疗组GFAP表达较对照组明显减少,NF标记轴突数显著多于对照组,皮质脊髓束部分再生并通过损伤区至远端0.5 cm.结论:外源性GDNF局部给药能减少胶质细胞增生,促进脊髓损伤大鼠皮质脊髓束轴突再生和神经骨架蛋白的修复.     

神经干细胞移植治疗脊髓损伤的实验研究

目的：观察神经干细胞（NSCs）移植对脊髓损伤（SCI）大鼠功能恢复的作用。方法：30只Wistar大鼠随机分为对照组、损伤组和移植组,每组10只;损伤组和移植组制作成L4平面的脊髓全横断模型,将培养的大鼠NSCs悬液注入移植组损伤脊髓处,损伤组注射等量的生理盐水。术后2mo,采用BBB评分、皮层体感诱发电位（CSEP）和辣根过氧化物酶（HRP）逆行示踪技术观察大鼠脊髓运动和传导功能的恢复程度。结果：术后2mo,BBB评分损伤组、移植组大鼠有所恢复,但都未达到正常水平,其中移植组的大鼠恢复较好,评分较高,与损伤组有统计学差异。SCI后,损伤组、移植组的CSEP波消失,术后2mo移植组的波形有所恢复,但潜伏期延长。对照组脊髓前角可见到许多HRP标记阳性神经元,损伤组未见阳性神经元,移植组可见有阳性神经元,但数目较对照组少。结论：NSCs脊髓内移植能促进损伤脊髓运动和传导功能的部分恢复。     

大鼠脊髓损伤亚急性期NSCs脊髓内移植对脊髓神经电生理的影响

目的 研究神经脊髓损伤的亚急性期(第7天)神经干细胞(neural stem cells,NSCs)脊髓内移植对大鼠受损伤脊髓神经电生理的影响.方法 正常成年SD雌性大鼠15只,分为3组:单纯全横断组、假手术组和神经干细胞移植组.测定3组大鼠的皮层体感诱发电位(cortical somatosensory evoked potential,CSEP)、运动诱发电位(motion evoked potential,MEP)和电针刺激大脑皮质运动区.结果 神经干细胞移植组的诱发电位CSEP、MEP的潜伏期短于单纯脊髓全横断组大鼠,电针刺激大脑皮质运动区发现NSC移植组大鼠双后肢出现明显收缩,且以对侧更为明显.结论 神经干细胞脊髓内移植后能部分促进损伤脊髓的感觉和运动传导功能的恢复.     

大鼠皮质脊髓束选择性横断损伤模型的建立

目的 建立一种稳定可靠且简便实用的皮质脊髓束选择性横断损伤动物模型.方法 选择在延髓锥体切断左侧锥体束,制作大鼠皮质脊髓柬选择性横断损伤模型.采用Rivlin斜板实验评价模型的运动功能,运用Luxolfast blue(LFB)染色法、生物素化葡聚糖胺(BDA)神经示踪法和蛋白激酶Cγ(PKCγ)免疫组织化学染色法观察皮质脊髓束的形态学变化.结果 皮质脊髓束损伤组大鼠术后右侧前后肢瘫痪,运动功能受限,Rivlin试验显示倾斜平面临界角度明显低于对照组,差异有统计学意义(P<0.05);损伤区未见LFB阳性神经纤维束通过;在延髓锥体交叉平面,仅见一束BDA或PKCγ阳性纤维束经右侧锥体交叉至脊髓后索,在左侧脊髓后索最深层,紧邻脊髓灰质后连合背侧下行至骶段,而在损伤平面以下,未见有BDA或PKC γ阳性纤维束经左侧锥体交叉至右侧,在整个脊髓后索的右侧半未见有BDA或PKC γ阳性纤维束.结论 选择在延髓切断一侧锥体束,建立皮质脊髓束选择性横断损伤模型,方法 简便实用,结果 稳定可靠,是研究皮质脊髓束的可塑性和神经轴突再生的理想动物模型.     

