利用克隆羊进行各类神经干细胞移植治疗脑挫裂伤的研究
|
摘要
中枢神经系统神经组织的再生是困扰医学界的难题之一，神经干细胞(Neural Stem Cell，NSC)移植治疗是目前一种新兴的，具有良好应用前景的治疗方法，在神经科学领域的许多疾病当中已经有很多研究证实了其移植治疗的有效性。脑外伤是神经外科临床工作中最常见的一大类疾病，危害极大，治疗价值最高。虽然不同种类的脑外伤有各自不同的病理生理过程，但有相当一部分的脑外伤都伴有或轻或重的脑挫裂伤，故本实验就专门针对脑挫裂伤进行研究。
NSC依据其来源分为胚胎源性神经干细胞(embryonic-derived neural stem cells，EdNSC)和成体源性神经干细胞(adult derived neural stem cells，AdNSC)两大类。这两类神经干细胞细胞移植治疗脑挫裂伤时哪个效果更好?不同来源的各类神经干细胞移植治疗中枢神经系统损伤时是否发生免疫排斥反应?能否建立一种可控性、重复性好，观察指标明确、操作简单确实可靠的大动物脑挫裂伤模型?对于这些问题，目前尚无系统性的研究。本实验充分利用了克隆羊这一遗传背景清楚的大动物作为模型，同时利用克隆技术得到来源相同的“自体”EdNSC和AdNSC及异体AdNSC，来解决上述问题。
本实验的第一部分首先对成体克隆羊(编号：CL-4)及成体非克隆羊(编号：SP-3119)通过开颅手术的方法取得其皮层脑组织，进而在体外培养获得克隆羊及非克隆羊的AdNSC；然后对CL-4克隆羊通过体细胞核移植(somatic cell nuclear transfer，SCNT)技术获得其克隆胚胎羊(此步骤由本课题的合作单位完成)，当胚胎羊在“代理母亲”体内长至35天时，采用外科手术获取其脑组织，继而在体外培养得到EdNSC。这样对于与CL-4同一批次的克隆羊来说，所得到的EdNSC及克隆羊的AdNSC排除表观遗传学因素，可认为是“自体NSC”；而非克隆羊的AdNSC是“异体NSC”。我们对所得到的这三类NSC通过Nestin鉴定后，用绿色荧光蛋白(GFP)进行标记以用作移植治疗。另外在NSC培养过程中发现EdNSC体外培养速度比AdNSC快。
在本实验的第二部分中，我们随机选取与CL-4同一批次的CL系列克隆羊20只。通过外科手术开颅暴露出克隆羊大脑上薛氏沟周围的运动皮层区，然后使用50g×100cm力度作自由落体加速撞击，建立起观察指标明确的克隆羊脑挫裂伤模型。
第三部分中，我们将已建立模型的动物随机分成四组，分别用第一部分所得到的三类NSC：克隆羊的EdNSC、AdNSC和非克隆羊的AdNSC以及生理盐水进行移植治疗。观察比较移植治疗后各组动物的存活率；行为学恢复情况；脑损伤对侧肢体运动功能恢复情况及运动诱发电位的恢复情况；并测定各组动物头颅MRI影像学及病理显微观的改变；同时比较移植到脑内的各类神经干细胞在动物脑内的存活、迁移、分化情况。结果发现：克隆羊EdNSC、AdNSC移植组与非克隆羊AdNSC移植组及对照组相比，动物的存活率、行为学恢复情况及脑损伤灶对侧肢体运动功能恢复情况及运动诱发电位的恢复情况明显提高；头颅MRI影像学及损伤区域病理显微观表现明显改善；移植到脑内的神经干细胞均可良好存活、迁移、分化；这两组比较而言，EdNSC的移植治疗效果相对更好一些，表现在脑损伤灶对侧肢体运动功能恢复更快；影像学及病理显微观显示脑挫裂伤灶吸收、恢复更快。而非克隆羊AdNSC移植组与对照组相比，移植治疗效果差别不大。
在实验的第四部分中，我们专门对各类NSC移植治疗中枢神经系统损伤后的免疫排斥反应问题进行了研究。实验中我们检测了细胞移植前后不同时间点(造模前1天、细胞移植前、细胞移植后3天、1个月及3个月时)各组动物血液中细胞因子(IL-2，IL-6，IL-10)水平，以及细胞移植后急性期(细胞移植后1周)和慢性期(细胞移植后3个月)，移植局部病理切片中CD3+细胞浸润的免疫组化染色情况，力争从全身和移植局部两个方面探讨各类神经干细胞移植后免疫排斥反应这一问题。结果发现：非克隆羊AdNSC组移植后3天、1个月、3个月时IL-2水平较其它组明显升高(P＜0.05)；细胞移植3个月时该组动物血中IL-6水平较其它组明显升高(P＜0.05)；而在移植后3天、1个月、3个月时该组动物血中IL-10水平较其它组明显下降(P＜0.05)。而克隆羊EdNSC及AdNSC移植组之间及它们与对照组相比，各时间点动物血液中细胞因子(IL-2，IL-6，IL-10)水平均没有统计学差异(P＞0.05)。非克隆羊AdNSC(异体细胞移植组)在细胞移植后的急性期(移植后1周)和慢性期(移植后3个月)，移植局部CD3+细胞的数量明显比其它组多【与其它组相比有统计学差异(P＜0.05)】，说明该组移植部位发生了免疫排斥反应。而两个“自体”细胞移植组(克隆羊EdNSC和AdNSC移植组)，移植局部CD3+细胞的数量与对照组相比无明显的增多，这说明这两个“自体”神经干细胞移植组局部未发生明显免疫排斥反应。这两个“自体”细胞移植组之间相比，可以看到EdNSC组CD3+细胞的数量稍多，但二者之间没有统计学差异(P＞0.05)。
综上，通过本实验我们得出了以下结论：1对胚胎克隆羊、成体克隆及非克隆羊通过外科手术取其皮层脑组织，在体外培养均能得到生长良好的胚胎神经干细胞(EdNSC)及成体神经干细胞(AdNSC)。在相同的条件下，EdNSC体外培养速度比成AdNSC快。2对羊脑上薛氏沟周围运动皮层区使用50g×100 cm力度作自由落体加速撞击，可产生观察指标明确且存活率高的羊脑挫裂伤模型，有望广泛用于神经干细胞移植及药物等治疗的实验研究。3胚胎神经干细胞(EdNSC)及成体神经干细胞(AdNSC)移植治疗脑挫裂伤后均有较明显的治疗效果；二者相比较而言胚胎神经干细胞(EdNSC)稍好于成体神经干细胞(AdNSC)。4中枢神经系统异体神经干细胞移植存在着明显的免疫排斥反应，而自体神经干细胞移植基本无免疫排斥反应
The regeneration of nervous tissue in central nervous system is a tough problemin medical science. Neural Stem Cell(NSC)transplantation serves as a newly emergingand a good perspective therapeutic method, has been confirmed in many diseasetherapy in neuroscience domain. Brain trauma is the most frequent disease in clinicalneurosurgery and has largest harm to mankind. It has the topmost therapeutic value.Even though different brain trauma has different pathological process, but most ofthem have the brain contusion and laceration. So We researched the brain contusionand laceration restrictly in our study.
