Endogenous plasminogen activator expression after embolic focal cerebral ischemia in mice
Endogenous plasminogen activator expression after embolic focal cerebral ischemia in mice
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DOI：
10.1016/s0006-8993(99)01645-5
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发表时间：
1999-08-07
期刊：
影响因子：
2.9
通讯作者：
Chopp, M
中科院分区：
文献类型：
--
作者：
Ahn, MY;Zhang, ZG;Chopp, M
尿激酶型纤溶酶原激活剂（u-PA）和组织型纤溶酶原激活剂（t-PA）在纤溶、细胞迁移、组织破坏、血管生成和组织重塑中发挥重要作用。组织中的 u-PA 和 t-PA 活性受到纤溶酶原激活剂抑制剂-1 (PAI-1) 的严格调节。然而，人们对缺血性脑中内源性纤溶酶原激活剂 (PA) 和 PAI-1 的活性知之甚少。为了评估脑缺血损伤是否诱导内源性 PA 和 PAI-1，我们测量了脑匀浆中的 PA 活性，并检查了体重 29-35 g 的 C57BL/6J 小鼠 (n = 45) 脑匀浆中 t-PA mRNA、u-PA mRNA 和 PAI-1 mRNA 的表达，其中大脑中动脉 (MCA) 被富含纤维蛋白的凝块闭塞。对照非缺血小鼠 (n = 4) 和 MCA 闭塞 (MCAO) 后 2 小时 (n = 5)、4 小时 (n = 5) 和 24 小时 (n = 4) 的小鼠制备脑匀浆用于直接酪蛋白酶谱分析。此外，MCAO 后 4 小时（n = 2）的 u-PA 和 t-PA 敲除小鼠用作直接酪蛋白酶谱分析的阴性对照。用于原位酶谱分析的冰冻切片是从对照小鼠 (n = 2) 和凝块闭塞后 2 小时、4 小时和 24 小时的小鼠 (n = 2，每个时间点) 获得的。制备脑匀浆用于逆转录聚合酶链反应 (RT-PCR)，以检查对照非缺血小鼠 (n = 4) 和 MCAO 后 2 小时 (n = 5)、4 小时 (n = 5) 和 24 小时 (n = 5) 的小鼠的 t-PA mRNA、u-PA mRNA 和 PAI-1 mRNA 表达。通过直接酪蛋白酶谱分析，与非缺血小鼠相比，缺血半球中风后 4 小时（P < 0.05）和 24 小时（P < 0.05）u-PA 活性增加。缺血脑中t-PA的活性与对照组没有显着差异。通过原位酶谱测定，PA 活性（很可能是 u-PA）存在于缺血半球中。通过 RT-PCR 发现，与对照小鼠相比，中风后 2 小时、4 小时和 24 小时缺血半球中 PAI-1 mRNA 的表达分别增加了 3 倍、15 倍和 25 倍，而 u-PA mRNA 和 t-PA mRNA 则没有。这项研究表明，中风后小鼠大脑中的 PAI-1 mRNA 和 u-PA 活性增加。 (C) 1999 Elsevier Science B.V. 保留所有权利。
Urokinase-type plasminogen activator (u-PA) and tissue-type plasminogen activator (t-PA) play important roles in fibrinolysis, cell migration, tissue destruction, angiogenesis and tissue remodeling. u-PA and t-PA activity in tissue are tightly regulated by plasminogen activator inhibitor-1 (PAI-1). However, little is known of the activity of endogenous plasminogen activators (PAs) and PAI-1 in ischemic brain. To evaluate whether cerebral ischemic injury induces endogenous PAs and PAI-1, we measured PA activity from brain homogenates, and examined the expression of t-PA mRNA, u-PA mRNA and PAI-1 mRNA from brain homogenates in C57BL/6J mice (n = 45) weighing 29-35 g in which the middle cerebral artery (MCA) was occluded by a fibrin-rich clot. Brain homogenates were prepared for direct casein zymography from control non-ischemic mice (n = 4) and mice at 2 h (n = 5), 4 h (n = 5), and 24 h (n = 4) after MCA occlusion (MCAO). Also, u-PA and t-PA knockout mice at 4 h (n = 2, each) after MCAO were used as a negative control for direct casein zymography. Frozen sections for in situ zymography were obtained from control mice (n = 2) and mice at 2 h, 4 h, and 24 h (n = 2, per time point) after clot occlusion. Brain homogenates were prepared for reverse transcriptase-polymerase chain reaction (RT-PCR) to examine t-PA mRNA, u-PA mRNA and PAI-1 mRNA expression from control non-ischemic mice (n = 4) and mice at 2 h (n = 5), 4 h (n = 5), and 24 h (n = 5) after MCAO. By direct casein zymography, u-PA activity increased at 4 h (P < 0.05), and 24 h (P < 0.05) after stroke in the ischemic hemisphere compared with the non-ischemic mice. Activity of t-PA in ischemic brain was not significantly different from the control group. As measured by in situ zymography, PA activity, most likely u-PA, was present in the ischemic hemisphere. By RT-PCR, expression of PAI-1 mRNA, but not u-PA mRNA and t-PA mRNA, increased 3-, 15- and 25-folds in the ischemic hemisphere at 2 h, 4 h and 24 h after stroke, respectively, compared with control mice. This study demonstrates that PAI-1 mRNA and u-PA activity increase in mouse brain after stroke. (C) 1999 Elsevier Science B.V. All rights reserved.