神经干细胞移植对脑损伤大鼠整合素表达的影响

目的探讨神经干细胞（NSCs）移植对大鼠创伤性脑损伤（TBI）整合素（integrin）表达的影响。方法从E14大鼠胚胎分离NSCs,进行原代培养及传代培养;对NSCs进行诱导分化;采用免疫细胞化学技术对NSCs和其分化为神经元的表型进行鉴定。采用改良的Feeney法制备创伤性脑损伤模型。利用脑立体定位仪和微量注射泵进行NSCs脑内移植。采用免疫组织化学技术、免疫印迹技术和RT—PCR技术检测在移植后不同时间脑组织损伤区整合素的表达。结果在培养基中,NSCs呈球团状悬浮生长,Nestin表达阳性。用含10％胎牛血清的培养基对NSCs进行体外诱导分化后第2d,多数细胞伸出突起,以后突起逐渐延长,分支增加。分化后第5d,部分细胞呈βⅢ-微管蛋白阳性。整合素阳性产物主要表达于细胞膜,呈棕黄色。在对照组及移植组均可见阳性细胞表达。在不同时间点,NSCs移植组移植点及其周围脑组织中整合素的mRNA表达均显著高于对照组（P〈O．01）。整合素的蛋白表达结果和tuRNA表达结果相一致。结论移植NSCs至TBI大鼠损伤脑组织,在移植点周围脑组织中整合素的表达显著增加。     

NAAGS基因修饰神经干细胞在创伤性颅脑损伤治疗中的研究

目的：探讨移植NAAG合酶（NAAG synthetase,NAAGS）基因修饰的神经干细胞（Neural Stem Cells,NSCs）能否促进创伤性颅脑损伤大鼠神经功能的恢复。方法：利用电穿孔转染大鼠NSCs,通过脑立体定向仪分别将PBS（模型组）、NSCs（NSCs组）、转基因NSCs（NAAGS＋NSCs组）移植到创伤性颅脑损伤（Traumatic Brain Injury,TBI）大鼠局部损伤灶边缘,通过NSS评分评价移植后大鼠神经功能的变化以及用TUNEL法检测NSCs的凋亡情况,并采用放射免疫法分析脑组织中促炎因子水平。结果：Nss评分结果显示NAAGs＋NSCs组和NSCs组在第7、14、21天神经功能评分均低于模型组（P〈0．05）;NAAGS＋NSCs组在第14和21天神经功能评分低于NSCs组（P〈0．05）;在各时间点细胞移植组比模型组的神经细胞凋亡数明显减少;转基因NSCs移植能明显降低TBI脑组织中促炎因子水平。结论：转基因NSCs移植后可以合成NAAGS促进TBI大鼠神经功能的恢复。     

小鼠脊髓损伤模型的建立及神经干细胞移植后运动功能恢复情况的研究

目的制作小鼠脊髓损伤打击模型,观察神经干细胞(NSCs)移植对脊髓损伤小鼠运动功能恢复及Nestin表达的影响。方法将50只小鼠随机分为空白组(5只)、模型组(15只)、对照组(15只)、治疗组(15只),运用改良Allen's法制备小鼠T10脊髓损伤模型并立即在损伤节段进行NSCs移植,于损伤后1、3、7、14、21d进行BBB评分,并通过免疫荧光法及荧光定量PCR检测Nestin的表达情况。结果所有脊髓打击后小鼠均出现双后肢瘫痪,但随时间延长运动功能可有不同程度恢复,NSCs移植14d后治疗组较模型组及对照组BBB评分显著增高(P0.05),且治疗组Nestin表达量也高于模型组及对照组。结论成功建立了小鼠脊髓损伤打击模型;移植的外源性神经干细胞在脊髓损伤处存活并促进损伤后小鼠运动功能的恢复。     

