Leukotriene B4 receptor 1 antagonist in combination with kinase MST1/2 inhibitor has potential for treatment of myocardial infarction

Timely inhibition of inflammation and initiation of resolution are important for the repair of injured tissues. Pharmacological inhibition of mammalian STE20-like protein kinase 1/2 (MST1/2) with XMU-MP-1 might augment tissue regeneration and repair by suppressing apoptosis and increasing cell proliferation. However, MST1 has anti-inflammatory activity, inhibition of which may result in therapy failure. Here, we identified an approach with the potential to overcome this limitation by protecting against cardiac inflammation resulting from inhibition of MST1 in macrophages. We found exacerbation of cardiac dysfunction in LysMCre-mediated Mst1/2-deficient mice after myocardial infarction (MI). This effect was attributed to a shift of macrophage subtypes from those expressing Cxcl2 and Cd163 toward those with Ccl2 and Ccl4 expression. Mass spectrometry identified leukotriene B4 (LTB4) as the lipid mediator that was upregulated in the absence of MST1. We found that MST1 phosphorylated 5-lipoxygenase (5-LOX) at its T218 residue, disrupting the interaction between 5-LOX and 5-LOX-activating protein, and resulting in reduction of LTB4 production. By contrast, a 5-LOXT218A variant showed no response to MST1. Moreover, treatment of peritoneal macrophages with LTB4 or with medium conditioned by Mst1-deficient macrophages resulted in high Ccl2 and Ccl4 expression and low Cxcl2 and Cd163 expression, except when the cells were co-treated with the LTB4 receptor 1 (BLT1) antagonist CP105696. Furthermore, CP105696 ameliorated cardiac dysfunction in LysMCre-mediated Mst1/2-deficient mice and enhanced cardiac repair in wild-type mice treated with XMU-MP-1 after MI. The combination of an MST1/2 inhibitor and a BLT1 antagonist represents a promising strategy for combatting the effects of MI. Overall design: Single cell RNA-Seq of mouse hearts

及时抑制炎症并启动炎症消退程序，对受损组织的修复具有关键意义。采用XMU-MP-1对哺乳动物STE20样蛋白激酶1/2（MST1/2）进行药理学抑制，可通过抑制细胞凋亡、促进细胞增殖，增强组织再生与修复能力。但MST1本身具备抗炎活性，对其进行抑制可能导致治疗失败。在此，我们报道了一种有望克服该局限的策略：通过拮抗巨噬细胞中MST1抑制所诱发的心脏炎症反应。我们发现，在心肌梗死（MI）模型中，LysMCre介导的Mst1/2基因敲除小鼠的心脏功能障碍会进一步加重。该现象的机制为巨噬细胞亚型发生极化偏移：从表达Cxcl2与Cd163的亚型，转向表达Ccl2与Ccl4的亚型。质谱分析发现，在MST1缺失的情况下，白三烯B4（LTB4）作为脂质介质表达上调。我们证实，MST1可在5-脂氧合酶（5-LOX）的T218位点对其进行磷酸化，从而破坏5-LOX与5-脂氧合酶激活蛋白的相互作用，最终减少LTB4的生成。相反，5-LOX的T218A突变体（5-LOXT218A）则不受MST1调控。此外，用LTB4或Mst1基因敲除巨噬细胞的条件培养基处理腹膜巨噬细胞，会使细胞高表达Ccl2与Ccl4、低表达Cxcl2与Cd163；但若同时加入白三烯B4受体1（BLT1）拮抗剂CP105696，则可阻断该效应。进一步实验表明，CP105696可改善LysMCre介导的Mst1/2基因敲除小鼠的心脏功能障碍，并在心肌梗死模型中增强XMU-MP-1处理的野生型小鼠的心脏修复能力。综上，MST1/2抑制剂与BLT1拮抗剂联合使用，有望成为对抗心肌梗死损伤的潜在治疗策略。实验整体设计：小鼠心脏单细胞RNA测序。