基于PPARγ信号通路和脂质组学技术探讨心肌缺血再灌注损伤细胞凋亡机制及芪参益气滴丸干预研究

Inflammatory mediated apoptosis is one of the major pathologic mechanisms of myocardial ischemia-reperfusion injury (MIRI), which can improve MIRI by regulating the inflammatory pathway. Lipid has a regulatory signal transduction function, there is no related researchs that lipid can regulate inflammatory response during the process of MIRI. Based on the result of our preliminary research,our group found that hexadecyl azelaoyl phosphatidylcholine (AzPC) was the major lipid marker which had close relationshiop with MIRI by lipidomics technique ,observed the PPARγ gene synchronous expression with AzPC. It suggested that azPC probably could be regulated by PPARγ- mediated inflammatory signaling pathways,and further influenced MIRI.In this study, We will first prepare the cardiomyocyte model of MIRI,treat the cardiomyocyte of MIRI with different concentrations of azPC,observe the interaction relationship between AzPC and the key targets of PPARγ- mediated inflammatory pathways by protein lipid overlay assay and siRNA, clarify its effect on PPARγ- regulated inflammatory signaling pathways.And further in the Qi deficiency and blood stasis rats model of MIRI,we will observe the effect of Qishenyiqi dripping Pills on the related lipid network of azPC and the related targets of PPARγ signaling pathways by lipidomics and other techniques,clarify the mechanisms that Qishenyiqi dripping Pills by interfering with azPC regulate the inflammatory signaling pathways of PPARγ and improve MIRI.Our study will probably provide a new lipid targe for drug treatment in the process of MIRI and also provide the programs that Chinese traditional medicine treat MIRI.

炎症介导的细胞凋亡是心肌缺血再灌注损伤(MIRI)主要病理机制之一，通过调控炎症反应通路可改善MIRI。脂质具有调节信号转导功能，尚无关于脂质调控MIRI炎症反应的报道。前期通过脂质组学发现十六烷基壬二酸基卵磷脂(azPC)是MIRI相关的主要脂质标志物，并观察到其与PPARγ基因同步表达的现象，提示azPC可能通过调控PPARγ介导的炎症通路影响MIRI。本研究拟首先制备MIRI心肌细胞模型，以不同浓度azPC处理，采用脂质-蛋白覆盖法及siRNA等技术观察其与PPARγ相关炎症通路关键靶点的作用关系，明晰其对PPARγ炎症通路影响。进一步在气虚血瘀型MIRI大鼠模型中，利用脂质组学等技术观察芪参益气滴丸对azPC相关脂质网络及PPARγ通路相关靶点表达影响，明确其通过干预azPC调控PPARγ炎症通路改善MIRI的作用机制。研究将为MIRI提供新的脂质干预靶点，并提供中医药防治方案。

炎症介导的细胞凋亡是心肌缺血再灌注损伤(MIRI)主要病理机制之一，通过调控炎症通路可改善MIRI。脂质具有调节信号转导功能，尚无关于脂质调控MIRI炎症反应的报道。本课题首先制备MIRI大鼠模型，应用脂质组学检测血及心肌中脂质代谢标志物，然后制备MIRI心肌细胞模型，应用已确定的脂质标志物，以不同浓度，不同时间刺激MIRI心肌细胞，利用分子生物学技术观察脂质标志物与PPARγ相关炎症通路的作用关系，明晰其对PPARγ炎症通路影响。进一步在MIRI大鼠中，观察芪参益气滴丸对脂质标志物相关脂质网络及PPARγ通路相关靶点的表达影响，明确其干预脂质标志物调控PPARγ炎症通路减轻MIRI细胞凋亡的作用机制。课题研究结果显示：在血清中共检测到28类977种，心脏中共检测到32类1141种脂质分子，在血清脂质中，与假手术组比，模型组PE降低；PIP2升高。与模型组比，芪参益气滴丸中剂量组PIP2降低，CerG2、LPC升高。在心脏的脂质中，与假手术组比，模型组Cer、CerG2、CerG2GNAc1、Co、DG、PC、PE、PIP、So、WE降低；FA、GM3、LPE、LPS、SQDG升高。与模型组比，芪参益气滴丸中剂量组Cer、CerG2GNAc1、Co、LPC、So、WE升高，与模型组差异有显著性，表明在MIRI时，血及心脏的脂质类型发生了改变，芪参益气滴丸中剂量组对血及心脏的脂质亚类均有调节作用。不同浓度的鞘磷脂刺激MIRI心肌细胞实验显示：鞘磷脂能够激活PPARγ表达，从而抑制炎症通路NF-κB、JAK-STAT 等介导的细胞凋亡。MIRI大鼠模型中，芪参益气滴丸能够通过调节脂质标志物组成的脂质网络来调控PPARγ多途径信号通路达到调控细胞凋亡的作用，降低MIRI大鼠心肌梗死面积，揭示了芪参益气滴丸通过调控脂质靶点进而调控PPARγ炎症通路介导心肌细胞凋亡的效应与机制。