胚胎神经干细胞移植及胶质细胞源性神经营养因子对大鼠脊髓损伤的修复作用

将胚胎神经干细胞(neural stem cells,NSCs)移植至成年大鼠损伤的脊髓,观察移植后NSCs的存活、迁移以及损伤后的功能恢复。实验结果显示：动物NSCs移植4周后,斜板实验平均角度和运动评分结果比对照组均有明显增高(P＜0．05),而脊髓损伤(spinal cord injury,SCI)处的空洞面积显著减小(P＜0．05);在NSCs中加入胶质细胞源性的神经营养因子(glial cell line-derived neurotrophic factor,GDNF)后,上述改变更加显著。移植后的NSCs不仅能存活,而且向损伤的头端和尾端迁移达3mm之远。这些结果表明,移植的NSCs不仅可以存活、迁移,还可减小SCI空洞面积,促进动物神经功能的恢复;此外,我们的结果还表明GDNF对SCI功能恢复有促进作用。     

神经干细胞移植对AD模型大鼠SNAP-25表达的影响

目的：观察神经干细胞对AD大鼠海马周围微环境中SNAP-25 表达及其认知功能的影响。方法：取成年雄性Wistar大鼠30 只,随机分为对照组、AD模型组、细胞移植组,每组10 只。采用凝聚态Abeta1-42 注射到大鼠海马组织内建立阿尔茨海默病(AD)大 鼠动物模型,通过Y 迷宫测试大鼠学习记忆能力和Western blot技术检测大鼠海马组织内SNAP-25 的表达。结果：Y 迷宫测试结 果显示术后4 周时AD模型组和细胞移植组大鼠学习记忆均低于对照组,与AD模型组比较,细胞移植组大鼠学习记忆能力明显 高于AD模型组,差异有统计学意义（P＜ 0.05）;Western blot 检测结果显示术后4 周时AD模型组和细胞移植组大鼠海马组织内 SNAP-25 蛋白表达量均低于对照组,与AD 模型组比较,细胞移植组大鼠海马组织SNAP-25 蛋白表达量高于AD 模型组差异有 统计学意义（P＜0.05）。结论：移植的NSCs 可改善AD 大鼠的学习和记忆能力,其机制可能是通过改变海马区周围的微环境并上 调了海马组织内SNAP-25 表达。     

人源胚胎神经干细胞移植可对抗大鼠的脑缺血/再灌注损伤

本文旨在研究人源胚胎神经干细胞(human embryonic neural stem cells,h NSCs)移植到脑缺血/再灌注损伤大鼠脑内后的迁移、分化,以及对大鼠脑卒中的疗效。我们在大脑中动脉栓塞(middle cerebral artery occlusion,MCAO)1 h的大鼠模型上,于血流再灌注后第7天注射h NSCs到缺血侧侧脑室,通过焦油紫染色测量大鼠的脑梗死体积,通过检测大鼠的感觉运动行为评估其神经功能的恢复水平,通过免疫荧光共标观察移植后的h NSCs在脑内的迁移与分化。结果显示,h NSCs移植后能够显著减小脑卒中大鼠脑梗死体积,并改善脑卒中大鼠的转棒、错步和转角等运动行为能力;侧脑室注射的h NSCs优先向胼胝体以及梗死区周边迁移,迁移到胼胝体的h NSCs可以分化成少突胶质细胞和星形胶质细胞,迁移到梗死区周边的细胞能够分化成神经元。以上这些结果提示,侧脑室移植的h NSCs可能通过向特定脑区的迁移和分化发挥对脑缺血/再灌注损伤大鼠的保护作用。     

缺氧预处理神经干细胞移植对大鼠急性脊髓损伤后神经胶质细胞凋亡及脊髓空洞形成的影响

目的:研究缺氧预处理神经干细胞(NSCs)移植对大鼠急性脊髓损伤(ASCI)后神经胶质细胞凋亡及脊髓空洞形成的影响。方法:将30只SD大鼠分为假手术对照组;脊髓损伤组;去铁敏组;普通NSCs组;缺氧预处理NSCs组。制成脊髓损伤模型,移植后观察脊髓神经胶质细胞凋亡情况及脊髓空洞形成情况。结果:经去铁敏缺氧预处理培养的NSCs与常规培养的NSCs无明显形态学变化。移植术后7d,缺氧预处理NSCs移植能显著减少脊髓损伤周围区神经胶质细胞的凋亡数量,减少脊髓空洞形成。结论:缺氧预处理NSCs移植能明显抑制大鼠急性脊髓损伤后神经胶质细胞凋亡,减少脊髓空洞的形成。     