Based on its resource, NSC can be divided into two groups: embryonic derivedneural stem cell(EdNSC)and adult derived neural stem cell(AdNSC). Whosetherapeutic effect on brain contusion and laceration is better? Do immunologicalrejection generate When various kinds of NSC transplanted into brain of animal? Canestablished a brain contusion and laceration model in big animals that have highsurvival rate and can be detected clearly? To these problems, there has no convinceresearch now. In our study, We utilize the cloning goats Whose heredity backgroundare very clearly as the models to estabilsh the brain contusion and laceration ofcloning goats, and investigate the immunological rejection of EdNSC and AdNSCtransplantation.
In the first part of our study, We obtained the cerebral cortex of adult cloninggoat(NO:CL-4) and non-cloning goat(NO:SP-3119) by surgical intervention. Weobtained the AdNSC of them by cell culture in vitro. Then We obtained the cerebralcortex of the fetal cloning goat(35days)Which gained by somatic cell nucleartransfer(SCNT) of CL-4 cloning goat and obtained the EdNSC by cell culture in vitro.To the CL series cloning goats,the EdNSC and AdNSC of cloning goat could be deemas "autoallergic NSC", and AdNSC of non-cloning goat was alloallergic NSC.On theother hand, in the cell culture process, the EdNSC growed faster than AdNSC in thesame cell culture method.
In the second part of our study, We chosed 20 CL series cloning goats at random.After exposed the upper Sylvii sulcus moter cortex of brain of the cloning goats bysurgical intervention, 50 gram of steel ball was used to produce a free fallingaccelerate strike on the brain from 100cm height respectively and established thebrain contusion and laceration goat models that had high survival rate and could be detected clearly.
In the third part of our study, We divided the cloning goat models into 4 groups.We transplanted the EdNSC, AdNSC of cloning goat and AdNSC of non-cloning goatand saline into the around areas of brain contusion and laceration of each groupmodels respectively. We observed each group models' survival rate, ethologicalrecovery; The Motion Evoked Potential(MEP), brain Magnetic ResonanceImaging(MRI) image, pathology changes of brain contusion and laceration in eachgroup models were also recorded. We then observed the survival, immigration anddifferentiation of all kinds of stem cells in goats' brains. We found: all kinds of stemcells can survival, immigrate and different well in the transplanted brain. The EdNSCand AdNSC of cloning goat transplantation groups had good survival rate, ethologyrecovery. Their MEP recovery of the motor cortex, brain MRI image and pathologychanges of brain contusion and laceration were better than AdNSC of non-cloninggoat transplantation group and control group; The EdNSC transplantation group hadthe best therapeutic effect on brain contusion and laceration in all transplantationgroups and control group.
In the fourth part of our study, We investigated the acute(1 week after celltransplantation) and chronic(3 months after cell transplantation) immunologicalrejection of each group of the models Who received various transplanted stem cells.We detected the various cytokine(IL-2,IL-6 and IL-10) in each group models' blood atdifferent time (1 day before model established; before, 3 days, 1 month and 3 monthsafter cell transplantation).We also detected CD3+ cells in the cell transplantation areasaround brain contusion and laceration in all groups goats by the method ofimmunohistochemistry. We found:in the AdNSC of non-cloning transplantationgroup,the level of IL-2 was obviously high and the level of IL- 10 was obviously lowthan other groups at 3 days, 1 month, and 3 months after celltransplantation(P＜0.05).The level of IL-6 was obviously high than other groups at 3months after cell transplantation(P＜0.05). The level of these cytokines in the EdNSC,AdNSC of cloning goat transplantation groups and control group had no statisticsdifference(P＞0.05) at every time. The quantity of CD3+ cells in the AdNSC ofnon-cloning goat group was much more than that of other groups(P＜0.05) at the acuteperiod(1 week after cell transplantation) and chronic period(3 months after celltransplantation). The quantity of CD3+ cells in the EdNSC, AdNSC of cloning goattransplantation group and control group had no statistics difference(P＞0.05) at acute and chronic period.
Overall, in our study, We got the conclusions as follow: 1,The EdNSC, AdNSCof cloning goat and the AdNSC of the non-cloning goat were successful obtained;EdNSC growed faster than AdNSC in the same cell culture method. 2,The braincontusion and laceration model of cloning goats were successful established by freefalling of 50 gram of steel ball accelerate strike on the goat brain from 100cm height.3, EdNSC and AdNSC transplantation both had obvious therapeutic effect on braincontusion and laceration of cloning goats; The therapeutic effect of EdNSCtransplantation was better. 4, Allograft of stem cells has acute and chronicimmunological rejection and autografting of stem cells has no acute and chronicimmunological rejection.
NSC依据其来源分为胚胎源性神经干细胞(embryonic-derived neural stem cells，EdNSC)和成体源性神经干细胞(adult derived neural stem cells，AdNSC)两大类。这两类神经干细胞细胞移植治疗脑挫裂伤时哪个效果更好?不同来源的各类神经干细胞移植治疗中枢神经系统损伤时是否发生免疫排斥反应?能否建立一种可控性、重复性好，观察指标明确、操作简单确实可靠的大动物脑挫裂伤模型?对于这些问题，目前尚无系统性的研究。本实验充分利用了克隆羊这一遗传背景清楚的大动物作为模型，同时利用克隆技术得到来源相同的“自体”EdNSC和AdNSC及异体AdNSC，来解决上述问题。
本实验的第一部分首先对成体克隆羊(编号：CL-4)及成体非克隆羊(编号：SP-3119)通过开颅手术的方法取得其皮层脑组织，进而在体外培养获得克隆羊及非克隆羊的AdNSC；然后对CL-4克隆羊通过体细胞核移植(somatic cell nuclear transfer，SCNT)技术获得其克隆胚胎羊(此步骤由本课题的合作单位完成)，当胚胎羊在“代理母亲”体内长至35天时，采用外科手术获取其脑组织，继而在体外培养得到EdNSC。这样对于与CL-4同一批次的克隆羊来说，所得到的EdNSC及克隆羊的AdNSC排除表观遗传学因素，可认为是“自体NSC”；而非克隆羊的AdNSC是“异体NSC”。我们对所得到的这三类NSC通过Nestin鉴定后，用绿色荧光蛋白(GFP)进行标记以用作移植治疗。另外在NSC培养过程中发现EdNSC体外培养速度比AdNSC快。
在本实验的第二部分中，我们随机选取与CL-4同一批次的CL系列克隆羊20只。通过外科手术开颅暴露出克隆羊大脑上薛氏沟周围的运动皮层区，然后使用50g×100cm力度作自由落体加速撞击，建立起观察指标明确的克隆羊脑挫裂伤模型。
第三部分中，我们将已建立模型的动物随机分成四组，分别用第一部分所得到的三类NSC：克隆羊的EdNSC、AdNSC和非克隆羊的AdNSC以及生理盐水进行移植治疗。观察比较移植治疗后各组动物的存活率；行为学恢复情况；脑损伤对侧肢体运动功能恢复情况及运动诱发电位的恢复情况；并测定各组动物头颅MRI影像学及病理显微观的改变；同时比较移植到脑内的各类神经干细胞在动物脑内的存活、迁移、分化情况。结果发现：克隆羊EdNSC、AdNSC移植组与非克隆羊AdNSC移植组及对照组相比，动物的存活率、行为学恢复情况及脑损伤灶对侧肢体运动功能恢复情况及运动诱发电位的恢复情况明显提高；头颅MRI影像学及损伤区域病理显微观表现明显改善；移植到脑内的神经干细胞均可良好存活、迁移、分化；这两组比较而言，EdNSC的移植治疗效果相对更好一些，表现在脑损伤灶对侧肢体运动功能恢复更快；影像学及病理显微观显示脑挫裂伤灶吸收、恢复更快。而非克隆羊AdNSC移植组与对照组相比，移植治疗效果差别不大。
