曲美他嗪通过促进中性粒细胞迁移改善脓毒症心肌病的作用及机制研究

Septic myocardial dysfunction is a fatal and severe complication for sepsis. So far, there was still no specific and targeted therapeutic options for septic myocardial dysfunction. The function of neutrophil impaired during severe sepsis and the failure of neutrophils migration to the infection site, resulted in multiple organ dysfunction and even death in patients . However, up to now, it has not been reported whether the failure of neutrophils migration contributes to the occurrence and development of septic myocardial dysfunction. Our previous studies found trimetazidine (TMZ) dramatically attenuated LPS-induced septic myocardial dysfunction and enhanced recruitment of neutrophils into heart, in turn, reducing myocardial pyroptosis, which indicated that the promoted neutrophils migration to heart was associated with improved cardiac function after TMZ treatment. Moreover, the failure of neutrophils migration was probably due to the down-regulation of neutrophil surface receptor CXCR2 and our previous studies showed TMZ obviously increased CXCR2 expression in neutrophils. Thus, we put forward our hypothesis: TMZ increased CXCR2 expression in neutrophils, which enhanced recruitment of neutrophils into heart, leading to decreased myocardial pyroptosis, finally improving cardiac function. In this project, we used both CLP- (cecal ligation and puncture) and LPS-induced mouse septic myocardial dysfunction model and mouse bone marrow derived neutrophils to investigate the role of TMZ in preventing septic myocardial dysfunction， and further to elucidate the molecular mechanisms or signaling pathways of TMZ in regulating neutrophil migration.

脓毒症心肌病是脓毒症的致命性严重并发症，目前仍缺乏特异性的针对性治疗手段。严重脓毒症时中性粒细胞迁移功能受损，不能有效募集至感染部位，导致多器官功能障碍，甚至引发死亡。然而，中性粒细胞的迁移受损在脓毒症心肌病的发生发展过程中的作用，目前仍未有文献报道。我们前期发现曲美他嗪显著缓解脓毒症心肌功能障碍且促进中性粒细胞募集至心脏，并减轻心肌细胞的焦亡，提示TMZ促进中性粒细胞迁移与改善脓毒症心肌功能障碍有关。而中性粒细胞迁移功能受损可能与其表面受体CXCR2表达下调有关。我们前期也发现，TMZ显著增加中性粒细胞CXCR2的表达。因此我们提出假设：TMZ上调中性粒细胞CXCR2的表达，促进中性粒细胞迁移，减轻心肌细胞焦亡，改善脓毒症心肌功能障碍。本项目利用CLP及LPS诱导的小鼠脓毒症心肌病模型和小鼠骨髓来源的中性粒细胞，研究TMZ对脓毒症心肌病的保护作用和调控中性粒细胞迁移的信号通路及分子机制。

项目背景：. 脓毒症是机体对感染的反应失调而导致危及生命的多器官功能障碍。近年来的临床研究表明，40%-50%脓毒症患者会出现心肌功能障碍，并且脓毒症所导致的心肌功能障碍与脓毒症死亡率呈明显正相关。曲美他嗪（Trimetazidine, TMZ）属于哌嗪类衍生物，目前在临床上主要用于治疗稳定性心绞痛。我们前期成果显示TMZ通过降低巨噬细胞的促炎细胞因子的产生，显著改善脓毒症心肌功能障碍（Br J Pharmacol（ 2016 Feb;173(3):545-61））。.中性粒细胞有效迁移至感染发生部位，是机体抵抗细菌、寄生虫及病毒等病原体引起的感染的第一道防线。中性粒细胞的迁移主要是通过其表面趋化因子受体识别感染部位的高浓度趋化因子实现的，其中CXC趋化因子受体CXCR2（CXC chemokine receptor 2）是介导中性粒细胞趋化、迁移的关键受体。研究表明，严重脓毒症时，中性粒细胞迁移能力显著受损，进一步导致凝血功能紊乱、心功能不全乃至多器官功能衰竭。而促进中性粒细胞的迁移，能够显著改善脓毒症所致的器官功能障碍。.主要研究内容：.1.建立盲肠穿孔结扎（cecal ligation puncture，CLP）及腹腔注射LPS两种脓毒症模型，研究TMZ对脓毒症小鼠心功能、中性粒细胞迁移及心肌细胞焦亡的作用及机制。.2.提取骨髓来源的中性粒细胞，研究TMZ影响中性粒细胞迁移的机制。.重要结果：.1.曲美他嗪预处理及治疗均可缓解LPS和盲肠穿孔结扎术（CLP）诱导的脓毒症心功能障碍，并能促进中性粒细胞迁移至心脏，减轻心肌细胞焦亡。.2.通过构建骨髓移植嵌合体小鼠，发现TMZ作用于骨髓源性细胞，增加脓毒症时中性粒细胞向心肌组织的迁移，进而改善脓毒症心肌病心功能损伤。.3.提取骨髓来源的中性粒细胞，结合生物信息学、生物化学、细胞生物学等方法证明TMZ通过增加Nrf2，进一步促进CXCR2的表达；.4.在小鼠体内阻断CXCR2受体表达后，TMZ促进中性粒细胞迁移至心脏、改善脓毒症心肌功能障碍的作用显著减弱。.科学意义：.本项目揭示了TMZ通过靶向CXCR2分子来促进中性粒细胞迁移至心脏，有效地改善了脓毒症所致心肌功能障碍，为TMZ治疗脓毒症所致心肌功能障碍提供了理论基础。