Becker Katrin Anne, Li Xiang, Seitz Aaron, Steinmann Joerg, Koch Anne, Schuchman Edward, Kamler Markus, Edwards Michael J, Caldwell Charles C, Gulbins Erich
Department of Molecular Biology, University Hospital Essen, University of Duisburg-Essen, Essen, Germany.
Department of Pharmacological & Pharmaceutical Sciences, College of Pharmacy, University of Houston, Houston, Texas, USA.
Cell Physiol Biochem. 2017;43(4):1603-1616. doi: 10.1159/000482024. Epub 2017 Oct 17.
BACKGROUND/AIMS: Cystic fibrosis (CF) is dominated by chronic inflammation and infection of the lung resulting in lung destruction and early death of patients. The lungs of CF patients are characterized by a massive accumulation of neutrophils. It requires definition why these massive numbers of neutrophils fail to eliminate typical CF pathogens like Staphylococcus aureus and Pseudomonas aeruginosa (P. aeruginosa) in CF lungs.
We determined ceramide, sphingosine and reactive oxygen species (ROS) in neutrophils from wildtype and CF mice and determined the effect of sphingosine and ROS alone or in combination on killing of different P. aeruginosa strains.
We demonstrate that wildtype neutrophils are able to kill non-mucoid and mucoid clinical P. aeruginosa strains, while neutrophils from CF mice are insufficient to kill these P. aeruginosa strains, although both types of neutrophils infected with P. aeruginosa produce comparable levels of superoxide. All three analyzed P. aeruginosa strains are resistant to reactive oxygen species. The inability of CF neutrophils to kill P. aeruginosa is caused by a marked decrease of surface sphingosine levels in CF neutrophils. Wildtype neutrophils contain much higher concentrations of surface sphingosine than CF neutrophils. Further, wildtype neutrophils, but not CF neutrophils, release sphingosine, most likely as microparticles, upon infection. Sphingosine kills P. aeruginosa in vitro at low micromolar concentrations. Reconstitution of sphingosine in CF neutrophils restores their ability to kill these pathogens, demonstrating the significance of sphingosine for bacterial killing.
The data provide evidence for a new paradigm explaining how neutrophils kill ROS-resistant P. aeruginosa, i.e. by sphingosine that kills P. aeruginosa at low concentrations. This mechanism is defective in CF neutrophils.
背景/目的：囊性纤维化（CF）以肺部慢性炎症和感染为主，导致肺部破坏和患者过早死亡。CF患者的肺部特征是中性粒细胞大量积聚。需要明确为什么在CF患者的肺部，如此大量的中性粒细胞无法清除典型的CF病原体，如金黄色葡萄球菌和铜绿假单胞菌（P. aeruginosa）。
我们测定了野生型和CF小鼠中性粒细胞中的神经酰胺、鞘氨醇和活性氧（ROS），并确定了单独或联合使用鞘氨醇和ROS对不同铜绿假单胞菌菌株杀伤作用的影响。
我们证明野生型中性粒细胞能够杀死非黏液型和黏液型临床铜绿假单胞菌菌株，而CF小鼠的中性粒细胞则不足以杀死这些铜绿假单胞菌菌株，尽管两种类型感染铜绿假单胞菌的中性粒细胞产生的超氧化物水平相当。所有三种分析的铜绿假单胞菌菌株都对活性氧具有抗性。CF中性粒细胞无法杀死铜绿假单胞菌是由于CF中性粒细胞表面鞘氨醇水平显著降低所致。野生型中性粒细胞所含的表面鞘氨醇浓度远高于CF中性粒细胞。此外，野生型中性粒细胞而非CF中性粒细胞在感染后会释放鞘氨醇，很可能是以微粒的形式。鞘氨醇在低微摩尔浓度下就能在体外杀死铜绿假单胞菌。在CF中性粒细胞中重建鞘氨醇可恢复其杀死这些病原体的能力，这证明了鞘氨醇对细菌杀伤的重要性。
这些数据为解释中性粒细胞如何杀死对ROS具有抗性的铜绿假单胞菌提供了新的范例，即通过低浓度杀死铜绿假单胞菌的鞘氨醇。这种机制在CF中性粒细胞中存在缺陷。
