包新杰;魏俊吉;冯铭;卢珊;赵春华;王任直;
BAO Xin-jie1,WEI Jun-ji1,FENG Ming1,LU Shan2,ZHAO Chun-hua2,WANG Ren-zhi11Department of Neurosurgery,Peking Union Medical College Hospital,Chinese Academy of Medical Sciences & Peking Union Medical College,Beijing 100730,China 2Center in Tissue Engineering,Institute of Basic Medical Sciences,Chinese Academy of Medical Sciences & Peking Union Medical College,Beijing 100730,China

• 1:中国医学科学院北京协和医学院北京协和医院神经外科
• 2:中国医学科学院北京协和医学院基础医学研究所组织工程中心

摘要(Abstract)：

目的探讨移植骨髓间充质干细胞(bone marrow-derived mesenchymal stemcells,BMSCs)治疗大鼠脑梗死及其减轻半暗带神经元的凋亡作用。方法采集成人志愿者骨髓,分离培养BMSCs;线栓法建立大鼠大脑中动脉栓塞(middle cerebral artery occlusion,MCAO)2h再灌注脑梗死模型;建模后第3天分别立体定向脑内移植5×105BMSCs和等量生理盐水,改良神经功能损害评分(modified neurological severity scores,mNSS)观察神经功能恢复;建模后第14天TUNEL法检测梗死周边神经细胞凋亡;建模后第28天免疫荧光染色检测梗死周边神经元的存活;采用依红-苏木素染色评估梗死体积。结果治疗组和对照组比较,梗死周边TUNEL阳性细胞显著减少(P=0.017),而神经元核抗原(neuronal nuclei antigen,NeuN)阳性细胞显著增多(P=0.015);同时,BMSCs可显著降低MCAO大鼠的相对梗死体积(P=0.041);mNSS评分显示,从MCAO后第14天开始,治疗组评分均显著低于对照组(P<0.05)。结论脑梗死急性期移植BMSCs可显著减少半暗带神经元凋亡,缩小梗死范围,有效促进神经功能的恢复。
Objective To investigate the apoptosis of neurons in the penumbra of cerebral ischemic rats transplanted with bone marrow derived-mesenchymal stem cells(BMSCs).Methods Bone marrow of a healthy adult volunteer was collected,and BMSCs were cultivated and harvested.Adult rats subjected to 2 h middle cerebral artery occlusion(MCAO)were randomly assigned to the BMSCs-treated group(n=12)and the control group(n=12),in which rats were transplanted with 5×105 BMSCs or saline,respectively,into the ipsilateral brain parenchyma on the 3rd post-MCAO day.The modified neurological severity scores(mNSS)was used to measure functional recovery.On the 14th post-MCAO day,apoptotic cells in the penumbra were detected by TUNEL analysis.On the 28th post-MCAO day,neurons were stained by immunoflu-orescence,and hematoxylin-eosin staining was used to evaluate the relative ischemic volume.Results TUNEL-positive cells were significantly reduced(P=0.017),and in contrast,neuronal nuclei antigen(NeuN)-positive cells were significantly increased(P=0.015)in the penumbra in BMSCs-treated group compared with control group.The relative ischemic volume was decreased in the BMSCs-treated group(28.3±4.5)% compared with the control group(36.8±5.6)%(P=0.041).The mNSS was significantly decreased in BMSCs-treated group at each time point starting on the 14th post-MCAO day compared with control group(P<0.05).Conclusion BMSCs treatment can decrease ischemic volume and improve functional recovery after cerebral ischemia,which may,at least partially,attribute to the reduced apoptosis of neurons in the penumbra.

关键词(KeyWords)： 脑梗死;间充质干细胞;神经元;凋亡
cerebral ischemia;mesenchymal stem cells;neuron;apoptosis

Abstract:

Keywords:

基金项目(Foundation): 国家863计划重点项目(2006AA02A115)

作者(Author): 包新杰;魏俊吉;冯铭;卢珊;赵春华;王任直;
BAO Xin-jie1,WEI Jun-ji1,FENG Ming1,LU Shan2,ZHAO Chun-hua2,WANG Ren-zhi11Department of Neurosurgery,Peking Union Medical College Hospital,Chinese Academy of Medical Sciences & Peking Union Medical College,Beijing 100730,China 2Center in Tissue Engineering,Institute of Basic Medical Sciences,Chinese Academy of Medical Sciences & Peking Union Medical College,Beijing 100730,China

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DOI:

参考文献(References)：

• [1]Paolo B,Pamela GR,Paul JS.Mesenchymal Stem Cells:Revisiting History,Concepts,and Assays[J].Cell Stem Cell,2008,2:313-319.
• [2]Liu L,Sun Z,Chen B,etal.Exvivo expansion and in vivo infusion of bone marrow-derived Flk-1+CD31-CD34-mes-enchymal stem cells:feasibility and safety from monkey to human[J].Stem Cells Dev,2006,15:349-357.
• [3]Zhang Z,Wang R,Wang R,et al.Transplantation of neu-ral stem cells modified by human neurotrophin-3promotes functional recovery after transient focal cerebral ischemia in rats[J].Neurosci Lett,2008,444:227-230.
• [4]Kim SS,Yoo SW,Park TS,et al.Neural induction with neurogenin1increases the therapeutic effects of mesenchymalstem cells in the ischemic brain[J].Stem Cells,2008,26:2217-2228.
• [5]Andrews EM,Tsai SY,Johnson SC,et al.Human adult bone marrow-derived somatic cell therapy results in function-al recovery and axonal plasticity following stroke in the rat[J].Exp Neurol,2008,211:588-592.
• [6]Shen LH,Li Y,Chen J,et al.Therapeutic benefit of bone marrowstromal cells administered1month after stroke[J].J Cereb Blood FlowMetab,2007,27:6-13.
• [7]Li J,Zhu H,Liu Y,et al.Human mesenchymal stem cell transplantation protects against cerebral ischemic injury and upregulates interleukin-10expression in Macaca fascicularis[J].Brain Res,2010,1334:65-72.
• [8]魏俊吉,曾立芬,樊晓彤,等.骨髓间充质干细胞治疗大鼠缺血性脑卒中的实验研究[J].中华医学杂志,2007,87:184-189.
• [9]Bang OY,Lee JS,Lee PH,et al.Autologous mesenchymal stem cell transplantation in stroke patients[J].Ann Neu-rol,2005,57:874-882.
• [10]Prockop DJ.“Stemness”does notexplain the repairof many tissues by mesenchymal stem/multipotent stromal cells(MSCs)[J].Clin Pharmacol Ther,2007,82:241-243.
• [11]Li Y,Chen J,Chen XG,et al.Human marrowstromal cell therapy for stroke in rat:neurotrophins and functional recov-ery[J].Neurology,2002,59:514-523.
• [12]Li J,Zhu H,Lu S,et al.Migration and differentiation of human mesenchymal stem cells in the normal rat brain[J/OL].Neurol Res,2010http://www.ingentaconnect.com/content/maney/nres.
• [13]Raichle ME.The pathophysiology of brain ischemia[J].Ann Neurol,1983,13:2-10.
• [14]The STEPS Participants.Stem cell therapies as an emerging paradigm in stroke(STEPS):bridging basic and clinical science for cellular and neurogenic factor therapy in treating stroke[J].Stroke,2009,40:510-515.