缺氧预处理神经干细胞移植对大鼠急性脊髓损伤后神经胶质细胞凋亡及脊髓空洞形成的影响

目的:研究缺氧预处理神经干细胞(NSCs)移植对大鼠急性脊髓损伤(ASCI)后神经胶质细胞凋亡及脊髓空洞形成的影响.方法:将30只SD大鼠分为假手术对照组;脊髓损伤组;去铁敏组;普通NSCs组;缺氧预处理NSCs组.制成脊髓损伤模型,移植后观察脊髓神经胶质细胞凋亡情况及脊髓空洞形成情况.结果:经去铁敏缺氧预处理培养的NSCs与常规培养的NSCs无明显形态学变化.移植术后7d,缺氧预处理NSCs移植能显著减少脊髓损伤周围区神经胶质细胞的凋亡数量,减少脊髓空洞形成.结论:缺氧预处理NSCs移植能明显抑制大鼠急性脊髓损伤后神经胶质细胞凋亡,减少脊髓空洞的形成.     

骨髓间充质干细胞经BDNF基因修饰后移植治疗大鼠半横断脊髓损伤的电生理研究

目的采用电生理的研究方法,观察脑源性神经营养因子(BDNF)基因修饰的骨髓间充质干细胞对脊髓损伤的修复作用。方法随机将大鼠分成3组:空白组10只(只切除椎板,暴露脊髓硬脊膜);SCI组10只;SCI术后细胞移植组10只;从以上三组大鼠随机抽取8只于细胞移植后1 d、7 d、14 d、21 d、30 d、60 d进行SEP(皮层体感诱发电位)、MEP(运动诱发电位)等电生理检测技术,并观察大鼠的运动评分恢复程度。结果细胞移植4d后,大鼠饮食和活动开始增加;后肢变化过程如下:损伤后1~4 d损伤侧后肢迟缓性瘫痪,拖地行走,损伤对侧后肢由损伤初期的运动减弱逐渐恢复,损伤后5~9 d损伤侧后肢痉挛性瘫痪;10~14 d损伤侧下肢恢复少量活动,损伤对侧后肢恢复至较损伤前稍弱的状态;15~21 d损伤侧后肢活动能力较之前有明显改善,至30 d损伤侧后肢活动能力及肌张力恢复程度最明显,30 d以后无更明显改善。免疫组化发现损伤处诱导标记的骨髓间充质干细胞存活,行为学观察发现细胞移植改善了损伤大鼠运动能力。结论骨髓间充质干细胞经BDNF基因修饰后可以促进脊髓损伤大鼠的神经再生及部分传导功能恢复。     

