选择性脑低温对脑缺血-再灌注大鼠皮质长链非编码RNA AK009271的影响  

Effect of selective brain hypothermia on long noncoding RNA AK009271 of cerebral cortex following cerebral ischemia-reperfusion in rats

作　　者：唐亚男 张高峰 王明山[1] Tang Ya′nan;Zhang Gaofeng;Wang Mingshan(Department of Anesthesiology,Qingdao Municipal Hospital Affiliated to Qingdao University,Qingdao 266071,China)

机构地区：[1]青岛大学附属青岛市市立医院麻醉科

出　　处：《中国脑血管病杂志》2019年第12期647-652,672,共7页Chinese Journal of Cerebrovascular Diseases

基　　金：青岛市民生科技计划项目(19-6-1-50-nsh)

摘　　要：目的观察脑缺血-再灌注大鼠皮质长链非编码RNA(LncRNA)AK009271的表达水平,并探讨选择性脑低温在脑缺血-再灌注损伤时对其表达的影响。方法SPF级健康雄性Sprague-Dawley(SD)大鼠75只,体质量200~250 g,按随机数字量表法随机分为假手术组(S组)、缺血-再灌注组(I/R组)、低温+缺血-再灌注组(HT组),每组25只。采用Longa线栓法栓塞大脑中动脉2 h后再灌注建立局部脑缺血-再灌注模型,HT组于再灌注即刻行颈内动脉灌注4℃等渗盐水15 min(20 ml/kg),术中持续监测脑温和肛温以确保选择性脑低温模型成功构建。S组仅分离颈动脉,而不进行大脑中动脉栓塞。再灌注6、24、48 h后行神经功能缺损评分,定量即时聚合酶链反应(qRT-PCR)测定LncRNA AK009271的表达水平;再灌注24 h Western Blot测定线粒体分裂蛋白1(Fis1)的表达水平,电镜下观察线粒体超微结构,原位末端标记法(TUNEL)检测细胞凋亡。结果(1)与I/R组比较,HT组4℃等渗盐水灌注15 min时脑温降低[(32.27±0.15)℃比(34.52±0.22)℃,t=1.603,P<0.05],而肛温差异无统计学意义(P>0.05)。(2)与S组比较,其余两组神经功能缺损评分增高;与I/R组比较,HT组神经功能缺损评分降低;差异均有统计学意义(均P<0.05)。(3)与S组比较,I/R组再灌注6、24、48 h后LncRNA AK009271的表达下调(均P<0.05),于再灌注24 h表达最低(均P<0.05);与I/R组比较,HT组再灌注各时点LncRNA AK009271的表达上调(均P<0.05)。(4)与S组比较,再灌注24 h后其余两组Fis1的表达上调(均P<0.05),线粒体超微结构出现损伤,细胞凋亡率增加(均P<0.05);与I/R组比较,再灌注24 h后HT组Fis1的表达下调(P<0.05),线粒体超微结构损伤较轻,细胞凋亡率降低(P<0.05)。结论脑缺血-再灌注大鼠皮质LncRNA AK009271的表达下调,选择性脑低温通过上调LncRNA AK009271的表达水平,进而抑制Fis1的表达,减轻细胞凋亡,从而发挥脑保护作用。Objective To observe the expression of long noncoding RNA(LncRNA)AK009271 in the cortex of rats with cerebral ischemia-reperfusion,and to evaluate the effect of selective brain hypothermia on the its expression.Methods All 75 healthy male Sprague-Dawley(SD)rats of SPF grade,weighing 200-250 g,were randomly divided into the sham operation group(group S),the ischemia-reperfusion group(group I/R)and the hypothermia group(group HT)by random digital scale method,with 25 rats in each group.Focal cerebral ischemia-reperfusion model was induced by string Longa′s thread thrombus method of the middle cerebral artery for 2 hours.The group HT was infused with 4°C cold isotonic saline for 15 min(20 ml/kg)through internal carotid artery immediately after reperfusion.During the procedure,brain and rectal temperatures were monitored to ensure successful construction of selective brain hypothermia model.The group S only separated the carotid artery without performing middle cerebral artery embolization.At 6,24,and 48 h after reperfusion,the neurological deficit scores were measured and the expression of LncRNA AK009271 was evaluated by quantitative real-time polymerase chain reaction(qRT-PCR).At 24 h after reperfusion,the expression of Fis1 was determined by Western Blot,and mitochondrial morphology was observed by electron microscopy.The apoptosis rate of cells was detected by TUNEL staining.Results(1)Compared with the group I/R,the group HT had decreased brain temperature after perfusion of 4°C isotonic saline for 15 min(32.27±0.15 vs.34.52±0.22,t=1.603,P<0.05),but without significant changes in rectal temperature(P>0.05).(2)Compared with the group S,the neurological deficit scores were increased in the other two groups;the neurological deficit score in the group HT was lower than the group I/R;all P<0.05.(3)Compared with the group S,the expression of LncRNA AK009271 in the group I/R all decreased at 6,24 and 48 h after reperfusion,and reached the lowest at 24 h after reperfusion(all P<0.05);however,the expression of LncR

关 键 词：脑缺血 再灌注损伤 选择性脑低温 长链非编码RNA 凋亡 

分 类 号：R73[医药卫生—肿瘤]