神经干细胞移植治疗大鼠脊髓损伤的活体示踪研究

目的:探索利用MR技术活体追踪干细胞的可行性及神经干细胞(NSCs)移植对大鼠脊髓损伤(SCI)后功能恢复的影响。方法:66只SD大鼠随机分为假损伤组(A组)、SCI对照组(B组)和细胞移植治疗组(C组)3组,应用已包被多聚左旋赖氨酸(PLL)的SPIO标记NSCs,对标记细胞进行普鲁士蓝染色,分别在细胞移植后第1、2、3、4、5周对各组动物进行BBB评分,细胞移植后1、3、5周行MRI检查。结果:(1)普鲁士蓝染色证实该方法标记NSCs的有效率为100%。(2)细胞移植后1~5周,B、C组动物运动功能均有不同程度恢复,但B组恢复较慢,BBB评分差异有统计学意义(P<0.05)。(3)1.5 T MRI检查见移植处在T2WI序列呈低信号改变,第3周时低信号向损伤区扩大,第5周时可在损伤区见到低信号改变。结论:MR技术可以活体追踪干细胞,NSCs移植治疗SCI有利于大鼠后肢功能的恢复。     

Human neural stem cells promote corticospinal axons regeneration and synapse reformation in injured spinal cord of rats

Background Axonal regeneration in lesioned mammalian central nervous system is abortive, and this causes permanent disabilities in individuals with spinal cord injuries. This paper studied the action of neural stem cell （NSC） in promoting corticospinal axons regeneration and synapse reformation in rats with injured spinal cord. Methods NSCs were isolated from the cortical tissue of spontaneous aborted human fetuses in accordance with the ethical request. The cells were discarded from the NSC culture to acquire NSC-conditioned medium. Sixty adult Wistar rats were randomly divided into four groups （n=15 in each）： NSC graft, NSC medium, graft control and medium control groups. Microsurgical transection of the spinal cord was performed in all the rats at the T11. The NSC graft group received stereotaxic injections of NSCs suspension into both the spinal cord stumps immediately after transection; graft control group received DMEM injection. In NSC medium group, NSC-conditioned medium was administered into the spinal cord every week; NSC culture medium was administered to the medium control group. Hindlimb motor function was assessed using the BBB Locomotor Rating Scale. Regeneration of biotin dextran amine （BDA） labeled corticospinal tract was assessed. Differentiation of NSCs and the expression of synaptophysin at the distal end of the injured spinal cord were observed under a confocal microscope. Group comparisons of behavioral data were analyzed with ANOVA. Results NSCs transplantation resulted in extensive growth of corticospinal axons and locomotor recovery in adult rats after complete spinal cord transection, the mean BBB scores reached 12.5 in NSC graft group and 2.5 in graft control group （P〈 0.05）. There was also significant difference in BBB score between the NSC medium （11.7） and medium control groups （3.7, P〈 0.05）. BDA traces regenerated fibers sprouted across the lesion site and entered the caudal part of the spinal cord. Synaptophysin expression colocalized with BDA positive axons and neurons distal to the injury site. Transplanted cells were found to migrate into the lesion, but not scatter along the route of axon grows. The cells differentiated into astrocytes or oligodendrocytes, but not into the neurons after transplantation. Furthermore, NSC medium administration did not limit the degree of axon sprouting and functional recovery of the injured rats compared to the NSC graft group. Conclusions Human embryonic neural stem cells can promote functional corticospinal axons regeneration and synapse reformation in the injured spinal cord of rats. The action is mainly through the nutritional effect of the stem cells on the spinal cord.     

