验证“血瘀证”微观辨证新子证“假说”

Comparing " status of microinflammation, pathology of microvascular, dysfunction of microcirculation (3 types of micro-lesions) "with pathological features of blood stasis, we find that VEC plays a key role among them,then we propose" New sub- syndrome" hypothesis that the three micro-lesions are the early pathological aspects of blood stasis according to microcosmic syndrome? By the guidelines of traditional theory that＂the disease is determined by veins ", we focus our research on the VEC and the RBC AQPs, CRP, MCP-1, the CA II. Western blot, RT-PCR are used to determine "assay micro-inflammation", "VEC injury", "RBC damage stasis" according to the expression of AQPs and its functional parameter in vivo and in vitro respectively. Further, Rehmanniae,Salvia Miltiorrhizae are taken to explore the differential effects upon sub- syndrome , so as to discuss effective substances and mechanism of Chinese herb base on AQPS.Aims : 1 to observe the different characters among the three micro-lesions and the relationship with blood circulation, then to confirm the hypothesis; 2 to find the "activating-cum-cooling blood and alleviating water retention" indications, create the new system of blood stasis Treating; 3 to explore the effective substances of Chinese herbs effect on the the AQP, in purpose of developing Class Ⅰ diuresis medicine. This project could cast new lights in the meaning of Chinese medicine blood stasis theory, which might bring novel evidence for the clinical application.

比照"微炎症状态、微血管病变、微循环障碍（三微病变）"与血瘀证病理特征，VEC居其关键位点，我们提出"三微病变"属血瘀证早期病理环节及微观辨证新子证"假说"？依据"病在脉、调之血，病在血、调之络"理论，紧扣VEC和RBC，以AQPs和炎性等指标为证据，采用Western Blot、RT-PCR、流式细胞术与细胞色谱等法检测"微炎症、VEC损伤、RBC损伤"血瘀证模型大鼠肺、心、肾组织AQPs表达特征与功能变化，验证"生地、丹参"差异效应，参照其成分代谢，结合体外试验，探索中药调节AQP效应成分与机制。旨在：①观察三微病变之差异效应及其与活血的相关性，证实假说；②寻找活血兼"凉血、利水"法应用指征，创建血瘀证辨治新体系；③发现中药作用于AQP最佳效应成分，奠定研发Ⅰ类利水中药基础。本项目结果从微观与整体、结合与转化角度全新拓展中医"血瘀证"理论内涵，诠释"活血"法本质并为其临床新用提供依据。

背景：以“血液”为重心的血瘀证研究已50年余，成果丰硕！但血瘀证不仅关乎“血”、更关联“脉”，应以“血脉”关链新点深化血瘀证研究。据此，依据“病在脉、调之血，病在血、调之络”理论，比照血瘀证与“三微病变（微炎症状态/微循环障碍/微血管病变）”特征，提出血瘀证“亚型（热瘀/水瘀/虚瘀）”假说。研究内容：①寻找血瘀证“亚型--热/水/虚瘀”存在的依据：制备血瘀证三模型，以“法（凉血活血/活血利水/益气活血）与药（生地/丹参/丹参+黄芪）”测证，借助血管内皮细胞及其膜蛋白AQP特异性表达验证之。②探索“水瘀”的分子靶标：通过丹参调节不同部位AQP1表达与其“活血利水”效应的关系，诠释水瘀本质。重要结果与数据：①凉血活血因应微炎症状态：糖尿病血瘀证模型呈现微炎症状态；地黄降炎性因子水平，减轻阴虚与血瘀表象；微炎症状态与凉血活血相关，反证“热瘀”。②活血利水因应微循环障碍：盐酸肾上腺素+冰泳法制备血瘀证模型呈现微循环障碍；丹参下调粘附分子/VEGF等表达，上调肺/肾组织AQP1表达、以减轻肺水肿与利尿，提示“水瘀与活血利水“亚法”存在；微循环障碍与活血利水相关，反证“水瘀”。③益气活血因应微血管病变：去甲肾上腺素法制备慢性血瘀证模型呈现微血管病变；丹参+黄芪2:1配比降低全血黏度及血浆黏度效应最佳、降低ET-1并升高NO、上调肺/肾组织AQP1表达；微血管病变与益气活血相关，反证“虚瘀”。结论：①以不同“法与药”调治三微病变，反证血瘀证“亚型”客观存在。②AQP1可能是“水瘀与活血利水”的分子靶标。③通过丹参调节不同部位AQP1的“消肿与利尿”效应，首次提出活血利水“亚法（活血消肿与活血利尿）”。科学意义：首次验证血瘀证“亚型”并提出活血利水“亚法”，创新中医学“证与法”理论，预示“血瘀证”研究新方向。