缺乏针对多种病理过程的有效神经保护剂是缺血性脑卒中治疗的紧迫挑战。克雷巴宁是一种具有优越药理活性的异喹啉类生物碱，可成为神经保护的有前景候选药物。然而，克雷巴宁在缺血性脑卒中中的影响和机制尚不清楚。我们使用大鼠大脑中动脉阻塞再灌注（MCAO/R）模型评估了克雷巴宁对缺血性脑损伤的影响。我们以MCAO/R大鼠和脂多糖（LPS）活化的BV-2细胞为研究对象，探究克雷巴宁的作用机制。我们首先证明了克雷巴宁有效改善了MCAO/R大鼠的神经功能缺陷，同时减轻了脑水肿和梗死。克雷巴宁治疗导致了NeuN+荧光密度的上调和FJB+细胞计数的下调，并缓解了突触损伤。克雷巴宁通过下调NADP+和NADPH水平、抑制gp91phox和p47phox的表达、减少p47phox在Iba-1+ 细胞中的膜转位，从而调控了NADPH氧化酶2（NOX2）的过度活化。此外，克雷巴宁减少了Iba-1+细胞的数量和蛋白表达。相关分析表明，抑制小胶质细胞中NOX2活化有助于减轻I/R脑损伤。此外，克雷巴宁具有显著的抗氧化性能，通过调节体内和体外的过氧化物质和细胞内活性氧自由基的表达，减少脂质和DNA过氧化反应。克雷巴宁还表现出抗炎作用，具体表现为减少NO、白细胞介素-1β、肿瘤坏死因子α、白细胞介素-6和诱导型一氧化氮合酶的表达。克雷巴宁通过抑制核因子κB（NF-κB）和丝裂原活化蛋白激酶（MAPK）信号通路发挥作用。这一结论得到了NF-κB p65启动子活性和核转位的减少、IκBα磷酸化和降解的抑制的支持。此外，它还抑制了ERK、JNK和p38 MAPK的磷酸化。重要的是，在gp91phox和p47phox静默后，克雷巴宁的抗氧化应激和神经炎症效应进一步增强。克雷巴宁通过抑制小胶质细胞中NOX2介导的氧化应激和神经炎症缓解了MCAO/R大鼠的脑损伤，这表明克雷巴宁可能是治疗脑缺血的潜在天然药物。 版权所有©2023 Elsevier GmbH。保留所有权利。

The urgent challenge for ischemic stroke treatment is the lack of effective neuroprotectants that target multiple pathological processes. Crebanine, an isoquinoline-like alkaloid with superior pharmacological activities, presents itself as a promising candidate for neuroprotection. However, its effects and mechanisms on ischemic stroke remain unknown.The effects of crebanine on brain damage following ischemic stroke were evaluated using the middle cerebral artery occlusion and reperfusion (MCAO/R) model. Mechanism of action was investigated using both MCAO/R rats and lipopolysaccharide (LPS)-activated BV-2 cells.We initially demonstrated that crebanine effectively ameliorated the neurological deficits in MCAO/R rats, while also reducing brain edema and infarction. Treatment with crebanine resulted in the up-regulation of NeuN+ fluorescence density and down-regulation of FJB+ cell count, and mitigated synaptic damage. Crebanine attenuated the hyperactivation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (NOX2) by downregulating NADP+ and NADPH levels, suppressing gp91phox and p47phox expressions, and reducing p47phox membrane translocation in Iba-1+ cells. Additionally, crebanine reduced the quantity of Iba-1+ cells and protein expression. Correlation analysis has demonstrated that the inhibition of NOX2 activation in microglia is beneficial for mitigating I/R brain injuries. Moreover, crebanine exhibited significant antioxidant properties by down-regulating the expression of superoxide anion and intracellular reactive oxygen species in vivo and in vitro, and reducing lipid and DNA peroxidation. Crebanine exerted anti-inflammatory effect, as evidenced by the reduction in the expressions of nitric oxide, interleukin 1β, tumor necrosis factor α, interleukin 6, and inducible nitric oxide synthase. The effect of crebanine was achieved through the suppression of nuclear factor-kappa B (NF-κB) and mitogen-activated protein kinases (MAPK) signaling pathway. This is supported by evidence showing reduced NF-κB p65 promoter activity and nucleus translocation, as well as suppressed IκBα phosphorylation and degradation. Additionally, it inhibited the phosphorylation of ERK, JNK, and p38 MAPKs. Importantly, the anti-oxidative stress and neuroinflammation effects of crebanine were further enhanced after silencing gp91phox and p47phox.Crebanine alleviated the brain damages of MCAO/R rats by inhibiting oxidative stress and neuroinflammation mediated by NOX2 in microglia, implying crebanine might be a potential natural drug for the treatment of cerebral ischemia.Copyright © 2023 Elsevier GmbH. All rights reserved.