Functional recovery following traumatic spinal cord injury mediated by a unique polymer scaffold seeded with neural stem cells and Schwann cells

Background The most important objective of transplant studies in the injured spinal cord has been to provide a favorable environment for axonal growth. Moreover, the continuing discovery of new grafts is providing new potentially interesting transplant candidates. Our purpose was to observe the morphological and functional repair effects of the co-transplantation of neural stem cell （NSC）, Schwann ceils （SCs） and poly lactide-co-glycolide acid （PLGA） on the spinal cord injury of rats.Methods A scaffold of PLGA was fabricated. NSCs and SCs were cultured, with the NSCs labeled with 5-bromodeoxyuridine, and the complex of NSC/PLGA or NSC＋SCs/PLGA were constructed. Thirty-six Wistar rats were randomly divided into three groups： group A （transplantation of PLGA）, group B （transplantation of NSC/PLGA） and group C （transplantation of NSC＋SCs/PLGA）. The 3 mm length of the right hemicord was removed under the microscope in all rats. The PLGA or the complex of PLGA-celIs were implanted into the injury site. Basso-Beattie-Bresnahan （BBB）locomotion scores, motor and somatosensory evoked potential of lower limbs were examined to learn the rehabilitation of sensory and motor function at 4 weeks, 8 weeks, 12 weeks and 24 weeks after injury. All the recovered spinal cord injury （SCI） tissues were observed with HE staining, immunohistochemistry, and transelectronmicroscopy to identify the survival, migration and differentiation of the transplanted cells and the regeneration of neural fibres at 4 weeks, 8 weeks,12 weeks and 24 weeks after injury.Results （1） From 4 weeks to 24 weeks after injury, the BBB locomotion scores of cell-transplanted groups were better than those of the non-cell-transplanted group, especially group C （P 〈0.05）. The amplitudes of the somatosensory evoked potential （SEP） and motor-evoked potential （MEP） were improved after injury in groups B and C, but the amplitude of SEP and MEP at 4 weeks was lower than that at 12 weeks and 24 weeks after injury. Compared with group B, the amplitude of SEP and MEP in group C was improved. The amplitude of SEP and MEP was not improved after injury in group A. （2） HE staining revealed the volume of the scaffold decreased and the number of cells in the scaffold increased. Newly-grown capillaries also could be seen. Immunohistochemistry staining showed the transplanted NSCs could survive and migrate until 24 weeks and they could differentiate into neurons and oligodendrocytes. The regenerated axons were observed in the scaffold-cell complex with transelectronmicroscopy. The above manifestations were more extensive in group C.Conclusions The transplanted NSC can survive and migrate in the spinal cord of rats up to 24 weeks after injury, and they can differentiate into various neural cells. Co-transplantation of cells/PLGA can promote the functional recovery of the injured spinal cord. The effect of co-transplanting NSC＋SCs/PLGA is better than transplanting NSC/PLGA alone.     