在实验的第四部分中，我们专门对各类NSC移植治疗中枢神经系统损伤后的免疫排斥反应问题进行了研究。实验中我们检测了细胞移植前后不同时间点(造模前1天、细胞移植前、细胞移植后3天、1个月及3个月时)各组动物血液中细胞因子(IL-2，IL-6，IL-10)水平，以及细胞移植后急性期(细胞移植后1周)和慢性期(细胞移植后3个月)，移植局部病理切片中CD3+细胞浸润的免疫组化染色情况，力争从全身和移植局部两个方面探讨各类神经干细胞移植后免疫排斥反应这一问题。结果发现：非克隆羊AdNSC组移植后3天、1个月、3个月时IL-2水平较其它组明显升高(P＜0.05)；细胞移植3个月时该组动物血中IL-6水平较其它组明显升高(P＜0.05)；而在移植后3天、1个月、3个月时该组动物血中IL-10水平较其它组明显下降(P＜0.05)。而克隆羊EdNSC及AdNSC移植组之间及它们与对照组相比，各时间点动物血液中细胞因子(IL-2，IL-6，IL-10)水平均没有统计学差异(P＞0.05)。非克隆羊AdNSC(异体细胞移植组)在细胞移植后的急性期(移植后1周)和慢性期(移植后3个月)，移植局部CD3+细胞的数量明显比其它组多【与其它组相比有统计学差异(P＜0.05)】，说明该组移植部位发生了免疫排斥反应。而两个“自体”细胞移植组(克隆羊EdNSC和AdNSC移植组)，移植局部CD3+细胞的数量与对照组相比无明显的增多，这说明这两个“自体”神经干细胞移植组局部未发生明显免疫排斥反应。这两个“自体”细胞移植组之间相比，可以看到EdNSC组CD3+细胞的数量稍多，但二者之间没有统计学差异(P＞0.05)。
综上，通过本实验我们得出了以下结论：1对胚胎克隆羊、成体克隆及非克隆羊通过外科手术取其皮层脑组织，在体外培养均能得到生长良好的胚胎神经干细胞(EdNSC)及成体神经干细胞(AdNSC)。在相同的条件下，EdNSC体外培养速度比成AdNSC快。2对羊脑上薛氏沟周围运动皮层区使用50g×100 cm力度作自由落体加速撞击，可产生观察指标明确且存活率高的羊脑挫裂伤模型，有望广泛用于神经干细胞移植及药物等治疗的实验研究。3胚胎神经干细胞(EdNSC)及成体神经干细胞(AdNSC)移植治疗脑挫裂伤后均有较明显的治疗效果；二者相比较而言胚胎神经干细胞(EdNSC)稍好于成体神经干细胞(AdNSC)。4中枢神经系统异体神经干细胞移植存在着明显的免疫排斥反应，而自体神经干细胞移植基本无免疫排斥反应
The regeneration of nervous tissue in central nervous system is a tough problemin medical science. Neural Stem Cell(NSC)transplantation serves as a newly emergingand a good perspective therapeutic method, has been confirmed in many diseasetherapy in neuroscience domain. Brain trauma is the most frequent disease in clinicalneurosurgery and has largest harm to mankind. It has the topmost therapeutic value.Even though different brain trauma has different pathological process, but most ofthem have the brain contusion and laceration. So We researched the brain contusionand laceration restrictly in our study.
Based on its resource, NSC can be divided into two groups: embryonic derivedneural stem cell(EdNSC)and adult derived neural stem cell(AdNSC). Whosetherapeutic effect on brain contusion and laceration is better? Do immunologicalrejection generate When various kinds of NSC transplanted into brain of animal? Canestablished a brain contusion and laceration model in big animals that have highsurvival rate and can be detected clearly? To these problems, there has no convinceresearch now. In our study, We utilize the cloning goats Whose heredity backgroundare very clearly as the models to estabilsh the brain contusion and laceration ofcloning goats, and investigate the immunological rejection of EdNSC and AdNSCtransplantation.
In the first part of our study, We obtained the cerebral cortex of adult cloninggoat(NO:CL-4) and non-cloning goat(NO:SP-3119) by surgical intervention. Weobtained the AdNSC of them by cell culture in vitro. Then We obtained the cerebralcortex of the fetal cloning goat(35days)Which gained by somatic cell nucleartransfer(SCNT) of CL-4 cloning goat and obtained the EdNSC by cell culture in vitro.To the CL series cloning goats,the EdNSC and AdNSC of cloning goat could be deemas "autoallergic NSC", and AdNSC of non-cloning goat was alloallergic NSC.On theother hand, in the cell culture process, the EdNSC growed faster than AdNSC in thesame cell culture method.
In the second part of our study, We chosed 20 CL series cloning goats at random.After exposed the upper Sylvii sulcus moter cortex of brain of the cloning goats bysurgical intervention, 50 gram of steel ball was used to produce a free fallingaccelerate strike on the brain from 100cm height respectively and established thebrain contusion and laceration goat models that had high survival rate and could be detected clearly.
In the third part of our study, We divided the cloning goat models into 4 groups.We transplanted the EdNSC, AdNSC of cloning goat and AdNSC of non-cloning goatand saline into the around areas of brain contusion and laceration of each groupmodels respectively. We observed each group models' survival rate, ethologicalrecovery; The Motion Evoked Potential(MEP), brain Magnetic ResonanceImaging(MRI) image, pathology changes of brain contusion and laceration in eachgroup models were also recorded. We then observed the survival, immigration anddifferentiation of all kinds of stem cells in goats' brains. We found: all kinds of stemcells can survival, immigrate and different well in the transplanted brain. The EdNSCand AdNSC of cloning goat transplantation groups had good survival rate, ethologyrecovery. Their MEP recovery of the motor cortex, brain MRI image and pathologychanges of brain contusion and laceration were better than AdNSC of non-cloninggoat transplantation group and control group; The EdNSC transplantation group hadthe best therapeutic effect on brain contusion and laceration in all transplantationgroups and control group.
In the fourth part of our study, We investigated the acute(1 week after celltransplantation) and chronic(3 months after cell transplantation) immunologicalrejection of each group of the models Who received various transplanted stem cells.We detected the various cytokine(IL-2,IL-6 and IL-10) in each group models' blood atdifferent time (1 day before model established; before, 3 days, 1 month and 3 monthsafter cell transplantation).We also detected CD3+ cells in the cell transplantation areasaround brain contusion and laceration in all groups goats by the method ofimmunohistochemistry. We found:in the AdNSC of non-cloning transplantationgroup,the level of IL-2 was obviously high and the level of IL- 10 was obviously lowthan other groups at 3 days, 1 month, and 3 months after celltransplantation(P＜0.05).The level of IL-6 was obviously high than other groups at 3months after cell transplantation(P＜0.05). The level of these cytokines in the EdNSC,AdNSC of cloning goat transplantation groups and control group had no statisticsdifference(P＞0.05) at every time. The quantity of CD3+ cells in the AdNSC ofnon-cloning goat group was much more than that of other groups(P＜0.05) at the acuteperiod(1 week after cell transplantation) and chronic period(3 months after celltransplantation). The quantity of CD3+ cells in the EdNSC, AdNSC of cloning goattransplantation group and control group had no statistics difference(P＞0.05) at acute and chronic period.
