血红素结合蛋白通过上调HO-1改善大鼠脑缺血再灌注后神经行为学

Hemopexin in HO-1 up-regulation for improving rat neuroethology after cerebral ischemia reperfusion injury

目的探讨血红素结合蛋白（HPX）是否通过上调血红素氧合酶-1（HO-1）改善大鼠脑缺血再灌注（IR）损伤。方法 60只雄性SD大鼠,7~8周龄,体重250~280 g,采用随机数字表法分为五组：假手术（Sham）组、缺血再灌注（MCAO）组、溶剂（叠氮钠）（MCAO＋Vehicle）组、HPX（MCAO＋HPX）组和HO-1抑制剂组（MCAO＋HPX＋ZnppⅨ）组,每组12只。采用大脑中动脉栓塞（MCAO）法制备大鼠缺血再灌注模型,MCAO组、MCAO＋Vehicle组、MCAO＋HPX组、MCAO＋HPX＋ZnppⅨ组大鼠分别在IR即刻经侧脑室单次注射生理盐水（10μl,0.9%）、叠氮钠（10μl,0.1%）、HPX（10μl,1.86 g/L）和10μl HPX＋ZnppⅨ混合液（HPX 1.86 g/L＋叠氮钠0.1%）。IR后24 h、7 d分别采用Carcia评分法评价大鼠神经行为学评分（NBS）,采用实时荧光定量聚合酶链式反应（RT-PCR）法检测半暗带区脑组织中HO-1 m RNA水平。结果 IR后24 h Sham组、MCAO组、MCAO＋Vehicle组、MCAO＋HPX组、MCAO＋HPX＋ZnppⅨ组大鼠的NBS评分分别为（15.50±2.43）、（5.17±1.60）、（5.00±2.10）、（10.83±1.47）、（5.33±1.97）分,比较差异有统计学意义（F=34.819,P〈0.05）;IR后7 d Sham组、MCAO组、MCAO＋Vehicle组、MCAO＋HPX组、MCAO＋HPX＋ZnppⅨ组大鼠的NBS评分分别为（16.83±1.60）、（8.83±3.31）、（9.17±3.19）、（12.50±3.08）、（9.33±2.07）分,比较差异有统计学意义（F=13.113,P〈0.05）。与Sham组相比,MCAO组大鼠IR后24 h、7 d缺血半暗带区脑组织HO-1 m RNA水平显著增加（P〈0.05）;与MCAO组相比,MCAO＋Vehicle组大鼠IR后24 h、7 d缺血半暗带区脑组织HO-1 m RNA水平无明显变化（P〉0.05）;与MCAO＋Vehicle组大鼠相比,MCAO＋HPX组大鼠IR后24 h、7 d缺血半暗带区脑组织HO-1 m RNA水平显著升高（P〈0.05）。结论 HPX可通过上调HO-1表达改善大鼠脑缺血再灌注后神经行为学表现。

Objective To investigate whether or not hemopexin（HPX） can improve rat cerebral ischemia reperfusion（IR） injury by its heme oxygenase-1（HO-1） up-regulation. Methods A total of 60 male SD rats, aging 7~8 weeks and weighting 250~280 g, were divided by random number table into sham operation（Sham） group, ischemia reperfusion（MCAO） group, solvent（sodium azide）（MCAO＋Vehicle） group, HPX（MCAO＋HPX） group and HO-1 inhibitor（MCAO＋HPX＋ ZnppⅨ） group, with 12 rats in each group. Middle cerebral artery occlusion（MCAO） was applied to prepare model of rat ischemia reperfusion. MCAO group, MCAO＋Vehicle group, MCAO＋HPX group and MCAO＋HPX＋Znpp Ⅸ group received respectively single injection through paracele immediate at IR by normal saline（10 μl, 0.9%）, sodium azide（10 μl, 0.1%）, HPX（10 μl, 1.86g/L） and 10 μl HPX＋Znpp Ⅸ（HPX 1.86 g/L＋ sodium azide 0.1%）. Carcia evaluation was applied in 24 h and 7 d after IR for neurobehavior score（NBS）, along with real-time fluorescent quantitative polymerase chain reaction（RTPCR） for HO-1 m RNA level in ischemic penumbra brain tissue. Results In 24 h after IR, Sham group, MCAO group, MCAO＋Vehicle group, MCAO＋HPX group and MCAO＋HPX＋Znpp Ⅸ group had NBS scores respectively as（15.50±2.43）,（5.17±1.60）,（5.00±2.10）,（10.83±1.47） and（5.33±1.97） points, and the difference had statistical significance（F=34.819, P〈0.05）. In 7 d after IR, Sham group, MCAO group, MCAO＋Vehicle group, MCAO＋HPX group and MCAO＋HPX＋ZnppⅨ group had NBS scores respectively as（16.83±1.60）,（8.83±3.31）,（9.17±3.19）,（12.50±3.08） and（9.33±2.07） points. Their difference had statistical significance（F=13.113, P〈0.05）. Comparing with Sham group, MCAO group had obviously higher HO-1m RNA level in ischemic penumbra brain tissue after 24 h and 7 d of IR（P〈0.05）. Comparing with MCAO group, MCAO＋Vehicle group had no obvious change in HO-1 m RNA level in ischemic penumbra brain tissue after 24 h and 7 d of IR（P〉0.05）. Comparing with MCAO＋Vehicle group, MCAO＋HPX group had remarkably higher HO-1 m RNA level in ischemic penumbra brain tissue after 24 h and 7 d of IR（P〈0.05）. Conclusion HPX can improve rat neurobehavioral manifestation after ischemia reperfusion by its HO-1 up-regulation.