嗅鞘细胞移植促进急性脊髓损伤的修复作用

目的研究嗅鞘细胞(OEG)移植后在体内存活情况以及对大鼠脊髓损伤长期的修复作用.方法用新生Wistar大鼠嗅球做OEG培养,大量增殖并标记.80只大鼠,随机分为4组,每组20只.A、B、C组以250 mm·g打击T13脊髓致运动诱发电位和体感诱发电位完全消失,A组在打击区中央用尖刀横行切断脊髓;D组仅做与A、B组相同节段的椎板减压,不挫伤脊髓.A、B组在脊髓远近断端距离挫伤区边缘1 mm的脊髓中线上,深度分别为1.75、1.5、1.0、0.5 mm处,各注入200 000个OEG;C组用同样的方法注射等量的DMEM培养液;D组不做处理.术后观察脊髓功能恢复情况(BBB运动功能评分法)和体重变化.术后24周取脊髓标本,做HE染色、嗜银染色、抗神经原纤维(NF)免疫组化染色、荧光显微镜下观察Hoechst标记的OEG在体内存活情况.结果手术后4周,A、B、C组开始有运动功能恢复,术后4～24周BBB运动功能评分A、B组均高于C组(P＜0.01),16周以后BBB评分变化较小.HE染色A、B两组脊髓损伤区结构紊乱,纤维走行方向扭曲不一致,细胞数目较多,C组大鼠脊髓损伤段常有明显空洞存在,纤维含量少且扭曲.嗜银染色和免疫组化抗NF染色显示无论在损伤区还是损伤头、尾端,A、B组神经纤维数量多于C组,但都少于相同节段的D组(P＜0.01).荧光显微镜下观察到Hoechst标记的OEG大量存在于脊髓损伤段周围.术后各组大鼠均出现体重减轻.术后2周,D组大鼠体重恢复至术前重量,此后逐渐增加.术后4周,A、B、C组大鼠体重降至最低,与D组(Sham组)比较差异有显著性(P＜0.01);此后,A、B组大鼠体重逐渐增加,而C组大鼠体重增加不明显;A、B组大鼠体重与C组比较差异有显著性(P＜0.05).结论OEG移植具有一定促进脊髓损伤神经纤维再生和功能恢复的作用.     

Co-transplantation of neural stem cells and Schwann cells within poly (L-lactic-co-glycolic acid) scaffolds facilitates axonal regeneration in hemisected rat spinal cord

Background  Various tissue engineering strategies have been developed to facilitate axonal regeneration after spinal cord injury. This study aimed to investigate whether neural stem cells (NSCs) could survive in poly(L-lactic-co-glycolic acid) (PLGA) scaffolds and, when cografted with Schwann cells (SCs), could be induced to differentiate towards neurons which form synaptic connection and eventually facilitate axonal regeneration and myelination and motor function.

Methods  NSCs and SCs which were seeded within the directional PLGA scaffolds were implanted in hemisected adult rat spinal cord. Control rats were similarly injured and implanted of scaffolds with or without NSCs. Survival, migration, differentiation, synaptic formation of NSCs, axonal regeneration and myelination and motor function were analyzed. Student’s t test was used to determine differences in surviving percentage of NSCs. One-way analysis of variance (ANOVA) was used to determine the differences in the number of axons myelinated in the scaffolds, the mean latency and amplitude of cortical motor evoked potentials (CMEPs) and Basso, Beattie & Bresnahan locomotor rating scale (BBB) score. The χ² test was used to determine the differences in recovery percentage of CMEPs.

Results  NSCs survived, but the majority migrated into adjacent host cord and died mostly. Survival rate of NSCs with SCs was higher than that of NSCs without SCs ((1.7831±0.0402)% vs. (1.4911±0.0313)%, P <0.001). Cografted with SCs, NSCs were induced to differentiate towards neurons and might form synaptic connection. The mean number of myelinated axons in PLGA+NSCs+SCs group was more than that in PLGA+NSCs group and in PLGA group ((110.25±30.46) vs. (18.25±3.30) and (11.25±5.54), P <0.01). The percentage of CMEPs recovery in PLGA+NSCs+SCs group was higher than in the other groups (84.8% vs. 50.0% and 37.5%, P <0.05). The amplitude of CMEPs in PLGA+NSCs+SCs group was higher than in the other groups ((1452.63±331.70) µV vs. (428.84±193.01) µV and (117.33±14.40) µV, P <0.05). Ipsilateral retransection resulted in disappearance again and functional loss of CMEPs for a few days. But contralateral retransection completely damaged the bilateral motor function.

Conclusions  NSCs can survive in PLGA scaffolds, and SCs promote NSCs to survive and differentiate towards neurons in vivo which even might form synaptic connection. The scaffolds seeded with cells facilitate axonal regeneration and myelination and motor function recovery. But regenerating axons have limited contribution to motor function recovery.

