基于JAK-STAT信号通路研究当归-川芎药对补血、活血作用机制

Under the guidance of the theory of traditional Chinese medicine (TCM) and on basis of four formulae (Foshousan, Shenyantaidongfang, Xiongqiongtang and Xiongguisan) consisted of Danggui (DG) and Chuanxiong (CX), it is proved that the compatibility of DG and CX had synergetic effects for tonifying blood, activating blood circulation and dissipating blood stasis; and different compatibilities (ratios, preparations and dosages) of DG and CX had different effect intensities. We proposed that DG-CX herb pair had the transformation characteristics of mutual assistance and enhancement; and the synergetic effects possibly were achieved through the same pathway closely related to blood deficiency and blood stasis syndromes. On basis of literatures, JAK-STAT signaling pathway had close relationship with blood deficiency and blood stasis. Therefore, the material basis, action and molecular mechanisms of DG-CX with different compatibilities on basis of JAK-STAT signaling pathway from levels of whole animal and cell were researched by using pharmacological and molecular biological methods. In order to further elucidate the efficacy material basis of DG-CX; reveal the scientific connotation of mutual assistant compatibility of DG-CX; provide new targets for prevention and treatment for diseases related to blood deficiency and blood stasis; provide new ideas and methods for explanation the action mechanism of drugs with effects of tonifying blood and activating blood stasis; and provide an effective way to develop the theory of TCM and create modern Chinese medicines.

申请者前期在中医药理论指导下，以当归、川芎配伍组成的佛手散、神验胎动方、芎汤和芎归散四首方为基础，研究发现当归、川芎配伍可协同增强补血、活血功效，且两药味配比、制法和用量不同其整体功效具有明显的偏向性。提出：当归-川芎配伍具有相使转化特征，可能通过与血虚、血瘀均密切相关的作用通路而实现协同增效。文献调研发现，JAK-STAT信号通路与血虚、血瘀均有密切联系。据此，本项目拟采用药理学和分子生物学方法，基于JAK-STAT信号通路从整体动物和细胞层面研究探索当归-川芎不同配伍补血、活血功效物质、作用机理和分子机制，进一步阐明当归-川芎药对的功效物质基础，揭示当归-川芎相使配伍的科学内涵，为血虚、血瘀相关疾病的预防和治疗提供新的靶点，为补血、活血类药物的作用机制阐释提供新的思路与方法，为发展中医药配伍理论和创制现代中药提供有效途径。

当归-川芎药对作为理血药对中的相使药对之一，课题组前期研究发现两药味配伍可协同增强补血、活血功效，且其配比、制法和用量不同其整体功效具有明显的偏向性。通过在整体动物和细胞水平，系统进行归芎药对补血和活血作用的疗效评价及作用机制研究，可为阐释该药对治疗血虚和血瘀相关疾病提供实验支撑。课题组基本按照计划完成预定任务。.确定当归-川芎配伍在临床中的实际应用规律是开展其效应评价和作用机制研究的必要前提。故我们首先对归芎药对在中医临床上的应用特征进行了分析。结果发现：归芎1︰1配比在临床中使用的频次远高于其它配比；当归量<川芎量时，以2︰3配比的应用频次最高；当归量>川芎量时，以3︰2配比的应用频次最高，更强有力支持了本课题的设计方案。.课题组前期已进行了归芎药对对血虚动物血常规和造血器官湿重指数的研究，及其对急性血瘀大鼠血液流变学和凝血功能指标的研究。故本课题①通过归芎药对对慢性失血性血虚大鼠造血和免疫相关细胞因子表达的影响研究，发现：GM-CSF、TNF-α、IL-3、TGF-α和IL-2指标对归芎药对各组补血效应差异的贡献较大；归芎3︰2的先水提后醇提组的总效应最强。②通过归芎药对对急性血瘀大鼠纤溶和黏附系统因子表达的影响研究，发现：t-PA、TXB2、ICAM-1、PAI-1、6-keto-PGF1和PGI2指标对归芎药对各组活血效应差异的贡献较大；GX1︰1和GX0︰1的先水提后醇提组的总效应明显高于其它各给药组。③通过血管组织病理及其主要黏附分子表达变化的研究，发现归芎药对的整体作用稍优于川芎组，当归组的作用最弱。④归芎药对及其组成药味给药后对模型组大鼠PBMCs和血管组织中降低的JAK2、STAT3、ERK和AKT蛋白表达均有升高作用。⑤绿原酸和洋川芎内酯H对HA-VSMCs中AKT、ERK、JAK2和STAT3蛋白表达均有促进作用，但其作用规律不一致，课题组正在进行阿魏酸、咖啡酸、丁基苯酞、丁烯基苯酞等更多芳香酸类和苯酞内酯类成分对上述蛋白表达的影响研究，以期寻找其规律，阐释其物质基础与作用机制的关系。