Overall, in our study, We got the conclusions as follow: 1,The EdNSC, AdNSCof cloning goat and the AdNSC of the non-cloning goat were successful obtained;EdNSC growed faster than AdNSC in the same cell culture method. 2,The braincontusion and laceration model of cloning goats were successful established by freefalling of 50 gram of steel ball accelerate strike on the goat brain from 100cm height.3, EdNSC and AdNSC transplantation both had obvious therapeutic effect on braincontusion and laceration of cloning goats; The therapeutic effect of EdNSCtransplantation was better. 4, Allograft of stem cells has acute and chronicimmunological rejection and autografting of stem cells has no acute and chronicimmunological rejection.
引文
1 《神经干细胞》，徐如祥主编．北京：军事医学科学出版社，2006
2 Isakova IA, Baker K, Dufour J, et al. Preclinical evaluation of adult stem cell engraftment and toxicity in the CNS of rhesus macaques. Mol Ther, 2006; 13(6): 1173-84
3 Aleksandrova MA, Saburina IN, Poltavtseva RA, et al. Behavior of human neural progenitor cells transplanted to rat brain. Dev Brain Res, 2002; 134: 143-8
4 Svendsen CN, Ter Borg MG, Armstrong RJ, et al. A new method for the rapid and long-term growth of human neural precursor cells. J Neurosci Methods, 1998; 85(2): 141-152
5 Flax JD, Aurora S, Yang C, et al. Engraftable human neural stem cells respond to developmental cues, replace neurons, and express foreign genes. Nat Biotechnol, 1998; 16(11): 1033-1039
6 Vescovi AL, Parati EA, Gritti A, et al. Isolation and cloning of multipotential stem cells from the embryonic human CNS and establishment of transplantable human neural stem cell lines by epigenetic stimulation. Exp Neurol, 1999; 156(1): 71-83
7 许汉鹏．第四军医大学博士论文．西安；2001
8 Jianhong zhu, Liangfu zhou, Xing wu, et al. Tracking Neural Stem Cells in Patients with Brain Trauma. the new England journal of medicine. 2006; 355:2376-2378
9 Chojnacki A, Weiss S. Production of neurons, astrocytes and oligodendrocytes from mammalian CNS stem cells. Nat Protoc, 2008; 3(6): 935-40
10 Temple S. Division and differentiation of isolated CNS blast cells in microculture. Nature, 1989; 340: 471-473
11 Elena C, Serena G, Giannellia, et al.Embryonic StemCDerived Versus Somatic Neural Stem Cells:A Comparative Analysis of Their Developmental Potential and Molecular Phenotype. Stem cell, 2006; 24(4): 234-240
12 Soong H, Yu S, Ing I, et al.Whole analysis of Human Neural Stem Cells drived from Embryonic Stem Cells and Stem and Progenitor Cells Isolated from Fetal Tissue, Stem Cells 2007; 25: 1298-1306
13 Conti L, Pollard SM, Gorba T, et al. Niche-independent symmetrical self-renewal of a mammalian tissue stem cell. PLoS Biol, 2005; 3:e283
14 Lendahl U, Zimmerman LB, McKay RDG.. CNS stem cell express a new class of intermediate filament protein. Cell, 1990; 60:585-595
15 Chalfie M, YuTu, Euskirchen G, et al. Green fluorescent protein as a marker for gene expression. Science, 1994; 263 (5148): 802-805
16 Gubin AN, Reddy B, Njoroge JM, et al. Long-term stable expression of green fluorescent protein in mammalian cells. Biochem Biophys Res Commun, 1997; 236(2): 347-350
17 陈治标，陈谦学，张宇蓝．中华实验外科杂志，2003，20：179-180
18 Povlishock JT, Hayes RL, Michel ME, et al. Workshop on animal models of traumatic brain injury. J Neurotrauma, 1994; 11: 723-732
19 Robert WG, Anderson, Christopher J, et al. Impact mechanics and axonal injury in a sheep model. J Neurotrauma, 2003; 20: 961-975
20 陈建良，吕文，郭伟．中华实验外科杂志，2003，20：281-282
21 彭钢，张彭跃，沙红英．山羊脑挫裂伤模型的建立．中华实验外科杂志，2009；26(2)：253-255
22 Hall ED. Hish-dose glucocorticoid treatment improves neurological recovery in head-injured mice. J Neurosurg, 1985; 62: 882-887
23 江基尧，朱诚，罗其中．现代颅脑损伤学[M]．上海：第二军医大学出版社，2004：661
24 Reynolds BA, Weiss S. Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science, 1992; 255: 1707-1709
25 Weiss S. Isolation of a stem cell for neurons and glia from the mammalian neural crest. Cell, 1992; 71: 973-985
26 Mckay R. Stem cells in the central nervous system. Science, 1997; 276:66-71
27 张旺明．脑组织移植与免疫排斥．国外医学：创伤与外科基本问题分册，1998；19(1)：15-8
28 马希峰，阮丽荣，杜英等．人胚脑源性神经干细胞特性及免疫原性研究．中国临床康复杂志．2004；8(28)：6058-9
29 Pakzaban P, Isacson O. Neural xenotransplantation: reconstruction of neuronal circuitry across species barriers. Neuroscience, 1994; 62: 989-1001
30 Gao J, Prough DS, McAdoo DJ. Transplantation of primed human fetal neural stem cells improves cognitive function in rats after traumatic brain injury. Exp Neurol, 2006; 201(2): 281-92
31 Richardson RM, Sun D, Bullock MR. Neurogenesis after traumatic brain injury. Neurosurg Clin N Am, 2007 Jan; 18(1): 169-81
32 Kelly S, Bliss TM, Shah AK, et al. Transplanted human fetal neural stem cells survive, migrate, and differentiate in ischemic rat cerebral cortex. Proc Natl Aced Sci , 2004; 101(32): 11839-11844
33 Blesch A, Lu P, Tuszynski MH. Neurotrophic factors,gene therapy, and neural stem cells for spinal cord repair. Brain Res Bull, 2002; 57: 833-8
34 Ogawa Y, Sawamoto K, Miyata T, et al. Transplantation of in vitroexpanded fetal neural progenitor cells results in neurogenesis and functional revovery after spinal cord contusion injury in adult rats. J Neurosci Res, 2002; 69(6): 561 -563
35 Cao Q, Benton RL, Whittemore SR, et al. Stem cell repair of central nervous system injury. J Neurosci Res, 2002; 68(5): 501-510
36 Stein DG, Palatucci C, Kahn D, et al. Temporal factors influence recovery of function after embryonic brain tissue transplants in adult rats with frontal cortex lesion. Behav Neurosci, 1998; 102(2): 260-267, 325-326
37 Olanow CW, Kowdower JH, Freeman TB. Fetal nigral transplantation as a therapy for Parkinson' s disease. TINS, 1996; 19: 102-109
38 Freeman TB, Olanow CW, Hanser GM, et al. Bilateral fetal nogral transplantation into the postcommissural putamen in Parkinson' s disease. Ann Neurol, 1995; 38:379-388
39 Kelly S, Bliss TM, Shah AK. Transplanted human fetal neural stem cells survive,migrate, and differentiate in ischemic rat cerebral cortex. Proc Natl Acad Sci USA,2004; 101(32): 11839-44
40 Billingham RW, Boswell T. Studies on the problem of corneal honografts. Proc R Soc Lond B Biol Sci, 1953; 141: 392-406
41 Barker CF, Billingham RE. Immunologically privileged sites. Adv Immunol,1977; 25: 1-54
