永生化神经前体细胞治疗大鼠局灶性脑缺血的实验研究

目的 探讨永生化神经前体细胞(INPC)移植对局灶性脑缺血大鼠神经功能的影响及移植细胞在模型大鼠脑内的存活以及分化情况.方法 雄性SD大鼠24只,均采用线栓法建立大脑中动脉缺血(MCAO)模型,随机分为2组(每组12只):脑缺血对照组、INPC移植组.缺血后3 d通过立体定位注射方法分别将等体积的磷酸盐缓冲液、5-溴脱氧尿嘧啶核苷(BrdU)标记的INPC悬液注射到2组大鼠纹状体缺血半暗带区.缺血再灌注后对大鼠进行神经损害严重程度评分(NSS).细胞移植后1和4周分别随机处死2组大鼠各6只,取脑组织制作冰冻切片,通过免疫荧光双标技术检测BrdU、胶质纤维酸性蛋白(GFAP)双阳性细胞和BrdU、神经元特异性烯醇化酶(NSE)双阳性细胞,观察INPC在移植区域的存活及分化情况.结果 脑缺血对照组与INPC移植组比较,细胞移植前后各时点的NSS评分差异均无统计学意义(均P＞0.05).INPC移植组细胞移植后1及4周,均可在移植针道附近及缺血灶周围检测到聚集较明显的BrdU免疫荧光染色阳性的植入细胞,并观察到BrdU染色阳性细胞扩散至全脑,免疫荧光双标技术检测见部分细胞为BrdU、GFAP双阳性的星形胶质细胞或BrdU、NSE双阳性神经元.结论 INPC可在局灶性脑缺血大鼠脑内存活并分化为神经元和星形胶质细胞.

Transplantation of immortalized neural progenitor cells to treat focal cerebral ischemia: experiment with rats

OBJECTIVE: To investigate the impact of transplantation of immortalized neural progenitor cells (INPCs) into the brain with focal cerebral ischemia and the survival and differentiation thereof. METHODS: Twenty-four male SD rats underwent middle cerebral artery occlusion (MCAO) and were randomly divided into 2 equal groups: INPC group undergoing transplantation of BrdU-labeled INPCs into the penumbra zone in striatum using stereotaxic apparatus 3 days after brain ischemia, and control group undergoing transplantation of PBS. Neurological severity score (NSS) system was used 6 hours, 1 day, and 3 days after MCAO, and 1, 2, 3, and 4 weeks respectively after transplantation. One week after transplantation, 6 animals of each group were randomly chosen and killed with their brains taken out, the rest of the animals were killed four weeks after transplantation. The survival of INPCs in the brain was determined by double immunofluorescent labeling technique with BrdU + NSE or BrdU + GFAP antibody. RESULTS: The NSS values at different time points after transplantation of both groups were all higher than those before MCAO; however, no significant difference in NSS was detected between the two groups. BrdU + GFAP and BrdU + NSE positive astrocytes and neurons were detected in the INPC group by double immunofluorescent labeling technique 1 week and 4 weeks after transplantation. CONCLUSION: INPC can survive in penumbra zone in rats with focal cerebral ischemia and develop into neurons and astrocytes.