腺病毒介导过氧化物酶体增殖物激活受体-γ1基因脑室内转染对大鼠缺血／再灌注脑的保护作用  

The neuroprotection of adenoviral vector-mediated PPAR -γ1 intracerebroventricular transfection against the ischemia/ reperfusion injury in rats

作　　者：唐吉伟[1] 徐军美 钱自亮[1] 

机构地区：[1]湖南省脑科医院麻醉科,长沙410000 [2]中南大学湘雅二医院麻醉科

出　　处：《国际麻醉学与复苏杂志》2012年第1期1-5,21,共6页International Journal of Anesthesiology and Resuscitation

摘　　要：目的研究通过脑室内途径腺病毒载体转染过氧化物酶体增殖物激活受体-γ1（peroxisome proliferater-activated receptor-γ1，PPAR-γ1）到脑的可行性，如果可行则进一步观察其对缺血，再灌注（ischemia／reperfusion，I/R）脑是否具有保护作用。方法选择90只成年雄性SD大鼠，按随机数字表法分成5组（每组18只）。第1组脑室内注入生理盐水0．1ml，第2组注入不含PPAR-γ1基因的空腺病毒载体0．1ml，第3组注入表达PPAR-γ1的腺病毒载体（adeno virusvector encodingPPAR-γ1，Adv-PPAR-γ1）0．1ml，第4组注入腺病毒介导的增强型绿色荧光蛋白（adv encoding enhanced green fluorescence protein，Adv-EGFP）0．1ml，第5组吡格列酮5mg／kg灌胃qd3d。3d后建立脑I／R模型。第1组为假手术组，不阻断大鼠大脑中动脉。第2、3、5组阻断大脑中动脉90min再灌注24h。采集脑组织标本，第4组免疫荧光显微镜下观察腺病毒表达情况；1、2、3、5组进行2，3，5三苯基氯化四氮唑（triphenyltetrazolium chloride，TrC）染色法测量脑梗死体积、伊文氏蓝法测定血脑屏障通透性、干媪重法测定脑含水量、光镜、电镜下进行病理形态学观察；脑组织髓过氧化物酶（myeloperoxidase，MPO）活性测定；用Western Blot方法检测缺血区脑组织白介素-1β（IL-1β）、细胞黏附分子-γ（inter-cellular adhesion mdecule-1，ICAM-1）、水通道蛋白-4（aquapofinprotein，AQP-4）、基质金属蛋白酶-9（matrixmetalloproteinase-9，MMP-9）蛋白的表达。结果脑室内途径腺病毒载体转染Adv-EGFP，免疫荧光反应阳性。脑I／R后，脑梗死体积（42．3±2．6）％、血脑屏障通透性（0．0945±0．0095）、脑组织含水量（87．4±4．7）％、MPO活性明显增高（0．213±0．044），IL-1B（0．84±0．05）、ICAM-1（0．85±0．07）、AQP-4（0．8±0．06）和MMP-9（0．80±0．03）蛋白的表达增加。脑室内注射Adv-PPAR-γ1或吡格�Objective To explore the feasibility of PPAR-γ1 gene transfection via intracerebroventricular injection and possible protective effect against cerebral ischemia reperfusion injury. Methods Ninety adult male SD rats were randomly divided into 6 groups （n=18）. In group Ⅰ , the rats were intracerebroventricularly injected with normal saline 0.1 ml. In group Ⅱ , the rats were injected with empty adenoviral （Adv） vector 0.1 ml. In group m , the rats were injected with 0.1 ml Adv-PPAR-β1 and the group IV received Adv-EGFP 0.1 ml. The group V was intragastrically administered with Pioglitazone. After 3 days, middle cerebral artery occlusion model was established. The group Ⅰ was sham-operation group. In group Ⅱ -Ⅴ middle cerebral artery was obstructed for 90 rain followed by reperfusion for 24 h. Twenty-four hours after operation, samples were collected. The expression of green fluorescent protein in Group Ⅳ was observed with fluorescence microscope to assess the adenovirus-mediated transfection. The cerebral infarction volume was measured by triphenyltetrazolium chloride （Trc） staining. The permeability of blood-brain barrier was determined by Evan＇s blue. Brain water was calculated by wet-dry weight method. The histopathology was observed under light microscope and electron microscope. The activity of myeloperoxidase （MPO） was tested. The expressions of IL-1β, inter-cellular adhesion mdecule-1 （ICAM-1）, aquaporin protein （AQP-4） and matrix metalloproteinase-9 （MMP-9） protein were determined by Western Blotting. Results After the animals were intracerebroventricularly administered Adv-EGFP plasmid, that the fluorescence for EGFP was positive in brain tissue suggested the successful transfection of viral gene. In response to I/R injury, the permeability of blood-brain barrier（0.094 5±0.009 5）, brain water（87.4±4.7）, and the activity of MPO were dramatically increased（0.213±0.044） as welt as the cerebral infarction volume（42.3±2.6）. The expressions of IL-11

关 键 词：过氧化物酶增殖物激活受体-γ1 脑缺血/再灌注 基因转染 

分 类 号：R74[医药卫生—神经病学与精神病学]