花生四烯酸表氧化酶代谢物EETs抑制NLRP3炎症小体通路改善病毒性心肌炎的作用及机制

Viral myocarditis (VMC) endangers human health seriously, and it lacks specific and efficient treatment. Epoxyeicosatrienoic acids (EETs) are derived from arachidonic acid (AA) metabolized by epoxygenase. We have studied EETs for many years and demonstrated that EETs are significantly protective in myocardium and inflammation. But effects and mechanisms of EETs on VMC have not been elucidated. Preliminary results showed that EETs levels were decreased in plasma of viral myocarditis patients，and EETs treatment significantly inhibited CVB3 induced cardiomyocytes death, inflammatory injury and NLRP3 inflammasome upregulation. Therefore, we hypothesize that EETs may ameliorate viral myocarditis by inhibiting NLRP3 inflammasome signaling. In present project, CVB3 myocarditis model will be established. EETs levels will be elevated via cardiomyocytes CYP2J2 transgene and sEH knockout to investigate protective effects of EETs in VMC. Subsequently, effects and mechanisms of EETs in alleviating VMC by inhibiting NLRP3 inflammasome signaling will be elucidated in cardiomyocytes specific activation of NLRP3 mice. Meanwhile, protective effects and mechanisms of EETs in CVB3 infected cardiomyocytes will be investigated in vitro. This project is significantly important for elucidating the role of EETs in pathophysiology of VMC and developing new therapeutic drugs to this new target.

病毒性心肌炎严重威胁人类健康，且缺乏特异和有效的治疗手段。EETs是花生四烯酸经表氧化酶代谢产生的内源性小分子，我们多年研究证实EETs具有显著的心肌保护和抗炎作用，然而其在病毒性心肌炎中的作用及机制尚未阐明。我们初步证明病毒性心肌炎患者血浆EETs水平降低，EETs显著抑制柯萨奇B3病毒（CVB3）诱导的心肌细胞死亡、炎症性损伤和NLRP3炎症体上调。因此我们假设EETs通过抑制NLRP3通路改善病毒性心肌炎。本项目拟建立CVB3心肌炎模型，利用心肌CYP2J2转基因和sEH敲除等途径上调EETs，研究EETs抗病毒性心肌炎的作用。进一步利用心肌NLRP3激活小鼠研究EETs抑制NLRP3通路抗病毒性心肌炎的作用及机制。同时将在原代心肌细胞中研究EETs对CVB3感染心肌细胞的保护作用及机制。本研究对阐明EETs在病毒性心肌炎中的病理生理作用和未来针对这一新靶点开发新的药物具有重要意义。

病毒性心肌炎严重威胁人类健康，且缺乏特异和有效的治疗手段。EETs是花生四烯酸经表氧化酶代谢产生的内源性小分子，我们多年研究证实EETs具有显著的心肌保护和抗炎作用，然而其在病毒性心肌炎中的作用及机制尚未阐明。我们前期证明病毒性心肌炎患者血浆中EETs水平降低，同时NLRP3炎症小体下游细胞因子的升高，这表明EETs水平降低可能无法有效抑制NLRP3炎症小体活化。因此，我们提出EETs可能通过抑制NLRP3炎症小体活化进而改善病毒性心肌炎。通过本项目的研究，我们首先证明EETs显著降低心肌炎小鼠死亡率、改善心脏功能，显著降低小鼠心脏组织炎症浸润和纤维化程度，抑制IL-1β、IL-18等炎症因子释放。体外实验证明在CVB3干预后EETs促进心肌细胞存活。机制研究表明EETs不仅抑制NLRP3炎症小体组分表达，还抑制NLRP3炎症小体的组装。最后，通过筛选本研究发现S100A9和TRIM56为EETs抑制炎症小体活化的直接靶点，通过分子对接显示11,12-EET可以与TRIM56和S100A9紧密结合。这些结果提示EETs通过直接结合TRIM56和S100A9抑制NLRP3炎症小体的活化，从而减轻心肌炎症性损伤，最终改善病毒性心肌炎。本研究阐明了EETs这一组内源性活性小分子在病毒性心肌炎中的作用和机制，为干预CYP450表氧化酶-EETs-sEH途径治疗病毒性心肌炎提供了理论依据。

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