脐血源神经干细胞移植改善局灶性脑缺血大鼠神经功能障碍  

Umbilical cord blood neural stem cells therapy in improving neurological disorders in rats with focal cerebral ischemia

作　　者：马建华[1,2] 吴月奎[1] 易波[1] 张业森[1] 田中秋 刘震 戴宜武 徐如祥 

机构地区：[1]安徽医科大学北京军区总医院临床学院,北京100700 [2]八一脑科医院

出　　处：《中华神经医学杂志》2014年第4期353-357,共5页Chinese Journal of Neuromedicine

基　　金：国家自然科学基金（81271391）

摘　　要：目的探讨脐血源神经干细胞移植对局灶性脑缺血大鼠神经功能障碍的修复改善作用。方法无菌条件下收集脐带血．加入淋巴细胞分离液，进行梯度离心获取单个核细胞悬浮液，接种于专用培养基中培养，进行传代。并对获得的细胞进行nestin染色鉴定。将造模成功的大脑中动脉栓塞模型大鼠60只按随机数字表法分成移植组和对照组，每组30只，分别于造模后2d从侧脑室注射脐血源神经干细胞和磷酸盐缓冲液，同时腹腔注射5．溴脱氧尿嘧啶核苷（BrdU）标记内源性神经干细胞。分别于移植后1d、1周、2周对2组大鼠进行改良神经功能缺损评分（mNSS）；2周时处死全部大鼠，进行TTC染色观察脑梗死体积百分比，并进行血管内皮生长因子（VEGF）及双肾上腺皮质激素（DCX）、神经胶质纤维酸性蛋白（GFAP）免疫荧光染色观察。结果在移植后1周、2周时，移植组和对照组相比mNSS评分明显降低（6．88±0．35vs 7．75±0．46；4．38±0．52vs6．25±0．71），差异均有统计学意义（P〈0．05）；移植后2周时，移植组和对照组相比脑梗死体积明显缩小（16．99％±0134％vs 34．16％±0．49％），BrdU／DCX、BrdU／GFAP双重标记阳性细胞数量明显增多（72．50±2．72vs46．00±2．43：63．14±2．18vs32．29±1．76），VEGF表达量明显增高（112．43±4．24vs64．9±3．81），差异均有统计学意义（P〈0．05）。结论脐血源神经干细胞移植能促进神经发生和VEGF分泌，对局灶性脑缺血大鼠神经功能障碍有改善作用。Objective To investigate the effect of umbilical cord blood neural stem cells （UCBNSCs） on functional recovery after focal cerebral ischemia in rats. Methods Cord blood was collected under aseptic conditions and added into the lymphocyte separation medium. The single nuclear cell suspension was obtained by the gradient centrifugation. The cell suspension was inoculated in special culture medium to subculture. The neural stem cells were identified by nestin testing. Middle cerebral artery occlusion （MCAO） rat models were divided into two groups： those received UCBNSCs or phosphate buffer saline （PBS） through lateral ventricle 2 days after MCAO, and both injected 5-Bromo-2＇-Deoxyuridine （BrdU） to label neural stem cells. Animals were underwent modified neurological severity scale （mNSS） 1 d, 1 w and 2 w after cell transplantation, and then, these rats were sacrificed 2 w after cell transplantation. TTC staining was employed to observe the infarct volume; the vascular endothelial growth factor （VEGF）, BrdU, doublecortin （DCX） and glial fibrillary acidic protein （GFAP） expressions were detected by immunofluorescent method. Results As compared with those in the PBS transplantation group, the mNSS scores were significantly decreased 1 w and 2 w after transplantation（6.88±0.35 vs 7.75±0.46; 4.38±0.52 vs 6.25±0.71） and the infarct volume was obviouslydecreased 2 w after transplantation （16.99%±0.34% vs 34.16%±0.49%） in UCBNSCs transplantation group with statistical difference （P〈0.05）. UCBNSCs transplantation group had significantly increased endogenous neural stem cell proliferation and differentiation （72.50±2.72 vs 46.00±2.43 ;63.14±2.18 vs 32.29±1.76）, as well as increased VEGF secretion （112.43±4.24 vs 64.95±3.81） as compared with PBS transplantation group（P〈0.05）. Conclusion Transplantation of UCBNSCs can promote neurogenesis and VEGF secretion, and be benefit for focal cerebral ischemia.

关 键 词：脐血源神经干细胞 脑缺血 细胞移植 神经功能障碍 

分 类 号：R743.33[医药卫生—神经病学与精神病学]