创伤性脑损伤对大鼠海马骨架蛋白及学习记忆功能影响

创伤性脑损伤（traumatic brain injury,TBI）是极为常见的外伤性疾病,致死率和致残率很高。存活者伴随的空间认知功能障碍,给患者家庭和社会造成了极大的负担。目前,对TBI造成的空间记忆障碍缺乏系统研究。脑损伤后海马组织与记忆有关的分子以及组成神经元骨架的分子如何变化研究甚少。本研究采用Wistar大鼠为研究对象,并随机将其分为假手术（sham）组和创伤性脑损伤（TBI）组。TBI组再按致伤后时间长短分为6 h、12 h、24 h、72 h、15 d五个亚组。TBI组应用PinPointTM颅脑撞击器撞击而致伤,sham组不撞击。采用Morris水迷宫评价实验动物空间记忆能力;干湿重法测定脑含水量,评估脑水肿与海马水通道蛋白4（aquaporin-4,AQP-4）的相关性;海马神经元特异性核蛋白（neuron specific nuclear protein,NeuN）标记和免疫荧光检测评估TBI致大鼠神经元丢失情况;通过Western印迹检测TBI致海马骨架相关蛋白质和记忆相关蛋白质含量变化。本研究证实,与sham组相比,TBI组大鼠潜伏期明显增加［（61.98±12.82) s vs.(28.32±8.52) s,n=5,P<0.01,day 15］,探索时间明显缩短［（36.98±0.37) s vs. (73.68±5.09) s,n=5,P<0.01,day15］,表明脑创伤损害了动物的空间参考记忆能力和空间工作记忆能力。与sham组相比,TBI组大鼠海马AQP-4在蛋白质水平上的表达和脑含水量持续升高,15 d恢复正常;在12 h［（3.78±0.74),(83.78±0.35)%］和72 h［（3.49±0.85),(82.28±0.63)%］均形成两个波峰,n=5,P均<0.01,表明继发性脑损伤与持续脑水肿和海马AQP-4在蛋白质上的高表达有关。与sham组相比,NeuN标记和免疫荧光检测发现,TBI后24 h 致大鼠海马神经元丢失严重［（198.2±8.002) vs.(297.2±6.866) cells/mm2, n=5,P<0.01］,表明TBI动物的海马功能受损。与sham相比,TBI组海马神经元树突标志物微管结合蛋白2（microtubule associated proein 2,MAP2）和突触前终末特异性标记物突触素（synaptophysin,SYN）在蛋白质水平均伤后逐步降低（n=5,P均<0.01）,72 h［（0.55±0.05) vs.(1.27±0.08), (0.52±0.14) vs.(1.06±0.16), n=5,P均<0.01］降低最明显;TBI组形成神经元纤维缠结主要成分的过度磷酸化tau（ser404）,伤后逐步升高,72 h［（1.25±0.11)vs. (0.33±0.07), n=5,P<0.01］升高最明显。 MAP2、SYN和过度磷酸化的tau（ser404）检测指标的改变,表明脑损伤致神经元受损,神经元生长和损伤修复能力减弱,最终导致神经元骨架破环,TBI损害了动物的海马空间记忆能力。与sham组相比,TBI组大鼠海马环磷酸腺苷反应元件结合蛋白（cAMP response element binding protein,CREB）和磷酸化CREB ser133(phosphorylated CREB Ser133, pCREB Ser133)含量降低明显（n=5,P均<0.05）,表明脑损伤动物海马的存储记忆能力减弱;TBI组大鼠海马一般调控阻遏蛋白激酶2(general control nonderepressible 2 kinase,GCN2)蛋白质升高明显（n=5,P均<0.05）,表明脑损伤动物海马将新信息转化成长期记忆能力下降。本研究提示,创伤性脑损伤可使大鼠海马神经元骨架破坏,进而导致在学习记忆过程中起重要作用的分子蛋白质下调,抑制记忆储存的蛋白质（GCN2）上调,促使学习记忆功能障碍。     

Neural Stem Cells Over-Expressing Brain-Derived Neurotrophic Factor (BDNF) Stimulate Synaptic Protein Expression and Promote Functional Recovery Following Transplantation in Rat Model of Traumatic Brain Injury