42 Brabb T, von Dassow P, Ordonez N, Schnabel B, Duke B, Goverman J. In situ tolerance within the central nervous system as a mechanism for preventing autoimmunity. J Exp Med, 2000; 192: 871-880
43 Widner H, Brundin P. Immunological aspects of grafting in the mammalian central nervous system. A speculative synthesis. Brain Res Rev, 1988; 13: 287-324
44 Hickey WH. Basic principles of immunological surveillance of the normal central nervous system. Glia, 2001; 36: 118-124
45 Armstrong RJ, Harrower TP, Hurelbrink CB, et al. Porcine neural xenografts in the immunocompetent rat: immune response following grafting of expanded neural precursor cells. Neuroscience, 2001; 106: 201-216
46 Mirza B, Krook H, Andersson P,et al. Widner H. Intracerebral cytokine profiles in adult rats grafted with neural tissue of different immunological disparity. Brain Res Bull, 2004: 63: 105-118
47 刘伟国，郑学胜，杨小锋等．颅脑损伤后移植神经干细胞诱发的淋巴细胞浸润《中华急诊医学杂志》，2007；16(4)：402-405
48 Nimer AF, Wennersten A, Holmin S, et al. MHC expression after human neural stem cell transplantation to brain contused rats. Neuroreport, 2004; 15(12): 1871-1875
49 Modo M, Mellodew K, Rezaie P. In vitro expression of major histocompatibility class Ⅰ and class Ⅱ antigens by conditionally immortalized murine neural stem cells. Neurosci Lett, 2003; 337(2): 85-88
50 McLaren FH, Svendsen CN, Van der Meide P, et al. Analysis of neural stem cells by flow cytometry: cellular differentiation modifies patterns of MHC expression. J Neuroimmunol, 2001; 112 (1/2): 35-46
51 Maier B, Schwerdtfeger K, Mautes A, et al. Differential release of interleukines 6,8, and 10 in cerebrospinal fluid and plasma after traumatic brain injury. Shock, 2001; 15 (6): 421-426
52 Mertelsmann R, Welte K. Human interleukin - 2: molecular biology, physiology and clinical possibilities. Immunbiology, 1986; 172: 400
53 姜宗培．血清及尿白介素-6检测在肾移植中的意义．中华肾脏病杂志，1998；14(4)：240-242
54 Kutukcuter N, Clark K, Rigg KM, et al. Transplantation, 1995; 59: 333-340
55 Wong PK, Quinn JM, Sims NA,et al. Interleukin-6 modulates production of T lymphocyte-derived cytokines in antigen-induced arthritis and drives inflammation-induced osteoclastogenesis. Arthritis Rheum, 2006; 54(1): 158-168
56 Kordower JH, Freeman TB, Snow BJ, et al. Neuropathological evidence of graft survival and striatal reinnervation after the transplantation of fetal mesencephalic tissue in a patient with Parkinson's disease. N Engl J Med, 1995; 332:1118-1124
57 Kordower JH, Styren S, Clarke M,et al. Fetal grafting for Parkinson's disease: expression of immune markers in two patients with functional fetal nigral implants. Cell Transplant, 1997; 6: 213-219
58 Brevig T, Holgersson J, Widner H. Xenotransplantation for CNS repair: immunological barriers and strategies to overcome them. Trends Neurosci, 2000; 23: 337-344
59 Fischer HG, Bielinsky AK. Antigen presentation function of brainderived dendriform cells depends on astrocyte help. Int Immunol, 1999; 11:1265-1274
60 Gould DS, Auchincloss H Jr. Direct and indirect recognition: the role of MHC antigens in graft rejection. Immunol Today, 1999; 20: 77-82
61 Baron JL, Madri JA, Ruddle NH, et al. Surface expression of a4 integrin by CD4 T cells is required for their entry into the brain parenchyma. J Exp Med, 1993; 177: 57-68
62 Merrill JE, Murphy SP. Inflammatory events at the blood brain barrier: regulation of adhesion molecules, cytokines, and chemokines by reactive nitrogen and oxygen species. Brain Behav Immun, 1997; 11: 245-263
63 Wang T, Donahue PK, Zervos AS. Specific interaction of type I receptors of the TGF-family with the immunophilin FKBR-12. Science, 1994; 265:674-676
64 Bechmann I, Mor G, Nilsen J, et al. FasL(CD95L, ApolL) is expressed in the normal rat and human brain:evidence for the existence of an immunological brain barrier. Glia, 1999; 27: 62-74
65 Freed CR, Greene PE, Breeze RE,et al. Transplantation of embryonic dopamine neurons for severe Parkinson's disease. N Engl J Med, 2001; 344:710-719
1 Altman J. Autoradiographic and histological studies of postnatal neurogenesis. Ⅳ. Cell proliferation and migration in the anterior forebrain, with special reference to persisting neurogenesis in the olfactory bulb. J Comp Neurol, 1969; 137:433-457
2 Kaplan MS, Hinds JW. Neurogenesis in the adult rat: electron microscopic analysis of ligh tradioautographs. Science, 1977; 197: 1092-1094
3 Reynolds B A, Weiss S. Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science, 1992; 255:1707-1710
4 Lendahl U, Zimmerman LB, Mckay RD. CNS stem cells express a new class of intermediate filament protein. Cell, 1990; 60:585-595
5 Svendsen CN, Ter Borg MG, Armstrong RJ, et al. A new method for the rapid and long-term growth of human neural precursor cells. J Neurosci Methods, 1998; 85(2): 141-152
6 Flax JD, Aurora S, Yang C, et al. Engraftable human neural stem cells respond to developmental cues, replace neurons, and express foreign genes. Nat Biotechnol, 1998; 16(11): 1033-1039
7 Vescovi AL, Parati EA, Gritti A, et al. Isolation and cloning of multipotential stem cells from the embryonic human CNS and establishment of transplantable human neural stem cell lines by epigenetic stimulation. Exp Neurol, 1999; 156(1): 71-83
8 Williams BP, Read J, Price J. The generation of neurons and oligodendrocytes from a common precursor cell. Neuron, 1991; 7(4): 685-693
9 Schultz SS et al. Adult stem cell application in spinal cord injury. Curr Drug Targets, 2005; 6(1): 63-73
10 Isakova IA, Baker K, Dufour J, et al. Preclinical evaluation of adult stem cell engraftment and toxicity in the CNS of rhesus macaques. Mol Ther, 2006; 13(6): 1173-84
11 肖露露，陈维，张成，等．干细胞移植治疗神经遗传疾病中HLA配合的几率分析．中国实验血液学杂志，2004；12(6)：845-8
12. Pessina A, Gribaldo L. The key role of adult stem cells: therapeutic perspectives. Curr Med Res Opin, 2006; 22(11): 2287-300
13 Ulrica Englund, Anders Bjorklund, Klas Wictorin, et al. Grafted neural stem cells develop into functional pyramidal neurons and integrate into host cortical circuitry. Proc Natl Acad Sci, 2002; 99(26): 7089
14 Kelly S, Bliss TM, Shah AK, et al. Transplanted human fetal neural stem cells survive, migrate, and differentiate in ischemic rat cerebral cortex. Proc Natl Aced Sci USA, 2004; 101(32): 11839-11844
15 Blesch A, Lu P, Tuszynski MH.Neurotrophic factors, gene therapy, and neural stem cells for spinal cord repair. Brain Res Bull, 2002; 57: 833-8
