肾上腺髓质素对局灶性脑缺血/再灌注损伤大鼠神经元凋亡及早期生长反应基因-1的影响

目的探讨肾上腺髓质素（ADM）对局灶性脑缺血/再灌注（I/R）损伤大鼠神经元凋亡、梗死体积及早期生长反应基因-1（Egr-1） mRNA表达的影响,进一步研究ADM在局灶性脑I/R损伤中的作用。方法将54只SD大鼠随机分为假手术组、I/R损伤组以及ADM股静脉组、颈内动脉组和侧脑室组。采用线栓法制备大鼠大脑中动脉（MCA）I/R损伤模型,于阻断血流2h后分别经股静脉、颈内动脉和侧脑室3条途径注射ADM进行干预后再灌注22h。应用氯化三苯四唑（TTC）染色法测定梗死体积,用末端脱氧核苷酸转移酶介导的dUTP缺口末端标记法（TUNEL）检测神经元凋亡,用原位杂交法检测Egr-1 mRNA阳性细胞表达。结果大鼠局灶性脑I/R损伤并经股静脉、颈内动脉、侧脑室注射ADM后,脑梗死体积显著小于I/R损伤组;且颈内动脉和侧脑室注射ADM在减少脑梗死体积方面明显优于股静脉注射ADM（P均〈0.05）。I/R损伤组大鼠缺血侧大脑皮质、海马CA1区凋亡阳性细胞数明显多于假手术组（P均〈0.01）,给予ADM后阳性细胞数显著少于I/R损伤组,以ADM颈内动脉组和侧脑室组更为明显（P均〈0.01）。假手术组大鼠大脑皮质有少量Egr-1 mRNA阳性细胞表达,I/R损伤后缺血侧大脑皮质、海马CA1区Egr-1 mRNA阳性细胞表达多于假手术组（P均〈0.01）,应用ADM的3组大鼠大脑皮质、海马CA1区Egr-1 mRNA阳性细胞的表达明显多于I/R损伤组（P均〈0.01）,但颈内动脉和侧脑室给予ADM组的Egr-1 mRNA阳性细胞表达增高最为明显（P均〈0.01）。结论股静脉、侧脑室和颈内动脉给予外源性ADM能减少神经元凋亡和梗死体积,激活Egr-1 mRNA,对局灶性脑I/R损伤可能有治疗作用。

Effect of adrenomedulin on neuron apoptosis and early growth response gene-1 after focal ischemia/ reperfusion in rats

OBJECTIVE: To explore the influence of adrenomedulin (ADM) on apoptosis of neuron, volume of infarction and the expression of early growth response gene-1 (Egr-1) mRNA in the rat with focal ischemia/reperfusion (I/R) injury. METHODS: Fifty-four SD rats were randomly divided into sham operation group, ADM femoral vein group, internal carotid artery group and lateral cerebral ventricle group. The model was reproduced by ligating the middle cerebral artery (MCA) with a ligature for 2 hours followed by injection of ADM through femoral artery, internal carotid artery and lateral cerebral ventricle before reperfusion for 22 hours. The volume of infarction was estimated with tetrazolium chloride (TTC) staining, apoptosis of the neuron was detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) method, the positive expression of Egr-1 mRNA was detected by in-situ hibridization. RESULTS: The volume of infarction were smaller after the injection of ADM through different ways than that of I/R group. The result was better when the internal carotid artery and the lateral cerebral ventricle were used than that after injection by the way of the femoral vein (both P<0.05). There were few positive cells with TUNEL staining in the cerebral cortex and hippocampus CA1 zone in the sham operation group, and more apoptotic cells were seen in the group with focal brain I/R injury (both P<0.01). After the administration of ADM, especially through the internal carotid artery and the lateral cerebral ventricle, the number of the positive cells with TUNEL staining was decreased obviously compared with I/R group (both P<0.01). There was a little positive expression of Egr-1 mRNA in the cerebral cortex and hippocampus CA1 zone in sham operation group. The expression was enhanced in the group with focal brain I/R injury (both P<0.01). With the injection of ADM, the expression was much more enhanced, especially when internal carotid artery and the lateral cerebral ventricle were used for injection compared with those in I/R group (both P<0.01). CONCLUSION: The injection of ADM through different ways can reduce the neural injury, decrease the apoptosis of the neurons and the volume of the infarction, and increase the expression of Egr-1 mRNA. Therefore, it is efficacious in the treatment of cerebral ischemia.