基于NF-κB通路与GATA-2去泛素化调控间crosstalk探究红景天苷对单核-内皮细胞粘附的影响机制

Lipid regulation still leaves 70% morbidity for atherosclerosis triggered endpoint, that challenges our existing therapeutic regimen. The outstanding effect of Salidroside on anti- atherosclerosis has been widely confirmed while its mechanism is still elusive. Monocyte-endotheliocyte adhesion has been considered to play a pivotal role in the early phase of atherosclerotic pathologic process. And we have found that Salidroside inhibit monocyte-endotheliocyte adhesion via downregulation NF-κB pathway modelated adhesion molecule expression in endotheliocytes. Based on the results from our previous research and bioinformatics analysis, we hypothesized that Salidroside blocks p65 modulated specific gene transcription via interacting with I2PP2A-PP2Ac axis, that not only gets in the way of VCAM-1 transcription and expression but also diminishes CSN5 transcription. Resultant low expression of CSN5 leads to a weak deubiquitinoylation of GATA-2, which in turn further suppresses VCAM-1 transcription. In the present study, Co-IP, ChIP, GST pull down, EMSA, TOP-Flash, western blot and BiFC are applied to investigate the mechanism underlying Salidroside triggerd inhibitory action in monocyte-endotheliocyte adhesion. Unveiling the crosstalk between NF-κB pathway and CSN5 modulated GATA-2 deubiquitinoylation is conducive to our understanding of monocyte-endotheliocyte adhesion, that offers us a new perspective to discover anti- atherosclerosis drugs.

血脂控制后动脉粥样硬化（AS）导致的终点事件残余风险仍高达70%，单纯降脂策略面临巨大挑战。中药活性成分红景天苷对AS疗效已获得广泛证实，但其作用机制尚不明确。单核-内皮细胞粘附是AS早期病理机制中关键环节，课题组前期研究表明红景天苷可通过下调NF-κB通路介导的VCAM-1表达进而抑制单核-内皮细胞粘附作用。结合前期研究基础及生物信息学分析结果，我们提出假说：红景天苷可通过直接作用于I2PP2A-PP2Ac轴，抑制p65转录因子活性，在抑制p65调控的VCAM-1转录同时还可通过CSN5介导的GATA-2去泛素化作用与GATA-2形成crosstalk机制，从而达到对VCAM-1基因转录的双重阻断作用。本项目拟运用Co-IP、ChIP、GST pull down、BiFC等试验探究红景天苷对单核-内皮细胞粘附的影响机制，为抗AS中药新药研发提供理论基础，发现潜在的抗AS治疗新靶点。

中药活性成分红景天苷对AS疗效已获得广泛证实，但其作用机制尚不明确。本项目通过运用Co-IP、ChIP、GST-pull down、BiFC等试验证实红景天苷可通过直接作用于I2PP2A-PP2Ac轴，继而抑制NF-κB亚基p65作为转录因子的转录活性，在直接抑制p65调控的VCAM-1基因转录的同时还可以通过CSN5介导的去泛素化作用与转录因子GATA-2形成crosstalk，从而达到对VCAM-1基因转录的双重阻断，并基于此抑制单核细胞与内皮细胞之间的粘附作用，发挥抗AS生物活性。本项目明确了红景天苷对单核-内皮细胞粘附的影响，探究红景天苷抗AS药理作用与单核-内皮细胞粘附抑制作用间的相关性，夯实红景天苷的抗AS药理作用的理论研究基础，为基于红景天苷的抗AS药物研发提供理论依据和研究策略，为基于中医理论的中药新药研发提供新的思路。本项目还首次证实CSN5介导的蛋白去泛素化机制在单核-内皮细胞粘附过程中的作用，挖掘潜在抗AS药物治疗新靶点，为临床抗AS药物提供新的研发管线和路径。本项目还首次揭示NF-κB通路与GATA-2转录因子间crosstalk机制，深化NF-κB通路精细调控网络，丰富单核-内皮细胞粘附作用的分子机制。