 

     

hTERT基因修饰神经干细胞移植修复大鼠脊髓损伤

目的 对神经干细胞的生物学特性进行观察,探讨hTERT基因修饰神经干细胞移植对修复大鼠脊髓损伤的影响。方法 体外培养的大鼠神经干细胞,用病毒PLXSN为载体介导hTERT基因反转录转染神经干细胞,分为阴性转染组、对照组与hTERT转染组3个组。经Western blot法检测分析hTERT蛋白的表达。运用细胞生长曲线、CCK-8比色法两种方法,分析细胞生长的优化作用。将造模成功的72只大鼠随机分为hTERT-NSCs、NSCs与对照组3组,每组各分24只,通过改良的Allen打击法来建立大鼠急性脊髓损伤模型。通过斜板试验,BBB评分给各组大鼠进行运动功能检测。通过HE染色及荧光显微镜研究,PKH-26标记的分布情况及NSC存活取材进行病理切片。术后72h通过RT-PCR分析脊髓损伤区周围MMP9/2、AQP/9基因的表达,运用UNEL法分析细胞凋亡情况。荧光显微镜观察PKH-26标记的分布情况及NSCs存活。结果 hTERT基因转染大鼠神经干细胞后,hTERT基因转染组蛋白水平有高表达、细胞的生长速度出现明显增快,相比与阴性hTERT转染组与对照组,各组间差异有统计学意义(P<0.05)。对照组的细胞凋亡指数略低于阴性转染组,阴性转染组略低于hTERT转染组大鼠下肢运动功能评价。与对照组相比hTERT转染组AQP4/9基因表达均较显著降低。PKH-26标记的阳性细胞数:对照组最少,hTERT转染组最多,阴性hTERT转染组次之,各组差异有统计学意义(P<0.05)。结论 通过PLXSN病毒为载体介导hTERT基因逆转染神经干细胞,可以促进大鼠神经干细胞增殖。hTERT基因修饰神经干细胞移植可促进脊髓损伤大鼠神经突触的再生,降低脊髓损伤区周围AQP/9、神经细胞凋亡和MMP9/2基因的表达,且能够促进大鼠的电生理及肢体运动功能的恢复。     

应用MRI和电镜观察pSVPoMcat微基因修饰雪旺氏细胞移植对损伤脊髓的修复作用

目的：探讨ｐＳＶＰｏＭｃａｔ微基因修饰雪旺氏细胞（ＳＣ）对损伤脊髓再生修复的作用。方法：采用切割法制备健康ＳＤ大鼠脊髓半横断损伤模型，随机植入ｐＳＶＰｏＭｃａｔ微基因修饰的ＳＣ（Ａ组），高纯化的ＳＣ（Ｂ组），仅植入明胶海绵的损伤对照组（Ｃ组）。受体鼠存活３月，行ＭＲＩ扫描，取材作电镜（ＴＥＭ）观察。结果：ＭＲＩ发现：Ａ组动物，脊髓损伤（ＳＣＩ）区脊髓信号已基本恢复正常，Ｂ组动物未恢复正常，而Ｃ组动     

羊膜间充质干细胞静脉移植治疗脊髓损伤的实验

目的:探讨羊膜间充质干细胞静脉移植对脊髓损伤修复的影响。方法:90只成年SD大鼠按改良Allen′s打击方法建立脊髓损伤模型,随机分为3组:对照组、假移植组和移植组,每组30只。于脊髓损伤后1周经尾静脉移植Brdu标记的羊膜间充质干细胞。3组大鼠分别于损伤后第1 d、3 d、1周及移植后1 d、3 d、1周、2周、4周采用BBB评分法对行为学进行评价。并于移植后1 d、3 d、1周、2周、4周分批处死,切片观察Brdu标记的细胞能否迁移到损伤区。结果:脊髓损伤后第1 d,3组实验动物后肢运动功能BBB评分皆为0分;细胞移植1周后移植组动物BBB评分为5.6±0.6,对照组和假移植组动物BBB评分分别为5.3±0.4和5.2±0.6;细胞移植2周后移植组、假移植组和对照组BBB评分分别为11.3±0.9、6.4±0.5和6.5±0.8;4周后3组实验动物评分分别为13.4±0.9、8.3±0.6和8.5±0.7;在损伤部位出现了Brdu标记的羊膜间充质干细胞。结论:经静脉移植的羊膜间充质干细胞能够迁移到损伤区并对脊髓损伤有一定的修复作用。     

