Introduction

Longxuetongluo Capsule (LTC) is wildly applied to treat ischemic stroke in clinical practice in China. However, the pharmacological mechanism of LTC on ischemic stroke is still unstated.

Objective

Our research was designed to study the protective effect of LTC against cerebral ischemia–reperfusion (I/R) injury and reveal the underlying mechanism both in vivo and in vitro.

Methods

PC12 cells treated with glucose deprivation/reperfusion (OGD/R) were used to simulate in vitro ischemia/reperfusion (I/R) injury. The cell viability, apoptosis rate, and protein expressions of PC12 cells were evaluated. In vivo validation of the protective effect of LTC was carried out by middle cerebral artery occlusion (MCAO)/reperfusion treatment, and the underlying mechanism of its anti-apoptosis ability was further revealed by immunohistochemistry staining and Western blotting.

Results

In the current study, we observed that LTC effectively inhibited oxygen-glucose deprivation/reperfusion (OGD/R) induced apoptosis of PC12 cells through suppressing the cleavage of poly ADP-ribose polymerase (PARP), caspase-3, and caspase-9. Further investigation revealed that OGD/R insult remarkably triggered the endoplasmic reticulum stress responses (ER stress) to induce PC12 cell apoptosis. LTC treatment alleviated OGD/R induced ER stress by inhibiting the activation of protein kinase RNA (PKR)-like ER kinase (PERK)/eukaryotic translation initiation factor 2 (eIF2α) and inositol requiring enzyme 1 (IRE1)/tumor necrosis factor receptor-associated factor 2 (TRAF2) pathways. Additionally, LTC also restrained the OGD/R-induced PC12 cell apoptosis by reversing the activated mitogen-activated protein kinase (MAPK) through IRE1/TRAF2 pathway. Animal studies demonstrated LTC significantly restricted the infarct region induced by middle cerebral artery occlusion (MCAO)/reperfusion, the activation of ER stress and apoptosis of neuronal cells had also been suppressed by LTC in the penumbra region.

Conclusion

LTC protects the cerebral neuronal cell against ischemia/reperfusion injury through ER stress and MAPK-mediated mechanisms.

中文翻译：

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龙血通络胶囊通过内质网应激和MAPK介导的机制保护脑缺血/再灌注损伤

介绍

龙血通络胶囊（LTC）在我国临床上广泛应用于治疗缺血性脑卒中。然而，LTC 治疗缺血性脑卒中的药理机制仍未阐明。

客观的

我们的研究旨在研究 LTC 对脑缺血再灌注 (I/R) 损伤的保护作用，并揭示体内和体外的潜在机制。

方法

用葡萄糖剥夺/再灌注 (OGD/R) 处理的 PC12 细胞用于模拟体外缺血/再灌注 (I/R) 损伤。评估PC12细胞的细胞活力、凋亡率和蛋白表达。通过大脑中动脉闭塞（MCAO）/再灌注治疗对LTC的保护作用进行体内验证，并通过免疫组织化学染色和Western印迹进一步揭示其抗细胞凋亡能力的潜在机制。

结果

在目前的研究中，我们观察到 LTC 通过抑制聚 ADP-核糖聚合酶 (PARP)、caspase-3 和 caspase-9 的切割，有效抑制氧-葡萄糖剥夺/再灌注 (OGD/R) 诱导的 PC12 细胞凋亡。进一步的研究表明，OGD/R 损伤显着触发了内质网应激反应（ER 应激），从而诱导 PC12 细胞凋亡。LTC 治疗通过抑制蛋白激酶 RNA (PKR) 样 ER 激酶 (PERK)/真核翻译起始因子 2 (eIF2α) 和肌醇需要酶 1 (IRE1)/肿瘤坏死因子受体的激活来缓解 OGD/R 诱导的 ER 应激相关因子 2 (TRAF2) 通路。此外，LTC 还通过 IRE1/TRAF2 通路逆转活化的丝裂原活化蛋白激酶 (MAPK) 来抑制 OGD/R 诱导的 PC12 细胞凋亡。

结论

LTC 通过 ER 应激和 MAPK 介导的机制保护脑神经元细胞免受缺血/再灌注损伤。