Brain-derived neurotrophic factor (BDNF) plays an essential regulatory role in the survival and differentiation of various neural cell types during brain development and after injury. In this study, we used neural stem cells (NSCs) genetically modified to encode BDNF gene (BDNF/NSCs) and naive NSCs transplantation and found that BDNF/NSCs significantly improved neurological motor function following traumatic brain injury (TBI) on selected behavioral tests. Our data clearly demonstrate that the transplantation of BDNF/NSCs causes overexpression of BDNF in the brains of TBI rats. The number of surviving engrafted cells and the proportion of engrafted cells with a neuronal phenotype were significantly greater in BDNF/NSCs than in naive NSCs-transplanted rats. The expression of pre- and post-synaptic proteins and a regeneration-associated gene in the BDNF/NSCs-transplanted rats was significantly increased compared to that in NSCs-transplanted rats, especially at the early stage of post-transplantation. These data suggest that neurite growth and overexpression of synaptic proteins in BDNF/NSCs-transplanted rats are associated with the overexpression of BDNF, which is hypothesized to be one of the mechanisms underlying the improved functional recovery in motor behavior at the early stage of cell transplantation following TBI. Therefore, the protective effect of the BDNF-modified NSCs transplantation is greater than that of the naive NSCs transplantation.     

神经干细胞在治疗脑损伤中的应用

神经干细胞（neural stem cells,NSCs）是中枢神经系统中既具有自我更新能力又能分化为神经系统各类细胞的细胞群。在体外一定条件下,NSCs能保持增殖能力,经定向诱导能分化为具有成熟神经细胞特征的各类细胞。NSCs移植治疗研究显示,植入的NSCs能分化为移植部位的神经细胞,并融入、整合该部位,重建受损神经网络,在一定程度上缓解病症。近年来,激活体内内源NSCs治疗神经损伤也逐渐得到广泛关注。因此,NSCs在治疗神经损伤中的应用研究已成为当前神经生物学基础理论和临床应用研究的热点。本文简要介绍了最近关于NSCs在治疗脑损伤中的应用研究进展。     

Ghrelin attenuates brain injury after traumatic brain injury and uncontrolled hemorrhagic shock in rats

Traumatic brain injury (TBI) and hemorrhagic shock often occur concomitantly due to multiple injuries. Gastrointestinal dysfunction occurs frequently in patients with TBI. However, whether alterations in the gastrointestinal system are involved in modulating neuronal damage and recovery after TBI is largely neglected. Ghrelin is a "gut-brain" hormone with multiple functions including antiinflammation and antiapoptosis. The purpose of this study was to determine whether ghrelin attenuates brain injury in a rat model of TBI and uncontrolled hemorrhage (UH). To study this, brain injury was induced by dropping a 450-g weight from 1.5 m onto a steel helmet attached to the skull of male adult rats. Immediately after TBI, a midline laparotomy was performed and both lumbar veins were isolated and severed at the junction with the vena cava. At 45 min after TBI/UH, ghrelin (4, 8 or 16 nmol/rat) or 1 mL normal saline (vehicle) was intravenously administered. Brain levels of TNF-α and IL-6, and cleaved PARP-1 levels in the cortex were measured at 4 h after TBI/UH. Beam balance test, forelimb placing test and hindlimb placing test were used to assess sensorimotor and reflex function. In additional groups of animals, ghrelin (16 nmol/rat) or vehicle was subcutaneously (s.c.) administered daily for 10 d after TBI/UH. The animals were monitored for 28 d to record body weight changes, neurological severity scale and survival. Our results showed that ghrelin downregulated brain levels of TNF-α and IL-6, reduced cortical levels of cleaved PARP-1, improved sensorimotor and reflex functions, and decreased mortality after TBI/UH. Thus, ghrelin has a great potential to be further developed as an effective resuscitation approach for the trauma victims with brain injury and severe blood loss.     

The Effects of Harvesting Media on Biological Characteristics and Repair Potential of Neural Stem Cells after Traumatic Brain Injury

