龙血通络胶囊对血管内皮细胞氧糖剥夺/复氧损伤后细胞凋亡的改善作用

Effect of Longxuetongluo capsule in protecting vascular endothelial cells against oxygen-glucose deprivation/reoxygenation-induced apoptosis

局灶性脑缺血及再灌注损伤是缺血性脑中风的重要发病过程,而缺血再灌注（I/R）损伤诱导的脑血管内皮细胞凋亡是局灶性脑缺血后脑组织损伤的重要诱因。龙血通络胶囊在临床上用于缺血性脑中风恢复期血瘀证的治疗,具有很好的治疗效果,然而其对大脑局灶性缺血后脑组织损伤的保护作用机制目前尚不清晰。该文采用体外氧糖剥夺/复氧（OGD/R）细胞模型模拟血管内皮细胞HUVEC缺血再灌注损伤,通过MTT实验、LDH实验、流式细胞术、AO/EB细胞染色以及Western blot等技术方法观察龙血通络胶囊对OGD/R诱导HUVEC细胞损伤的保护作用,并对其作用机制进行研究。MTT和LDH实验结果显示,龙血通络胶囊能剂量依赖性的提升HUVEC细胞OGD/R损伤后的细胞存活率并降低乳酸脱氢酶（LDH）的释放;流式细胞术与AO/EB细胞染色结果显示,龙血通络胶囊能浓度依赖性的抑制HUVEC细胞OGD/R损伤后的细胞凋亡;Western blot实验结果表明,龙血通络胶囊通过抑制HUVEC细胞caspase 3/9凋亡通路的激活进而抑制OGD/R损伤的细胞凋亡。总之,该研究表明龙血通络胶囊对OGD/R诱导的HUVEC细胞损伤具有明显的保护作用,其作用机制是通过抑制OGD/R诱导线粒体相关caspase 3/9通路的激活而实现的。

Focal cerebral ischemia reperfusion is an essential process during ischemic stroke. The apoptosis of vascular endothelial cells induced by ischemia/reperfusion（ I/R） injury is an important cause for brain injury after focal cerebral ischemia. Longxuetongluo capsule（ LTC） has been used for the treatment of ischemic stroke in clinic. However,its underlying action mechanism is still unclear.This study aimed to verify the protective effect and mechanisms of LTC on HUVEC cells against oxygen-glucose deprivation/reoxygenation（ OGD/R） injury through MTT,LDH,flow cytometry,AO/EB staining and western blot assays. As a result,OGD/R significantly decreased the viability of HUVEC cells,which was significantly improved by LTC. LDH release assay showed that OGD/R significantly increased the lactate dehydrogenase（ LDH） release,and LTC dramatically reduced the OGD/R-induced LDH release. Further mechanism study indicated that LTC dose-dependently inhibited the cleavage of PARP,caspase 3,and caspase 9 induced by OGD/R,suggesting that LTC could inhibit the activation of caspase 3/9 apoptosis pathway in the OGD/R-induced apoptosis of HUVEC cells. In conclusion,LTC could protect HUVEC cells against OGD/R injury by inhibiting the activation of mitochondria-related caspase 3/9 apoptosis pathway.