16 朱剑虹，周良辅等．人神经干细胞的分离、克隆和动物脑内移植及转基因表达．中国神经精神疾病杂志．2001：27(3)：183-185
17 Jianhong zhu, liangfu zhou, feng ge, et al. Tracking Neural Stem Cells in Patients with Brain Trauma. the new England journal of medicine, 2006; 355:2376-2378
18 Shihabuddin LS, Homer PJ, Ray J, et al. Adult spinal cord stem cells generate neuros after transplantation in the adult dentate syrus. J Neurosci, 2000; 20: 8727-8735
19 Gould E, Reeves AJ, Fallah M, et al. Hippocampal neurogenesis in adult old world primates. Proc Natl Acad Sci, 1999; 96: 5263-5267
20 Muotri AR, Nakashima K, Toni N, et al. Development of functional human embryonic stem cell-derived neurons in mouse brain. Proc Natl Acad Sci USA, 2005; 102 (51): 18644-18648
21 Lim DA, Huang YC, Alvarez-Buylla A, et al. The adult neural stem cell niche: lessons for future neural cell replacement strategies. Neurosurg Clin N Am, 2007; 18 (1) : 81-92
22 Masoud Tavazoie, Lieven Van der Veken, Violeta Silva-Vargas, et al. A Specialized Vascular Niche for Adult Neural Stem Cells. Cell Stem Cell, 2008; 3: 279-288
23 Zaman Mirzadeh, FlorianT, Merkle, et al. Neural Stem Cells Confer Unique Pinwheel Architecture to the Ventricular Surface in Neurogenic Regions of the Adult Brain. Cell Stem Cell, 2008; 3: 265-278
24 Ogawa Y, Okano H, Nakamura M, et al. Transplantation of in vitro expanded fetal neural progenitor cells results in neurogenesis and functional recovery after spinal cord contusion injury in rats. J Neurosci Res, 2002; 69: 925
25 Luciano Conti, Steven M, Pollard, et al. Niche Independent Symmetrical Self-Renewal of a Mammalian Tissue Stem Cell. PLoS Biol, 2005; 3 (9): e283
26 C(?)dric G Geoffroy, Olivier Raineteau. A Cre-lox approach for transient transgene expression in neural precursor cells and long-term tracking of their progeny in vitro and in vivo. BMC Dev Biol, 2007; 7: 45
27 Cau E, Casarosa S, Guillemot F. Mash land Ngnl control distinct step of determination and differentiation in the olfactory sensory neuron lineage.Development, 2002; 129: 1871-1880
28 Cai L, Morrow EM, Cepko CL. Misexpression of basic helix-loop-helix genes in the murine cerebral cortex affects cell fate choices and neuronal survival.Development, 2000; 127: 3021-3030
29 Milosevic J, Storch A, Schwarz J. Spontaneous apoptosis in murine freefloating neurospheres. Exp Cell Res, 2004; 294: 9-17
30 Micci MA, Pattillo MT, Kahrig KM, et al. Caspase inhibition increases survival of neural stem cells in the gastrointestinal tract. Neurogastroenterol Motil, 2005; 17:557-564
31 Maria-Adelaide Micci, Pankaj Jay Pasricha. Neural Stem Cells for the Treatment of Disorders of the Enteric Nervous System. Strategies and Challenges Developmental dynamics, 2007; 236: 33-43
32 Karlsson J, Petersen A, Gido G, et al. Combining neuroprotective treatment of embryonic nigral donor tissue with mild hypothermia of the graft recipient. Cell Transplant, 2005; 14: 301-309
33 Blesch A, Lu P, Tuszynski MH. Neurotrophic factors, gene therapy, and neural stem cells for spinal cord repair. Brain Res Bull, 2002; 57: 833-8
34Molcanyi M, Riess P, Bentz K, et al. Trauma-associated inflammatory response impairs embryonic stem cell survival and integration after implantation into injured rat brain. J Neurotrauma, 2007; 24 (4): 625-37
35 Taupin P. Autologous transplantation in the central nervous system. Indian J Med Res, 2006; 124 (6): 613-8
36 Hwang WS. Cloned human embryonic stem cells for tissue repair and transplantation. Stem Cell Rev, 2005; 1 (2): 99-109
37 张海燕，徐群渊等．温度敏感型基因永生化神经前体细胞系的建立．解剖学报，2005；36(5)：491-497
2 Isakova IA, Baker K, Dufour J, et al. Preclinical evaluation of adult stem cell engraftment and toxicity in the CNS of rhesus macaques. Mol Ther, 2006; 13(6): 1173-84
3 Aleksandrova MA, Saburina IN, Poltavtseva RA, et al. Behavior of human neural progenitor cells transplanted to rat brain. Dev Brain Res, 2002; 134: 143-8
4 Svendsen CN, Ter Borg MG, Armstrong RJ, et al. A new method for the rapid and long-term growth of human neural precursor cells. J Neurosci Methods, 1998; 85(2): 141-152
5 Flax JD, Aurora S, Yang C, et al. Engraftable human neural stem cells respond to developmental cues, replace neurons, and express foreign genes. Nat Biotechnol, 1998; 16(11): 1033-1039
6 Vescovi AL, Parati EA, Gritti A, et al. Isolation and cloning of multipotential stem cells from the embryonic human CNS and establishment of transplantable human neural stem cell lines by epigenetic stimulation. Exp Neurol, 1999; 156(1): 71-83
7 许汉鹏．第四军医大学博士论文．西安；2001
8 Jianhong zhu, Liangfu zhou, Xing wu, et al. Tracking Neural Stem Cells in Patients with Brain Trauma. the new England journal of medicine. 2006; 355:2376-2378
9 Chojnacki A, Weiss S. Production of neurons, astrocytes and oligodendrocytes from mammalian CNS stem cells. Nat Protoc, 2008; 3(6): 935-40
10 Temple S. Division and differentiation of isolated CNS blast cells in microculture. Nature, 1989; 340: 471-473
11 Elena C, Serena G, Giannellia, et al.Embryonic StemCDerived Versus Somatic Neural Stem Cells:A Comparative Analysis of Their Developmental Potential and Molecular Phenotype. Stem cell, 2006; 24(4): 234-240
12 Soong H, Yu S, Ing I, et al.Whole analysis of Human Neural Stem Cells drived from Embryonic Stem Cells and Stem and Progenitor Cells Isolated from Fetal Tissue, Stem Cells 2007; 25: 1298-1306
13 Conti L, Pollard SM, Gorba T, et al. Niche-independent symmetrical self-renewal of a mammalian tissue stem cell. PLoS Biol, 2005; 3:e283
14 Lendahl U, Zimmerman LB, McKay RDG.. CNS stem cell express a new class of intermediate filament protein. Cell, 1990; 60:585-595
15 Chalfie M, YuTu, Euskirchen G, et al. Green fluorescent protein as a marker for gene expression. Science, 1994; 263 (5148): 802-805
16 Gubin AN, Reddy B, Njoroge JM, et al. Long-term stable expression of green fluorescent protein in mammalian cells. Biochem Biophys Res Commun, 1997; 236(2): 347-350
17 陈治标，陈谦学，张宇蓝．中华实验外科杂志，2003，20：179-180
18 Povlishock JT, Hayes RL, Michel ME, et al. Workshop on animal models of traumatic brain injury. J Neurotrauma, 1994; 11: 723-732
19 Robert WG, Anderson, Christopher J, et al. Impact mechanics and axonal injury in a sheep model. J Neurotrauma, 2003; 20: 961-975
20 陈建良，吕文，郭伟．中华实验外科杂志，2003，20：281-282
21 彭钢，张彭跃，沙红英．山羊脑挫裂伤模型的建立．中华实验外科杂志，2009；26(2)：253-255