Various solutions are utilized widely for the isolation, harvesting, sorting, testing and transplantation of neural stem cells (NSCs), whereas the effects of harvesting media on the biological characteristics and repair potential of NSCs remain unclear. To examine some of these effects, NSCs were isolated from cortex of E14.5 mice and exposed to the conventional harvesting media [0.9% saline (Saline), phosphate-buffered saline (PBS) or artificial cerebrospinal fluid (ACSF)] or the proliferation culture medium (PCM) for different durations at 4°C. Treated NSCs were grafted by in situ injection into the lesion sites of traumatic brain injury (TBI) mice. In vitro, harvesting media-exposed NSCs displayed time-dependent reduction of viability and proliferation. S phase entry decreased in harvesting media-exposed cells, which was associated with upregulation of p53 protein and downregulation of cyclin E1 protein. Moreover, harvesting media exposure induced the necrosis and apoptosis of NSCs. The levels of Fas-L, cleaved caspase 3 and 8 were increased, which suggests that the death receptor signaling pathway is involved in the apoptosis of NSCs. In addition, exposure to Saline did not facilitate the neuronal differentiation of NSCs, suggesting that Saline exposure may be disadvantageous for neurogenesis. In vivo, NSC-mediated functional recovery in harvesting media-exposed NSC groups was notably attenuated in comparison with the PCM-exposed NSC group. In conclusion, harvesting media exposure modulates the biological characteristics and repair potential of NSCs after TBI. Our results suggest that insight of the effects of harvesting media exposure on NSCs is critical for developing strategies to assure the successful long-term engraftment of NSCs.     

Binge Ethanol Prior to Traumatic Brain Injury Worsens Sensorimotor Functional Recovery in Rats

A significant number of patients suffering from traumatic brain injury (TBI) have a high blood alcohol level at the time of injury. Furthermore, drinking alcohol in a binge-like pattern is now recognized as a national problem, leading to a greater likelihood of being injured. Our objective was to determine the consequences of a binge paradigm of alcohol intoxication at the time of TBI on long-term functional outcome using a sensitive test of sensorimotor function. We trained adult, male, Sprague Dawley rats on the skilled forelimb reaching task and then administered a single binge dose of ethanol (2g/kg, i.p.) or saline for three consecutive days (for a total of 3 doses). One hour after the final ethanol dose, rats underwent a TBI to the sensorimotor cortex corresponding to the preferred reaching forelimb. Animals were then tested for seven weeks on the skilled forelimb reaching task to assess the profile of recovery. We found that the group given ethanol prior to TBI displayed a slower recovery curve with a lower recovery plateau as compared to the control group. Therefore, even a relatively short (3 day) episode of binge alcohol exposure can negatively impact long-term recovery from a TBI, underscoring this significant public health problem.     

Pathway-Focused PCR Array Profiling of Enriched Populations of Laser Capture Microdissected Hippocampal Cells after Traumatic Brain Injury

Cognitive deficits in survivors of traumatic brain injury (TBI) are associated with irreversible neurodegeneration in brain regions such as the hippocampus. Comparative gene expression analysis of dying and surviving neurons could provide insight into potential therapeutic targets. We used two pathway-specific PCR arrays (RT2 Profiler Apoptosis and Neurotrophins & Receptors PCR arrays) to identify and validate TBI-induced gene expression in dying (Fluoro-Jade-positive) or surviving (Fluoro-Jade- negative) pyramidal neurons obtained by laser capture microdissection (LCM). In the Apoptosis PCR array, dying neurons showed significant increases in expression of genes associated with cell death, inflammation, and endoplasmic reticulum (ER) stress compared with adjacent, surviving neurons. Pro-survival genes with pleiotropic functions were also significantly increased in dying neurons compared to surviving neurons, suggesting that even irreversibly injured neurons are able to mount a protective response. In the Neurotrophins & Receptors PCR array, which consists of genes that are normally expected to be expressed in both groups of hippocampal neurons, only a few genes were expressed at significantly different levels between dying and surviving neurons. Immunohistochemical analysis of selected, differentially expressed proteins supported the gene expression data. This is the first demonstration of pathway-focused PCR array profiling of identified populations of dying and surviving neurons in the brain after TBI. Combining precise laser microdissection of identifiable cells with pathway-focused PCR array analysis is a practical, low-cost alternative to microarrays that provided insight into neuroprotective signals that could be therapeutically targeted to ameliorate TBI-induced neurodegeneration.     

