基于eNOS/NO信号通路探讨慢性高原低氧对痰浊证模型大鼠心肺血管内皮稳态的影响机制

Highland people are susceptible to phlegm because of pectoral-qi deficiency, blood stasis caused by rare air and disturbance of body fluid in transportation caused by high fat diet harming the spleen and stomach. Phlegm leads to various cardiovascular diseases by deposition blood vessel and damaging the structure and function of blood vessel. Hyperlipidemia belongs to phlegm turbidity syndrome in traditional Chinese medicine. Hyperlipidemia incidence increased significantly in plateau. Hyperlipidemia and chronic hypoxia causes vascular endothelial injury by eNOS/NO signaling pathways, but the pathogenesis of phlegm turbidity syndrome in the plateau hypoxic environment is unclear. Previous research, we found that the hypoxia combined with high-fat diet caused severe changes in aortic endothelial structures and impaired endothelium-dependent vasorelaxation responses associated with eNOS/NO signaling pathways. Establishing the phlegm turbidity syndrome animal model by high fat diet, study the effects and mechanism of chronic hypoxia on vascular endothelial homeostasis in phlegm turbidity syndrome model rats compared with the normoxic environment phlegm turbidity syndrome. We proposed to vascular endothelial homeostasis-the initiating step of cardiovascular disease as the research target, to cardiopulmonary vascular endothelium-dependent relaxing function, anti platelet aggregation, endothelial monocyte adhesion as index using laser scanning confocal microscope, RT-PCR detection based on eNOS/NO signaling pathway, to explain the pathogenesis of phlegm turbidity syndrome in plateau hypoxic environment, thus provide the early prevention and treatment of cardiovascular disease in plateau and help for making risk factors prevention strategy by traditional Chinese medicine.

高原气薄，宗气亏乏，血行不畅，加之高原人群嗜食肥甘，损伤脾胃，津液运化失常，易生痰浊。痰性粘腻，沉积脉道，损伤血管结构和功能，导致多种心血管疾病发生。高脂血症归于中医痰浊证范畴。高原高脂血症发生率明显增加。高脂血症及慢性低氧影响eNOS/NO信号通路，损伤血管内皮，但高原低氧环境痰浊证致病机理尚不清楚。我们前期研究发现，低氧联合高脂饮食引起大鼠主动脉内皮结构损伤和功能障碍，机制与eNOS/NO信号通路有关。以高脂饮食诱导痰浊证动物模型，与常氧环境痰浊证相比，慢性低氧对痰浊证模型大鼠血管内皮稳态的影响机制如何？我们基于eNOS/NO信号通路，以心血管疾病始动环节—血管内皮稳态为研究靶点，以心肺血管内皮依赖性舒张功能、血管内皮抗血小板聚集、抗单核细胞粘附等为指标，利用激光共聚焦、RT-PCR等手段，阐释低氧环境痰浊证致病机理，为早期防治高原心血管疾病及针对其危险因素制定中医药防治策略提供依据。

项目背景：高原气薄，宗气亏乏，血行不畅，加之高原人群嗜食肥甘，损伤脾胃，津液运化失常，痰浊内生。痰性粘腻，沉积脉道，损伤血管结构和功能，导致多种心血管疾病发生。高脂血症属中医痰浊证。高原高脂血症发生率明显增加。高脂血症及慢性低氧影响eNOS/NO信号通路，损伤血管内皮，但高原痰浊证致病机理尚不清楚，严重制约疾病防治。研究内容：（1）慢性低氧对痰浊证大鼠及EC血管内皮稳态的影响及机制：主动脉、肺、心肌形态学、肺动脉压及血脂、血压、心率；血管内皮依赖性舒张功能；血管内皮抗血小板粘附与抗炎性细胞浸润和吸附；血管平滑肌细胞增殖。（2）慢性低氧对痰浊证大鼠及EC的eNOS/NO信号通路影响：组织及EC的NOS表达、NO；ADMA；活性氧成分及超氧化物歧化酶。重要结果：相比于常氧，低氧环境高脂饮食促发大鼠TCH及LDL水平升高更明显。低氧痰浊证大鼠主动脉、心肌和肺组织结构损伤更严重，主、肺动脉血管内皮依赖性舒张功能明显降低，主动脉、心肌、肺组织及血浆和EC的TF、sVCAM-1、sE-Selectin水平升高，促发血小板、单核细胞与EC粘附，引起血管内皮功能障碍。低氧和痰浊证引起EC氧化压力增加，抗氧化压力降低，引起eNOS mRNA、eNOS蛋白、磷酸化eNOS（Ser1177）表达降低，磷酸化eNOS（Thr495）表达升高，ADMA水平升高，降低NO水平，影响EC活力，以低氧痰浊证组最明显。与细胞实验结果基本一致，低氧和痰浊证也不同程度引起心肌、肺组织和主动脉氧化-抗氧化失衡，引起eNOS/NO改变，导致内皮结构损伤和功能障碍。科学意义：本研究基于eNOS/NO信号通路，以心血管疾病始动环节—血管内皮稳态为研究靶点，以血管内皮依赖性舒张功能，血管内皮抗血小板聚集、抗单核细胞粘附、ADMA等为指标，阐释低氧环境痰浊证致病机理，为早期防治高原心血管疾病、制定中医药防治策略提供依据。