22 Hall ED. Hish-dose glucocorticoid treatment improves neurological recovery in head-injured mice. J Neurosurg, 1985; 62: 882-887
23 江基尧，朱诚，罗其中．现代颅脑损伤学[M]．上海：第二军医大学出版社，2004：661
24 Reynolds BA, Weiss S. Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science, 1992; 255: 1707-1709
25 Weiss S. Isolation of a stem cell for neurons and glia from the mammalian neural crest. Cell, 1992; 71: 973-985
26 Mckay R. Stem cells in the central nervous system. Science, 1997; 276:66-71
27 张旺明．脑组织移植与免疫排斥．国外医学：创伤与外科基本问题分册，1998；19(1)：15-8
28 马希峰，阮丽荣，杜英等．人胚脑源性神经干细胞特性及免疫原性研究．中国临床康复杂志．2004；8(28)：6058-9
29 Pakzaban P, Isacson O. Neural xenotransplantation: reconstruction of neuronal circuitry across species barriers. Neuroscience, 1994; 62: 989-1001
30 Gao J, Prough DS, McAdoo DJ. Transplantation of primed human fetal neural stem cells improves cognitive function in rats after traumatic brain injury. Exp Neurol, 2006; 201(2): 281-92
31 Richardson RM, Sun D, Bullock MR. Neurogenesis after traumatic brain injury. Neurosurg Clin N Am, 2007 Jan; 18(1): 169-81
32 Kelly S, Bliss TM, Shah AK, et al. Transplanted human fetal neural stem cells survive, migrate, and differentiate in ischemic rat cerebral cortex. Proc Natl Aced Sci , 2004; 101(32): 11839-11844
33 Blesch A, Lu P, Tuszynski MH. Neurotrophic factors,gene therapy, and neural stem cells for spinal cord repair. Brain Res Bull, 2002; 57: 833-8
34 Ogawa Y, Sawamoto K, Miyata T, et al. Transplantation of in vitroexpanded fetal neural progenitor cells results in neurogenesis and functional revovery after spinal cord contusion injury in adult rats. J Neurosci Res, 2002; 69(6): 561 -563
35 Cao Q, Benton RL, Whittemore SR, et al. Stem cell repair of central nervous system injury. J Neurosci Res, 2002; 68(5): 501-510
36 Stein DG, Palatucci C, Kahn D, et al. Temporal factors influence recovery of function after embryonic brain tissue transplants in adult rats with frontal cortex lesion. Behav Neurosci, 1998; 102(2): 260-267, 325-326
37 Olanow CW, Kowdower JH, Freeman TB. Fetal nigral transplantation as a therapy for Parkinson' s disease. TINS, 1996; 19: 102-109
38 Freeman TB, Olanow CW, Hanser GM, et al. Bilateral fetal nogral transplantation into the postcommissural putamen in Parkinson' s disease. Ann Neurol, 1995; 38:379-388
39 Kelly S, Bliss TM, Shah AK. Transplanted human fetal neural stem cells survive,migrate, and differentiate in ischemic rat cerebral cortex. Proc Natl Acad Sci USA,2004; 101(32): 11839-44
40 Billingham RW, Boswell T. Studies on the problem of corneal honografts. Proc R Soc Lond B Biol Sci, 1953; 141: 392-406
41 Barker CF, Billingham RE. Immunologically privileged sites. Adv Immunol,1977; 25: 1-54
42 Brabb T, von Dassow P, Ordonez N, Schnabel B, Duke B, Goverman J. In situ tolerance within the central nervous system as a mechanism for preventing autoimmunity. J Exp Med, 2000; 192: 871-880
43 Widner H, Brundin P. Immunological aspects of grafting in the mammalian central nervous system. A speculative synthesis. Brain Res Rev, 1988; 13: 287-324
44 Hickey WH. Basic principles of immunological surveillance of the normal central nervous system. Glia, 2001; 36: 118-124
45 Armstrong RJ, Harrower TP, Hurelbrink CB, et al. Porcine neural xenografts in the immunocompetent rat: immune response following grafting of expanded neural precursor cells. Neuroscience, 2001; 106: 201-216
46 Mirza B, Krook H, Andersson P,et al. Widner H. Intracerebral cytokine profiles in adult rats grafted with neural tissue of different immunological disparity. Brain Res Bull, 2004: 63: 105-118
47 刘伟国，郑学胜，杨小锋等．颅脑损伤后移植神经干细胞诱发的淋巴细胞浸润《中华急诊医学杂志》，2007；16(4)：402-405
48 Nimer AF, Wennersten A, Holmin S, et al. MHC expression after human neural stem cell transplantation to brain contused rats. Neuroreport, 2004; 15(12): 1871-1875
49 Modo M, Mellodew K, Rezaie P. In vitro expression of major histocompatibility class Ⅰ and class Ⅱ antigens by conditionally immortalized murine neural stem cells. Neurosci Lett, 2003; 337(2): 85-88
50 McLaren FH, Svendsen CN, Van der Meide P, et al. Analysis of neural stem cells by flow cytometry: cellular differentiation modifies patterns of MHC expression. J Neuroimmunol, 2001; 112 (1/2): 35-46
51 Maier B, Schwerdtfeger K, Mautes A, et al. Differential release of interleukines 6,8, and 10 in cerebrospinal fluid and plasma after traumatic brain injury. Shock, 2001; 15 (6): 421-426
52 Mertelsmann R, Welte K. Human interleukin - 2: molecular biology, physiology and clinical possibilities. Immunbiology, 1986; 172: 400
53 姜宗培．血清及尿白介素-6检测在肾移植中的意义．中华肾脏病杂志，1998；14(4)：240-242
54 Kutukcuter N, Clark K, Rigg KM, et al. Transplantation, 1995; 59: 333-340
55 Wong PK, Quinn JM, Sims NA,et al. Interleukin-6 modulates production of T lymphocyte-derived cytokines in antigen-induced arthritis and drives inflammation-induced osteoclastogenesis. Arthritis Rheum, 2006; 54(1): 158-168
56 Kordower JH, Freeman TB, Snow BJ, et al. Neuropathological evidence of graft survival and striatal reinnervation after the transplantation of fetal mesencephalic tissue in a patient with Parkinson's disease. N Engl J Med, 1995; 332:1118-1124
57 Kordower JH, Styren S, Clarke M,et al. Fetal grafting for Parkinson's disease: expression of immune markers in two patients with functional fetal nigral implants. Cell Transplant, 1997; 6: 213-219
58 Brevig T, Holgersson J, Widner H. Xenotransplantation for CNS repair: immunological barriers and strategies to overcome them. Trends Neurosci, 2000; 23: 337-344
59 Fischer HG, Bielinsky AK. Antigen presentation function of brainderived dendriform cells depends on astrocyte help. Int Immunol, 1999; 11:1265-1274
60 Gould DS, Auchincloss H Jr. Direct and indirect recognition: the role of MHC antigens in graft rejection. Immunol Today, 1999; 20: 77-82
61 Baron JL, Madri JA, Ruddle NH, et al. Surface expression of a4 integrin by CD4 T cells is required for their entry into the brain parenchyma. J Exp Med, 1993; 177: 57-68
62 Merrill JE, Murphy SP. Inflammatory events at the blood brain barrier: regulation of adhesion molecules, cytokines, and chemokines by reactive nitrogen and oxygen species. Brain Behav Immun, 1997; 11: 245-263
63 Wang T, Donahue PK, Zervos AS. Specific interaction of type I receptors of the TGF-family with the immunophilin FKBR-12. Science, 1994; 265:674-676
