人参大黄药对配伍“标本兼治”对局灶性脑缺血/再灌注损伤小鼠模型神经血管单元的保护作用及其分子机制

The project is based on traditional Chinese medicine theory, and puts forward the therapeutical rule of "treating both symptoms and root cause in acute condition" for stroke. The prescription with synergy of couplet medicines of Ginseng and rhubarb(SCGR) is chosen as the target prescription to clarify the protection effect of SCGR on neurovascular unit of mice model with cerebral ischemia /reperfusion injury, especially the effect to promote angiogenesis and neurogenesis, and thus to open new research ideas and method for the treatment of acute ischemic stroke with traditional Chinese medicine. Meanwhile, aquaporin (AQP)-4 knockout mice and connexin (Cx) 43 RNA interference technology are used to reveal that AQP4 and Cx43 and their interactions are involved in cerebral neurovascular unit protection and regulation of angiogenesis and neurogenesis in brain after stroke, and to provide potential targets for the treatment of diseases associated with neurovascular injury. In addition, the project is aimed to clarify the function mechanism of SCGR is possibly related with AQP4, Cx43 and their interaction, so as to provide reference for the study on the mechanism of TCM on cerebral ischemic injury in the future. Eventually, to verify the hypothesis of this project: SCGR may work through more comprehensive protective effect on the neurovascular unit, especially promoting angiogenesis and neurogenesis, to promote recovery of neurological function. The molecular mechanism may be related with AQP4, Cx43 and their interactions.

该项目立足传统中医理论，提出中风病“急则标本兼治”的治则，以人参大黄药对配伍组成复方作为目标方剂，阐明人参大黄药对配伍对局灶性脑缺血/再灌注损伤小鼠模型神经血管单元的保护作用，尤其是促进血管生成和神经再生作用，为中医药治疗急性缺血性中风开辟新的思路和方法。同时，采用水通道蛋白(AQP)4基因敲除小鼠和缝隙连接蛋白（Cx）43 RNA干扰技术，揭示AQP4与Cx43及其相互作用参与脑梗死后神经血管单元的保护及脑内血管生成和神经再生的调节，为神经血管损伤相关疾病的治疗提供潜在的作用靶点。此外，将阐明人参大黄药对配伍的作用机制可能与AQP4和Cx43及其相互作用有关，为今后研究中医药对脑缺血损伤的机制提供参考。最终，以验证课题假说：人参大黄药对配伍可能通过对脑梗死后神经血管单元更加全面的保护作用，尤其是促血管生成和神经再生，促进神经功能的恢复，其分子机制可能与AQP4和Cx43及其相互作用有关。

项目的背景：缺血性中风是当前严重危害人类生命与健康的主要公共卫生问题。该项目立足传统中医理论，提出中风病“急则标本兼治”的治则，以人参大黄药对配伍组成复方作为目标方剂，阐明其对局灶性脑缺血/再灌注（I/R）损伤模型神经保护作用及其机制，为中医药治疗缺血中风提供潜在的治疗靶点和开辟新的研究思路。.主要研究内容：(1) 探讨人参大黄对脑I/R损伤大鼠神经保护作用的配伍比例；(2) 人参大黄对脑I/R损伤大鼠的神经保护作用；(3) 人参大黄对脑I/R损伤大鼠神经保护作用的机制；(4) 人参大黄组分配伍对脑I/R损伤的神经保护作用；(5) 人参大黄组分配伍对脑I/R损伤神经保护作用的机制。.重要结果：(1) 人参酒大黄2:1配伍改善脑I/R损伤大鼠神经功能障碍和脑梗死面积作用最为明显；(2) 人参大黄配伍较单药更能减轻神经功能障碍、减小脑梗死面积、降低脑梗死灶周Cx43和AQP4 的表达；(3) 缝隙连接阻滞剂甘珀酸可降低Cx43的表达，同时下调AQP4，人参大黄配伍干预可进一步减轻神经功能障碍和减小脑梗死面积；(4)人参皂苷Rg1或人参皂苷Rb1和大黄素或大黄酚组分配伍对脑I/R损伤大鼠较单独活性成分更显著改善神经功能缺损、梗死体积和血脑屏障通透性，且显著减少AQP4和Cx43的表达；(5)大黄素和人参皂苷GRg1配伍较单独组使用更能减轻神经功能障碍，减小脑梗死面积，降低炎症因子水平，抑制HMGB的细胞移位，同时降低HMGB1和TLR4的表达。.关键数据及其科学意义：项目阐明了人参大黄药对配伍及其组分配伍对脑梗死后更显著的神经保护作用，为中医药治疗缺血性脑血管病开辟新的研究思路，为中风病“急则标本兼治”治则提供实验依据；同时项目阐明人参大黄药对配伍对脑梗死后神经保护作用的治疗学机制可能与 AQP4 和 Cx43 及其相互作用有关，为今后研究中医药对脑缺血损伤后的机制研究提供参考。