Klebsiella pneumoniae infection of murine neutrophils impairs their efferocytic clearance by modulating cell death machinery.
Klebsiella pneumoniae infection of murine neutrophils impairs their efferocytic clearance by modulating cell death machinery.
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DOI：
10.1371/journal.ppat.1007338
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发表时间：
2018-10
期刊：
影响因子：
6.7
通讯作者：
Sharma J
中科院分区：
文献类型：
--
作者：
Jondle CN;Gupta K;Mishra BB;Sharma J
中性粒细胞是第一种渗透细胞类型，对于对抗包括肺炎克雷伯菌(Kpn)在内的细菌病原体的肺部感染是必不可少的。在感染或损伤后，需要通过一种被称为胞吐作用的高度调控的过程来清除细胞凋亡，以恢复体内平衡，但关于细菌感染对这一过程的影响知之甚少。在这里，我们证明了KPN感染阻碍了巨噬细胞在体外和体内肺内对中性粒细胞的呼出细胞摄取。感染中性粒细胞的吞噬功能受损的同时，中性粒细胞表面的凋亡标志性磷脂酰丝氨酸(PS)暴露急剧减少，维持质膜内小叶PS的磷脂转运蛋白Flppase的活性增加。同时，药物抑制Flipase活性增强了PS外化，恢复了KPN感染的中性粒细胞的吞噬作用。我们进一步表明，KPN感染干扰了细胞凋亡的激活，而是通过激活中性粒细胞中的坏死性下垂机制激活了非凋亡性的程序性细胞死亡。相应地，RIPK1和RIPK3抑制剂对坏死性下垂的药理抑制在体外恢复了KPN感染的中性粒细胞对泡泡细胞的摄取。重要的是，用坏死下垂抑制剂治疗KPN感染的小鼠可以改善临床前KPN肺炎小鼠模型的体内疾病转归。据我们所知，这是KPN通过调节细胞死亡途径损害中性粒细胞排出功能的首次报道，这可能为这种感染的治疗干预提供新的靶点。由感染性或无菌损伤引起的炎症性疾病通常以死亡或死亡的浸润性细胞的病理性堆积为特征。肺炎克雷伯菌(Kpn)是一种条件致病菌，其引起的肺炎脓毒症具有相似的病因，但其潜在的发病机制尚不清楚。在这里，我们报告了KPN感染颠覆了一种称为胞吐作用的保护性宿主过程，吞噬细胞通过吞噬并清除死亡/死亡细胞，从而化解炎症和感染。我们的结果表明，KPN感染的中性粒细胞通过吞噬作用清除中性粒细胞的效率低于未感染的中性粒细胞。机制研究表明，在感染的中性粒细胞泡泡功能受损的情况下，“吃我”信号磷脂酰丝氨酸(PS)的暴露减少是通过Flppase活性的增加和细胞死亡途径向坏死性下垂的倾斜所致。因此，在肺炎败血症的临床前模型中，通过药物逆转PS暴露，用坏死下垂抑制剂治疗，恢复了KPN感染的中性粒细胞的泡腾清除，并改善了疾病结局。据我们所知，这是KPN通过损害嗜中性粒细胞前泡细胞的凋亡信号和坏死性下垂机制的激活来颠覆中性粒细胞的流出清除的第一个报告。这可能导致针对肺炎败血症的KPN感染和相关炎症的新的治疗靶点。
Neutrophils are the first infiltrating cell type essential for combating pneumoseptic infections by bacterial pathogens including Klebsiella pneumoniae (KPn). Following an infection or injury, removal of apoptotic infiltrates via a highly regulated process called efferocytosis is required for restoration of homeostasis, but little is known regarding the effect of bacterial infection on this process. Here we demonstrate that KPn infection impedes the efferocytic uptake of neutrophils in-vitro and in-vivo in lungs by macrophages. This impaired efferocytosis of infected neutrophils coincides with drastic reduction in the neutrophil surface exposure of apoptosis signature phospholipid phosphatidyserine (PS); and increased activity of phospholipid transporter flippases, which maintain PS in the inner leaflet of plasma membrane. Concomitantly, pharmacological inhibition of flippase activity enhanced PS externalization and restored the efferocytosis of KPn infected neutrophils. We further show that KPn infection interferes with apoptosis activation and instead activates non-apoptotic programmed cell death via activation of necroptosis machinery in neutrophils. Accordingly, pharmacological inhibition of necroptosis by RIPK1 and RIPK3 inhibitors restored the efferocytic uptake of KPn infected neutrophils in-vitro. Importantly, treatment of KPn infected mice with necroptosis inhibitor improved the disease outcome in-vivo in preclinical mouse model of KPn pneumonia. To our knowledge, this is the first report of neutrophil efferocytosis impairment by KPn via modulation of cell death pathway, which may provide novel targets for therapeutic intervention of this infection. Inflammatory diseases caused by infectious or sterile injuries are often characterized by pathological accumulation of dead or dying infiltrating cells. Pneumonic sepsis caused by Klebsiella pneumoniae (KPn), an opportunistic pathogen, has similar etiology, however, the underlying mechanism remains unknown. Here we report that KPn infection subverts a protective host process termed efferocytosis, by which the phagocytic cells engulf and clear dead/dying cells thereby resolving inflammation and infection. Our results show that KPn infected neutrophils are cleared less efficiently via efferocytosis than the uninfected neutrophils. Mechanistic studies implicated a reduced exposure of “eat me” signal phosphatidyleserine (PS) via increased flippase activity and skewing of cell death pathway toward necroptosis in impaired efferocytosis of infected neutrophils. Accordingly, pharmacological reversal of PS exposure by flippase inhibition, treatment with necroptosis inhibitors restored the efferocytic clearance of KPn infected neutrophils, and improved the disease outcome in a preclinical model of pneumonic sepsis. To our knowledge this is the first report of KPn subversion of efferocytic clearance of neutrophils by impairing pro-efferocytic apoptotic signatures and activation of necroptosis machinery. This could lead to novel therapeutic targets against KPn infection and associated inflammation in pneumonic sepsis.
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