未成熟大鼠脑白质损伤对miR-219变化的影响

Effects of white matter injury on miR-219 changes in premature rats

目的:研究miR-219在未成熟大鼠脑白质损伤时的变化,探索早产儿脑白质损伤后髓鞘化障碍的发病机制。方法:缺血缺氧(HI)法建立未成熟大鼠脑白质损伤的动物模型。4日龄SD大鼠随机分为实验组(n=80)和对照组(n=60)。实验组予右侧颈总动脉永久性结扎后,6%氧气、94%氮气吸入90 min。q PCR检测HI后1 d、3 d、7 d时两组胼胝体miR-219的动态表达变化,免疫印迹检测HI后3 d、7 d时PDGFRα在胼胝体中的表达,Morris水迷宫评估大鼠认知和记忆能力。结果:成功建立了未成熟大鼠脑白质损伤的动物模型,HI后7天两组大鼠胼胝体miR-219的相对表达量均较1天和3天时明显上升(P<0.01),但实验组7 d时miR-219表达量显著低于对照组(6.63±1.39 vs 13.19±2.23,P<0.01); HI后7 d时两组大鼠胼胝体PDGFRα蛋白表达比3天时均有所降低,但实验组7 d时PDGFRα蛋白表达降低较慢,其相对灰度值显著高于对照组(0.93±0.04 vs 0.76±0.05,P<0.01);实验组大鼠HI后14天髓鞘碱性蛋白表达降低,成年期Morris水迷宫实验显示其逃避潜伏期时间长于对照组(P<0.05),第5天60 s内穿越平台次数少于对照组(P<0.05),在目标象限停留的时间百分比低于对照组(P<0.01)。结论:未成熟大鼠脑白质损伤时miR-219的表达在发育过程中其上升趋势受到抑制,并伴有PDGFRα的表达相对增加,提示miR-219在早产儿脑白质损伤后髓鞘化障碍发生机制中发挥重要作用。

Objective:To determine the changes of miR-219 in white matter injury of premature rats,and to investigate the pathogenesis of myelination failure in premature infants with white matter injury.Methods:Hypoxia-ischemia(HI) method was used to establish animal model of white matter injury in premature rats.The4-day-old SD rats were randomly divided into the study group(n=80) and control group(n=60).After the permanent ligation of right common carotid artery was performed in the study group,6% oxygen and 94% nitrogen were inhaled for for 90 min.q PCR was used to determine the expression changes of miR-219 in corpus callosum of both groups at 1,3 and 7 d after HI.Western blotting was used to determine the expression level of PDGFRα in corpus callosum of both groups at 3 and 7 d after HI.Morris water maze was used to evaluate cognitive and memory abilities in rats.Results:The animal models of white matter injury in premature rats were successfully established.The relative expression level of miR-219 in corpus callosum of the two groups at 7 d after HI was significantly higher than that at 1 and 3 d after HI(P<0.01).The expression level of miR-219 at 7 d in the study group was significantly lower than that in the control group(6.63±1.39 vs 13.19±2.23,P<0.01).The protein expression level of PDGFRα in corpus callosum of the two groups decreased at 7 d after HI compared with that at3 d,but the protein expression level of PDGFRα decreased slower in the study group at 7 d,and the relative gray value was significantly higher than that in the control group(0.93±0.04 vs 0.76±0.05,P<0.01).The expression level of myelin basic protein decreased in the study group at 14 d after HI.The adult Morris water maze test showed that the escape latency in the study group was longer than that in the control group(P<0.05),with the times of crossing platform fewer than that in the control group(P<0.05),and the percentage of time in target quadrant lower than that in the control group(P<0.01).Conclusion:The increasing trend of miR-219 expression in premature rats with white matter injury during development was inhibited,with corresponding increase of PDGFRα expression,suggesting that miR-219 may play an important role in myelination failure of premature infants with white matter injury.