新生小鼠脑切片体外循环灌流模型中白质损伤的研究  

White matter injury after cardiopulmonary bypass in a brain slice model of neonatal rats with perfusion

作　　者：黄骏荣 祝忠群[1] 刘刚[1] 陈会文 冯林音[2] 

机构地区：[1]上海交通大学医学院附属上海儿童医学中心心胸外科,上海200127 [2]中国科学院上海药物研究所,上海201203

出　　处：《中国胸心血管外科临床杂志》2017年第4期295-300,共6页Chinese Journal of Clinical Thoracic and Cardiovascular Surgery

摘　　要：目的通过建立新生小鼠脑切片体外循环灌流模型,探讨该模型能否复制体外循环(CPB)和深低温停循环(DHCA)下脑损伤病理生理状态,并研究脑切片灌流及氧糖剥夺(OGD)是否造成白质中少突胶质前体细胞(pre OL)的损伤,为体外循环对白质损伤提供细胞学证据。方法将40只7天龄Sprague-Dawley大鼠,平均体质量(14.7±1.5)g,随机制作活体脑切片,每只大鼠约切出3~5片脑切片。根据灌注方式和温度不同分为5组,每组含脑切片24片:对照组全程人工脑脊液(a CSF)灌流(36℃);实验组在不同温度(36℃,32℃,25℃和15℃)OGD 60 min,建立离体脑切片灌流系统,模拟心脏手术CPB/DHCA整个过程。应用免疫组化检测髓鞘碱性蛋白MBP及少突胶质细胞标记物O4抗体评价少突胶质细胞损伤程度。结果在OGD实验组中,成熟少突胶质(MBP阳性)细胞存在明显损伤,其形态学明显变化和荧光表达明显减少,而且OGD温度越高,损伤也越严重;pre OL(O4阳性)细胞在36℃,32℃及25℃组均出现不同程度的荧光表达减少,且OGD温度越高,荧光表达减少越明显,O4阳性细胞的形态学和荧光表达改变在对照组及15℃OGD组之间差异无统计学意义。结论本模型能有效模拟CPB/DHCA下脑损伤病理生理状态,而且这种损伤可导致少突胶质细胞系发育关键阶段的pre OL的损伤,分化为下游的少突胶质细胞(OL)数量减少,最终导致轴突低髓鞘化和白质损伤。Objective Through establishment of brain slice model in rats with perfusion and oxygen glucose deprivation（OGD）,we investigated whether this model can replicate the pathophysiology of brain injury in cardiopulmonary bypass（CPB） and deep hypothermic circulatory arrest（DHCA） or not and whether perfusion and OGD can induce preoligodendrocytes（preOL） injury or not,to provide cytological evidence for white matter injury after cardiopulmonary bypass.Methods Three to five living brain slices were randomly obtained from each of forty sevenday-old（P7） Sprague-Dawley（SD） rats with a mean weight of 14.7±1.5 g.Brain slices were randomly divided into five groups with 24 slices in each group：control group with normothermic artificial cerebralspinal fluid（aCSF） perfusion（36℃） and DHCA groups：OGD at 15℃,25℃,32℃ and 36℃.The perfusion system was established,and the whole process of CPB and DHCA in cardiac surgery was simulated.The degree of oligodendrocyte injury was evaluated by MBP and O4 antibody via application of immunohistochemistry.Results In the OGD group,the mature oligodendrocytes（MBP-positive） cells were significantly damaged,their morphology was greatly changed and fluorescence expression was significantly reduced.The higher the OGD temperature was,the more serious the damage was;preOL（O4-positive） cells showed different levels of fluorescence expression reduce in 36℃,32℃ and 25℃ groups,and the higher the OGD temperature was,the more obvious decrease in fluorescence expression was.There was no statistically significant difference in the O4-positive cells between the control group and the 15℃ OGD group.Conclusion The perfused brain slice model is effective to replicate the pathophysiology of brain injury in CPB/DHCA which can induce preOL damage that is in critical development stages of oligodendrocyte cell line,and reduce differentiation of oligodendrocyte cells and eventually leads to hypomyelination as well as cerebral white matter injury.

关 键 词：先天性心脏病 体外循环 深低温停循环 白质 少突胶质前体细胞 

分 类 号：R-332[医药卫生] R654.1