Cultured Rat Astrocytes Give Rise to Neural Stem Cells

Previously, we reported the occurrence of neural stem cells (NSCs) around an area of damage after rat traumatic brain injury (TBI), but it was unclear if this was due to blastgenesis in astrocytes, or to NSCs migrating from the subventricular zone (SVZ). In this study, NSCs were isolated and cultured from cultured type 1 astrocytes taken from newborn rat cortex in which the subventricular zone and hippocampus had been discarded. All cultured type 1 astrocytes showed glial fibrillary acidic protein (GFAP) immunopositivity. Nestin immunopositive spheres were isolated from type 1 astrocytes and cultured in the presence of bFGF and EGF in the medium. Neurospheres differentiated into Tuj1-, GFAP- and A2B5-positive cells after 4 days of culture without bFGF and EGF. These results indicate that isolated neurospheres from brain cortex astrocytes can differentiate into neurons and glia and might contribute to neurogenesis and neuroplasticity.     

急性酒精中毒合并中度创伤性脑损伤大鼠海马AQP4的表达

目的:探讨大鼠急性酒精中毒合并颅脑外伤后AQP4在海马区表达的变化.方法:健康成年雄性SD大鼠96只,随机分为4组:假手术组(N组)、急性酒精中毒组(A组)、中度创伤性脑损伤组(T组)和急性酒精中毒合并中度创伤性脑损伤(AT组).腹腔注射酒精(2.5g/kg),2h后以重物自由落体击打大鼠头部建立急性酒精中毒合并中度创伤性脑损伤(traumatic brain injury,TBI)动物模型.各组动物分别存活1、3、5、14天.免疫组化方法检测海马CA1区AQP4的表达.结果:AQP4阳性产物分布于胶质纤维和毛细血管壁,各实验组表达均高于N组.术后1天T组比AT组表达显著增高(P＜0.01),术后3天AT组比T组表达增高(P＜0.05),术后14天AT组比T组表达显著增高(P＜0.01).结论:大鼠急性酒精中毒合并颅脑外伤后晚期,海马CA1区AQP4表达增高,可能加重晚期继发性脑水肿,是急性酒精中毒合并颅脑外伤预后不良的原因之一.     

Neural stem cells transplanted into intact brains as neurospheres form solid grafts composed of neurons, astrocytes and oligodendrocyte precursors

Neural stem cells (NSCs) are tissue-specific stem cells with self-renewal potential that can give rise to neurons and glia in vivo and in vitro. The aim of this study was to transplant NSCs as whole neurospheres into intact brain and assess the fate and phenotype of their progeny generated in vivo. We isolated NSCs from E14 foetal rat forebrains and cultured them in basic fibroblast and epidermal growth factor-supplemented serum-free medium in the form of neurospheres in vitro. Neurospheres were transplanted into the intact brains of 2 Wistar rats and after a period of 3 weeks, grafted brains were examined immunohistochemically. Neurospheres formed solid grafts that were found in the lateral ventricle and in the velum interpositum under the hippocampus. The majority of cells in the transplanted tissue were identified as beta-III-tubulin(+), NeuN(+), PanNF(+) and synaptophysin(+) neurons and were accumulated throughout the graft centre. GFAP(+) astrocytes were scattered throughout the entire graft and astrocyte processes delimited the outer and perivascular surfaces. A great number of NG2(+) oligodendrocyte precursors was detected. Nestin(+) endothelial cells were found to line capillaries growing in the transplant. These data indicate that nestin(+) NSCs prevailing in neurospheres differentiate following transplantation into nestin(-) neuronal and glial cells which confirms the multipotency of NSCs. Three weeks posttransplantation neuronal and astrocyte cells reached terminal differentiation (formation of synaptic vesicles and superficial and perivascular limiting membranes) while elements of oligodendroglial cell lineage remained immature. Grafting stem cells as non-dissociated neurospheres provide cells with favourable conditions which facilitate cell survival, proliferation and differentiation. However, in the intact brain, grafted neurosphere cells were not found to integrate with the brain parenchyma and formed a compact structure demarcated from its surroundings.