摘要
目的构建携带缺氧诱导因子-1α(HIF-1α)基因的重组腺病毒载体,探索HIF-1α对大鼠局灶性脑缺血的治疗作用。方法应用腺病毒表达系统AdEasySystem构建携带缺氧诱导因子-1α和绿色荧光蛋白(GFP)的重组腺病毒(Ad-HIF-1α)并行PCR鉴定。建立大鼠线栓法大脑中动脉缺血再灌注模型,分为Ad-HIF-1α、腺病毒空载体(Ad)、生理盐水(NS)3组;将Ad-HIF-1α、Ad和NS注射到模型鼠缺血侧侧脑室,观察Ad-HIF-1α的分布和绿色荧光持续时间、三组大鼠神经功能缺失评分和2,3,5-三苯基氯化四氮唑(TTC)染色评价Ad-HIF-1α对脑缺血的治疗效果。结果在荧光显微镜下绿色荧光蛋白的表达逐渐向远离侧脑室的部位扩展,在14d时荧光最强,随后逐渐减弱,至28d时荧光基本消失。Ad-HIF-1α治疗组24h大鼠神经功能缺失评分为2.4±0.5,与Ad组(2.6±0.5)和NS组(2.7±0.7)比较差异无统计学意义(P>0.05);48及72hAd-HIF-1α治疗组神经功能缺失评分分别为1.6±0.7和0.9±0.6,与Ad组(分别为2.9±0.6和3.2±0.6)和NS组(分别为3.0±0.7和3.2±0.8)比较差异均有统计学意义(P<0.05);72h时脑组织TTC染色显示Ad-HIF-1α治疗组梗死体积为81.2mm3±1.4mm3,与Ad组(173.9mm3±1.3mm3)和NS组(171.7mm3±6.2mm3)比较差异有统计学意义(P<0.05)。结论Ad-HIF-1α对大鼠局灶性脑缺血具有一定的治疗作用,为HIF-1α基因的进一步研究和临床应用奠定了基础。
Objective To construct a recombinant adenoviral vector carrying HIF-1α gene and explore the therapeutic effect of HIF-1α on focal cerebral ischemia in adult rats. Methods The AdEasy System was used to construct the recombinant adenoviral vector carrying HIF-1 α gene and green fluorescent protein and PCR was used to identify the HIF-1α gene. Middle cerebral artery occlusion（MCAo） and reperfusion models were established and divided into Ad-HIF-1α group,Ad group and NS group. After Ad-HIF-1α, Ad and NS were injected into the ischemic ventricle, expression of Ad-HIF-1α was observed and its therapeutic effect was evaluated by 2,3,5-triphenyltetrazolium chloride （ TTC ） staining and neurological severity scores. Results GFP expression distributed apart from the ventricle and reached a peak at 14 days and persisted for about 4 weeks under fluorescent microscope. The neurological severity scores was 2.4 ± 0.5 at 24 h in Ad-HIF-1α group and there was no statistical significance compared with Ad group（2.6 ± 0. 5） and NS group（2.7 ± 0. 7） （ P 〉 0. 05 ）. The scores were 1.6 ± 0. 7 at 48 h and 0. 9 ± 0. 6 at 72 h in Ad- HIF-1α group, and there were statistical significance compared with Ad group（2.9 ± 0.6 and 3.2 ± 0.6 respectively） and NS group （ 3.0 ± 0.7 and 3.2 ± 0.8 ） （ P 〈 0.05 ）. The infarct volume was 81.2 mm^3 ± 1.4 mm^3 at 72 h in Ad-HIF-1α group and there was statistical significance compared with Ad group （173.9 mm^3 ± 1.3 mm^3） and NS group（171.7 mm^3 ±6.2 mm^3）（p〈0.05）. Conclusion HIF-1α gene had definite therapeutic effect on focal cerebral ischemia in adult rats, which settles a foundation for next HIF-1α gene study and clinic application.
出处
《中华医学杂志》
CAS
CSCD
北大核心
2006年第15期1057-1060,共4页
National Medical Journal of China
基金
重庆市教委基金资助项目(渝科教2002-18号)
