脑缺血中12/15-脂氧合酶对过氧化物酶体增殖物激活受体γ调节作用的初步探讨  

The regulatory effect of 12/15-lipoxygenase on peroxisome proliferator-activated receptors γ in rats with cerebral ischemia

作　　者：徐艳炜[1] 孙莉[2] 梁浩[2] 程焱[1] 

机构地区：[1]天津医科大学总医院神经内科,300052 [2]天津市神经病学研究所

出　　处：《中华神经科杂志》2014年第2期123-127,共5页Chinese Journal of Neurology

基　　金：国家自然科学基金资助项目（81070968）

摘　　要：目的 观察大鼠局灶性脑缺血再灌注损伤中12/15-脂氧合酶表达及活性的改变,并就12/15-脂氧合酶对过氧化物酶体增殖物激活受体γ（peroxisome proliferator-activated receptors γ,PPARγ）的调节作用进行初步探讨.方法 将健康雄性SD大鼠用随机数字表法随机分为假手术组（n=12）、缺血再灌注组（n=18）、干预组（n=12）,制作大鼠大脑中动脉阻塞60 min再灌注24 h模型（MCAO/R）,干预组于MCAO/R前30 min给予12/15-脂氧合酶的产物15-羟基二十碳四烯酸（15-hydroxyeicosatetraenoic acid,15-HETE）.缺血60 min、再灌注24 h后,分别采用蛋白质印迹法和免疫荧光法检测缺血皮质中12/15-脂氧合酶、PPARγ的表达,通过ELISA法测定12/15-脂氧合酶产物15-HETE的表达水平.结果 （1）与假手术组相比,蛋白质印迹检测显示,缺血再灌注组12/15-脂氧合酶蛋白表达增高,差异有统计学意义（假手术组：0.458±0.026,缺血再灌注组：0.688±0.063,t=-8.195,P〈0.05）;免疫荧光检测显示12/15-脂氧合酶与PPARγ在缺血脑组织中具有定位一致性,即共表达性;酶联免疫检测显示,缺血再灌注组12/15-脂氧合酶产物15-HETE的含量（ng/mg）明显增加,差异有统计学意义（假手术组：31.202±2.188,缺血再灌注组：59.402±4.579,t=-12.787,P〈0.05）;（2）与缺血再灌注组相比,蛋白质印迹检测显示12/15-脂氧合酶产物15-HETE干预组PPARγ总蛋白（缺血再灌注组：1.584±0.116,15-HETE干预组：3.826±0.198,t=-23.884）表达增加,核蛋白（缺血再灌注组：12.167±1.131,15-HETE干预组：25.726±1.843,t=-15.360）表达增加、浆蛋白（缺血再灌注组：2.394±0.373,15-HETE干预组：鼠1.184±0.342,t=5.860）表达降低（即PPARγ活性增加）,差异均有统计学意义（均P〈0.05）.结论 脑缺血再灌注损伤时12/15-脂氧合酶表达和产物增加,12/15-脂氧合酶可能通过其产物对PPARγ表达和活性具有调节作用.Objective To examine the expression and activity of 12/15-1ipoxygenase（12/15-LOX） in rats exposed to cerebral ischemia-reperfusion （I/R）, and investigate the regulatory effect of 12/15-LOX on peroxisome proliferator-activated receptors γ （PPARγ ）. Methods Adult male Sprague-Dawley rats underwent 60 min middle cerebral artery occlusion followed by a 24-hour reperfusion （MCAO/R） and were treated with either vehicle （ I/R group ） or 15-hydroxyeicosatetraenoic （ 15-HETE ） group 30 min before operation. According to digital table method, rats were randomly separated into every group. There were 12 rats of Sham-operated group and 15-HETE group, 18 rats of I/R group. After 60 min ischemia and 24 h reperfusion, immunoflorescene staining and western blot were used to evaluate the expression of 12/15-LOX and PPARγin ischemia cortex. The content of 15-HETE was evaluated by enzyme immunoassay. Results （1）Western blot showed that the expression of 12/15-LOX whole protein was enhanced in I/R group compared with sham-operated group （ I/R group ： 0. 688 ± 0. 063 vs sham-operated group ： 0. 458 ± 0. 026,t =-8. 195,P 〈 0.05 ）. Immunoflorescence staining revealed that the localization of 12/15-LOX was consistent with PPARγ in ischemia brain. Enzyme immunoassay found the content of 15-HETE in 1/R group was dramatically increased compared with sham-operated group （ I/R group ： （ 59. 402 ± 4. 579 ） ng/mg vs sham-operated group ： （31. 202 ± 2. 188） ng/mg, t = - 12. 787, P 〈 0. 05 ）. （2） 15-HETE （ 12/15-LOX metabolite） treatment dramatically increased I/R-induced expression of PPARγ total protein （I/R group： 1. 584 ± 0. 116,15-HETE-group ： 3. 826 ± 0. 198, t = - 23. 884, P 〈 0. 05 ） and nuclear translocation （ I/R group：12. 167 ± 1. 131,15-HETE-group： 25. 726 ± 1. 843, t = - 15. 360, P 〈 0.05 ）, and suppressed PPARγ cytoplasmic retention （ that means the enhancement of PPARγ activity, I/R group ： 2. 394 ± 0. 373, 15-HETE-group：1. 18

关 键 词：脑缺血 再灌注损伤 花生四烯酸盐12-脂氧合酶 花生四烯酸盐15-脂氧合酶 羟基花生四烯酸类 PPARΓ 

分 类 号：R743.3[医药卫生—神经病学与精神病学]