bcl-2基因修饰神经干细胞移植修复大鼠脊髓损伤

目的探讨bcl-2基因修饰神经干细胞移植对脊髓损伤大鼠损伤神经功能恢复的影响。方法体外培养大鼠神经干细胞,经Ad-EGFP为载体介导端B淋巴细胞瘤-2基因(bcl-2)基因转染神经干细胞,分为3组:对照组、阴性转染组、bcl-2转染组。Western-blot检测神经干细胞在转染前后bcl-2蛋白的表达。成年雌性SD大鼠85只,造模成功72只,随机分为对照组,NSCs组,bcl-2-NSCs组,24只/组,按照改良的Allen打击法建立大鼠急性脊髓损伤模型。通过BBB评分、斜板试验进行运动功能评定。造模后7 d通过RT-PCR及Western-blot检测检测脊髓损伤区周围HSP27、c-fos基因的表达,TUNEL法检测细胞凋亡情况。造模后4周取材行病理切片HE染色及荧光显微镜观测EGFP标记的NSC存活及分布情况,通过SEP和MEP观察大鼠神经电生理恢复情况。结果 bcl-2基因转染大鼠神经干细胞后,bcl-2转染组与对照组、阴性转染组相比bcl-2基因和蛋白水平均有表达(P0.05);大鼠下肢运动功能评价bcl-2-NSCs组优于NSCs组,NSCs组优于对照组。造模后72 h,bcl-2-NSCs组细胞凋亡数均明显低于对照组和NSCs组(P0.05)。造模后7 d,与对照组和NSCs组相比,bcl-2-NSCs组HSP27基因和蛋白的表达均较显著升高(P0.05),bcl-2-NSCs组c-fos基因和蛋白的表达较显著降低(P0.05)。造模后4周,HE染色对照组可见脊髓组织缺失及脊髓空洞形成,无神经轴索通过。NSCs组损伤区可见少量神经轴索样结构,脊髓空洞较小,bcl-2-NSCs组可见较多神经轴索样结构,未见脊髓空洞。EGFP标记的阳性细胞数:bcl-2-NSCs组最多,NSCs组次之,对照组未见,且各组之间差异有显著性(P0.05)。造模后4周,SEP和MEP的潜伏期:bcl-2-NSCs组NSCs组对照组,且各组之间差异有显著性(P0.05);波幅:bcl-2-NSCs组NSCs组对照组,且各组之间差异有显著性意义(P0.05)。结论通过Ad-EGFP为载体介导端B淋巴细胞瘤-2基因(bcl-2)基因转染使神经干细胞能够促进体外培养的大鼠神经干细胞增殖。bcl-2基因修饰神经干细胞移植可促进脊髓损伤大鼠神经突触的再生,升高脊髓损伤区HSP27表达,降低脊髓损伤区bcl-2基因的表达和神经细胞凋亡,改善大鼠的肢体运动功能和电生理功能。     

BDNF基因修饰真皮多能干细胞移植对脊髓损伤的修复作用

目的 观察脑源性神经营养因子(BDNF)基因修饰的真皮多能干细胞(dMSCs)移植后对损伤脊髓的修复作用.方法 将72只大鼠采用改良Allen 重物打击法制备脊髓损伤模型,将其随机均分为A组[接受BDNF腺病毒表达载体(Adv-BDNF)感染的dMSCs移植]、B组(接受未感染Adv-BDNF的dMSCs移植)、C组(...