64 Bechmann I, Mor G, Nilsen J, et al. FasL(CD95L, ApolL) is expressed in the normal rat and human brain:evidence for the existence of an immunological brain barrier. Glia, 1999; 27: 62-74
65 Freed CR, Greene PE, Breeze RE,et al. Transplantation of embryonic dopamine neurons for severe Parkinson's disease. N Engl J Med, 2001; 344:710-719
1 Altman J. Autoradiographic and histological studies of postnatal neurogenesis. Ⅳ. Cell proliferation and migration in the anterior forebrain, with special reference to persisting neurogenesis in the olfactory bulb. J Comp Neurol, 1969; 137:433-457
2 Kaplan MS, Hinds JW. Neurogenesis in the adult rat: electron microscopic analysis of ligh tradioautographs. Science, 1977; 197: 1092-1094
3 Reynolds B A, Weiss S. Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science, 1992; 255:1707-1710
4 Lendahl U, Zimmerman LB, Mckay RD. CNS stem cells express a new class of intermediate filament protein. Cell, 1990; 60:585-595
5 Svendsen CN, Ter Borg MG, Armstrong RJ, et al. A new method for the rapid and long-term growth of human neural precursor cells. J Neurosci Methods, 1998; 85(2): 141-152
6 Flax JD, Aurora S, Yang C, et al. Engraftable human neural stem cells respond to developmental cues, replace neurons, and express foreign genes. Nat Biotechnol, 1998; 16(11): 1033-1039
7 Vescovi AL, Parati EA, Gritti A, et al. Isolation and cloning of multipotential stem cells from the embryonic human CNS and establishment of transplantable human neural stem cell lines by epigenetic stimulation. Exp Neurol, 1999; 156(1): 71-83
8 Williams BP, Read J, Price J. The generation of neurons and oligodendrocytes from a common precursor cell. Neuron, 1991; 7(4): 685-693
9 Schultz SS et al. Adult stem cell application in spinal cord injury. Curr Drug Targets, 2005; 6(1): 63-73
10 Isakova IA, Baker K, Dufour J, et al. Preclinical evaluation of adult stem cell engraftment and toxicity in the CNS of rhesus macaques. Mol Ther, 2006; 13(6): 1173-84
11 肖露露，陈维，张成，等．干细胞移植治疗神经遗传疾病中HLA配合的几率分析．中国实验血液学杂志，2004；12(6)：845-8
12. Pessina A, Gribaldo L. The key role of adult stem cells: therapeutic perspectives. Curr Med Res Opin, 2006; 22(11): 2287-300
13 Ulrica Englund, Anders Bjorklund, Klas Wictorin, et al. Grafted neural stem cells develop into functional pyramidal neurons and integrate into host cortical circuitry. Proc Natl Acad Sci, 2002; 99(26): 7089
14 Kelly S, Bliss TM, Shah AK, et al. Transplanted human fetal neural stem cells survive, migrate, and differentiate in ischemic rat cerebral cortex. Proc Natl Aced Sci USA, 2004; 101(32): 11839-11844
15 Blesch A, Lu P, Tuszynski MH.Neurotrophic factors, gene therapy, and neural stem cells for spinal cord repair. Brain Res Bull, 2002; 57: 833-8
16 朱剑虹，周良辅等．人神经干细胞的分离、克隆和动物脑内移植及转基因表达．中国神经精神疾病杂志．2001：27(3)：183-185
17 Jianhong zhu, liangfu zhou, feng ge, et al. Tracking Neural Stem Cells in Patients with Brain Trauma. the new England journal of medicine, 2006; 355:2376-2378
18 Shihabuddin LS, Homer PJ, Ray J, et al. Adult spinal cord stem cells generate neuros after transplantation in the adult dentate syrus. J Neurosci, 2000; 20: 8727-8735
19 Gould E, Reeves AJ, Fallah M, et al. Hippocampal neurogenesis in adult old world primates. Proc Natl Acad Sci, 1999; 96: 5263-5267
20 Muotri AR, Nakashima K, Toni N, et al. Development of functional human embryonic stem cell-derived neurons in mouse brain. Proc Natl Acad Sci USA, 2005; 102 (51): 18644-18648
21 Lim DA, Huang YC, Alvarez-Buylla A, et al. The adult neural stem cell niche: lessons for future neural cell replacement strategies. Neurosurg Clin N Am, 2007; 18 (1) : 81-92
22 Masoud Tavazoie, Lieven Van der Veken, Violeta Silva-Vargas, et al. A Specialized Vascular Niche for Adult Neural Stem Cells. Cell Stem Cell, 2008; 3: 279-288
23 Zaman Mirzadeh, FlorianT, Merkle, et al. Neural Stem Cells Confer Unique Pinwheel Architecture to the Ventricular Surface in Neurogenic Regions of the Adult Brain. Cell Stem Cell, 2008; 3: 265-278
24 Ogawa Y, Okano H, Nakamura M, et al. Transplantation of in vitro expanded fetal neural progenitor cells results in neurogenesis and functional recovery after spinal cord contusion injury in rats. J Neurosci Res, 2002; 69: 925
25 Luciano Conti, Steven M, Pollard, et al. Niche Independent Symmetrical Self-Renewal of a Mammalian Tissue Stem Cell. PLoS Biol, 2005; 3 (9): e283
26 C(?)dric G Geoffroy, Olivier Raineteau. A Cre-lox approach for transient transgene expression in neural precursor cells and long-term tracking of their progeny in vitro and in vivo. BMC Dev Biol, 2007; 7: 45
27 Cau E, Casarosa S, Guillemot F. Mash land Ngnl control distinct step of determination and differentiation in the olfactory sensory neuron lineage.Development, 2002; 129: 1871-1880
28 Cai L, Morrow EM, Cepko CL. Misexpression of basic helix-loop-helix genes in the murine cerebral cortex affects cell fate choices and neuronal survival.Development, 2000; 127: 3021-3030
29 Milosevic J, Storch A, Schwarz J. Spontaneous apoptosis in murine freefloating neurospheres. Exp Cell Res, 2004; 294: 9-17
30 Micci MA, Pattillo MT, Kahrig KM, et al. Caspase inhibition increases survival of neural stem cells in the gastrointestinal tract. Neurogastroenterol Motil, 2005; 17:557-564
31 Maria-Adelaide Micci, Pankaj Jay Pasricha. Neural Stem Cells for the Treatment of Disorders of the Enteric Nervous System. Strategies and Challenges Developmental dynamics, 2007; 236: 33-43
32 Karlsson J, Petersen A, Gido G, et al. Combining neuroprotective treatment of embryonic nigral donor tissue with mild hypothermia of the graft recipient. Cell Transplant, 2005; 14: 301-309
33 Blesch A, Lu P, Tuszynski MH. Neurotrophic factors, gene therapy, and neural stem cells for spinal cord repair. Brain Res Bull, 2002; 57: 833-8
34Molcanyi M, Riess P, Bentz K, et al. Trauma-associated inflammatory response impairs embryonic stem cell survival and integration after implantation into injured rat brain. J Neurotrauma, 2007; 24 (4): 625-37
35 Taupin P. Autologous transplantation in the central nervous system. Indian J Med Res, 2006; 124 (6): 613-8
36 Hwang WS. Cloned human embryonic stem cells for tissue repair and transplantation. Stem Cell Rev, 2005; 1 (2): 99-109
37 张海燕，徐群渊等．温度敏感型基因永生化神经前体细胞系的建立．解剖学报，2005；36(5)：491-497