Key Laboratory of Resource Biology and Biotechnology in Western China, Ministry of Education, School of Medicine, Northwest University, Xi'an, 710069, China.
Shaanxi Buchang Pharmaceutical Co. Ltd, Xi'an, 710075, China.
J Ethnopharmacol. 2021 Apr 24;270:113809. doi: 10.1016/j.jep.2021.113809. Epub 2021 Jan 12.
Naoxintong (NXT) is a traditional Chinese medicine preparation that is often used in combination with aspirin in the treatment of cardiovascular diseases (CVD). One of the main symptoms of CVD is hypoxic-ischemia (HI). The purpose of this study is to find out the molecular nodes targeted by NXT and its related molecular pathways in vascular repair.
First, human vein umbilical endothelial cells (EA.hy926) were utilized to set up the Oxygen-Glucose Deprivation-Reoxygenation (OGD/R) model and treated with NXT. Cell proliferation, damage and apoptosis were detected by MTT, LDH, and flow cytometry assays. Second, transcriptional responses of OGD/R cells to NXT treatment were investigated. qRT-PCR, western blotting and inhibitor assays were performed. Third, the anti-thrombotic effect of NXT was evaluated by the zebrafish thrombosis model. Morphological observation, histological staining and qRT-PCR assays were implemented on zebrafish model to further observe in vivo the therapeutic effects of NXT on ischemia and thrombosis.
In OGD/R EA.hy926 cells, NXT treatment could reduce ischemic vascular injury, increase cell viability and decrease the proportion of apoptosis. Through RNA-seq analysis, 183 differentially expressed genes (DEGs) were screened with 110 up-regulated genes and 73 down-regulated genes between OGD/R and OGD/R + NXT treated EA.hy926 cells. VEGF and NFκB pathways were enriched. Among these genes, COX2 was identified as one of important targets via which NXT could restore vascular injury. COX2 inhibitor (NS-398), and aspirin, a drug that prevents the development of CVD by targeting COX2, exhibited similar effects to NXT in the treatment of OGD/R EA.hy926 cells. In zebrafish thrombosis model, NXT could attenuate tail venous thrombus and recover the quantity of heart red blood cells. Furthermore, NXT could prevent the formulation of thrombosis and eliminate inflammation in zebrafish by COX2-VEGF/NFκB signaling.
Our studies implicated that NXT could restore HI injury and inhibit thrombosis through COX2-VEGF/NFκB signaling, which is consistent with the molecular target of aspirin. This finding might explain the principle of NXT combined with aspirin in the treatment of cardiovascular diseases.
脑心通（NXT）是一种中药制剂，常用于与阿司匹林联合治疗心血管疾病（CVD）。CVD 的主要症状之一是缺氧缺血（HI）。本研究的目的是确定 NXT 及其相关分子途径在血管修复中的分子靶点。
首先，利用人脐静脉内皮细胞（EA.hy926）建立氧葡萄糖剥夺-复氧（OGD/R）模型，并给予 NXT 处理。通过 MTT、LDH 和流式细胞术检测细胞增殖、损伤和凋亡。其次，研究了 OGD/R 细胞对 NXT 处理的转录反应。进行 qRT-PCR、western blot 和抑制剂实验。第三，通过斑马鱼血栓模型评估 NXT 的抗血栓作用。对斑马鱼模型进行形态观察、组织学染色和 qRT-PCR 检测，进一步观察 NXT 对缺血和血栓形成的体内治疗效果。
在 OGD/R EA.hy926 细胞中，NXT 处理可减轻缺血性血管损伤，增加细胞活力，降低细胞凋亡比例。通过 RNA-seq 分析，在 OGD/R 和 OGD/R+NXT 处理的 EA.hy926 细胞之间筛选出 183 个差异表达基因（DEGs），其中 110 个上调基因和 73 个下调基因。VEGF 和 NFκB 通路被富集。在这些基因中，COX2 被鉴定为 NXT 恢复血管损伤的重要靶点之一。COX2 抑制剂（NS-398）和阿司匹林（一种通过靶向 COX2 预防 CVD 发展的药物）在治疗 OGD/R EA.hy926 细胞方面表现出与 NXT 相似的效果。在斑马鱼血栓模型中，NXT 可减轻尾部静脉血栓形成并恢复心脏红细胞数量。此外，NXT 通过 COX2-VEGF/NFκB 信号通路可防止血栓形成并消除斑马鱼中的炎症。
我们的研究表明，NXT 可通过 COX2-VEGF/NFκB 信号通路恢复 HI 损伤并抑制血栓形成，这与阿司匹林的分子靶点一致。这一发现可能解释了 NXT 与阿司匹林联合治疗心血管疾病的原理。
