Department of Pharmacology, Monash University, Clayton, Victoria, Australia.
PLoS One. 2011;6(12):e28393. doi: 10.1371/journal.pone.0028393. Epub 2011 Dec 2.
Post-ischemic oxidative stress and vasomotor dysfunction in cerebral arteries may increase the likelihood of cognitive impairment and secondary stroke. However, the underlying mechanisms of post-stroke vascular abnormalities, as distinct from those causing primary brain injury, are poorly understood. We tested whether augmented superoxide-dependent dysfunction occurs in the mouse cerebral circulation following ischemia-reperfusion, and evaluated the role of Nox2 oxidase.
Cerebral ischemia was induced in male C57Bl6/J wild-type (WT) and Nox2-deficient (Nox2(-/-)) mice by middle cerebral artery occlusion (MCAO; 0.5 h), followed by reperfusion (23.5 h). Superoxide production by MCA was measured by L-012-enhanced chemiluminescence. Nitric oxide (NO) function was assessed in cannulated and pressurized MCA via the vasoconstrictor response to N(ω)-nitro-L-arginine methyl ester (L-NAME; 100 µmol/L). Expression of Nox2, the nitration marker 3-nitrotyrosine, and leukocyte marker CD45 was assessed in cerebral arteries by Western blotting.
Following ischemia-reperfusion, superoxide production was markedly increased in the MCA of WT, but not Nox2(-/-) mice. In WT mice, L-NAME-induced constriction was reduced by ∼50% in ischemic MCA, whereas ischemia-reperfusion had no effect on responses to L-NAME in vessels from Nox2(-/-) mice. In ischemic MCA from WT mice, expression of Nox2 and 3-nitrotyrosine were ∼1.4-fold higher than in the contralateral MCA, or in ischemic or contralateral vessels from Nox2(-/-) mice. Vascular CD45 levels were unchanged by ischemia-reperfusion.
Excessive superoxide production, impaired NO function and nitrosative stress occur in mouse cerebral arteries after ischemia-reperfusion. These abnormalities appear to be exclusively due to increased activity of vascular Nox2 oxidase.
脑动脉的缺血后氧化应激和血管舒缩功能障碍可能增加认知障碍和继发中风的可能性。然而，与原发性脑损伤不同的是，中风后血管异常的潜在机制还了解甚少。我们检测了在缺血再灌注后，小鼠大脑循环中是否会出现增强的超氧化物依赖性功能障碍，并评估了 Nox2 氧化酶的作用。
雄性 C57Bl6/J 野生型（WT）和 Nox2 缺陷（Nox2(-/-)）小鼠通过大脑中动脉闭塞（MCAO；0.5 h）诱导脑缺血，随后再灌注（23.5 h）。通过 L-012 增强化学发光法测量 MCA 中的超氧化物产生。通过 N（ω）-硝基-L-精氨酸甲酯（L-NAME；100 µmol/L）对插管和加压 MCA 的血管收缩反应评估一氧化氮（NO）功能。通过 Western blot 评估 Nox2、硝化标记物 3-硝基酪氨酸和白细胞标记物 CD45 在脑动脉中的表达。
缺血再灌注后，WT 但不是 Nox2(-/-) 小鼠的 MCA 中超氧化物产生明显增加。在 WT 小鼠中，缺血 MCA 中 L-NAME 诱导的收缩减少了约 50%，而缺血再灌注对 Nox2(-/-) 小鼠血管中 L-NAME 的反应没有影响。在 WT 小鼠的缺血 MCA 中，Nox2 和 3-硝基酪氨酸的表达比对侧 MCA 或 Nox2(-/-) 小鼠的缺血或对侧血管高约 1.4 倍。血管 CD45 水平不受缺血再灌注影响。
在缺血再灌注后，小鼠脑动脉中出现过度的超氧化物产生、NO 功能障碍和硝化应激。这些异常似乎完全归因于血管 Nox2 氧化酶活性的增加。
