Molecular mechanisms of sphingosine-mediated killing of bacteria

我们已经证明，微摩尔甚至纳摩尔浓度的鞘氨醇足以杀死许多病原体，包括铜绿假单胞菌、金黄色葡萄球菌（甚至耐甲氧西林的S。金黄色葡萄球菌[MRSA]）、鲍氏不动杆菌、流感嗜血杆菌、卡他莫拉菌和伯克霍尔德氏菌属，它们是引起肺炎的重要病原体。体内研究表明，鞘氨醇也能杀死铜绿假单胞菌和沙门氏菌。金黄色葡萄球菌在患有囊性纤维化（CF）的小鼠的肺中，对哺乳动物细胞没有可测量的不利影响。在小鼠和小型猪中吸入鞘氨醇（吸入溶液中浓度高达1 mM）以及局部给药鞘氨醇的其他研究证实，鞘氨醇对哺乳动物气管、支气管和鼻的上皮细胞无不良影响。因此，鞘氨醇似乎具有相对选择性的抗菌功能。鞘氨醇含有NH 2基团和OH基团。在中性或微酸性pH下，如在气道和许多上皮细胞表面上发现的，NH 2基团将被质子化，因此带正电。我们假设，鞘氨醇，根据其正电荷，结合带负电荷的磷脂，主要是心磷脂。初步研究证实了这一假设。心磷脂表达于细菌的质膜上，在呼吸过程中具有重要的功能。在哺乳动物细胞中，心磷脂仅在线粒体中表达，但它不存在于细胞质膜中，因此，以胶束形式应用于细胞的鞘氨醇更不容易接近。拟议的研究将调查的假设，鞘氨醇/鞘氨醇胶束结合心磷脂，并诱导心磷脂在细菌质膜的集群，随后形成大的凝胶，甚至晶体样结构，干扰膜的生物物理特性，迅速导致质膜的泄漏和细菌细胞死亡。

We have shown that micromolar or even nanomolar concentrations of sphingosine are sufficient to kill many pathogens, including Pseudomonas aeruginosa, Staphylococcus aureus (even methicillin-resistant S. aureus [MRSA]), Acinetobacter baumannii, Haemophilus influenzae, Moraxella catarrhalis, and Burkholderia species, which are important pathogens causing pneumonia. In vivo studies showed that sphingosine also kills P. aeruginosa and S. aureus in the lungs of mice with cystic fibrosis (CF), with no measurable adverse effects on the mammalian cells. Additional studies of the inhalation of sphingosine in mice and mini-pigs in concentrations as high as 1 mM in the inhalation solution and also of locally administered sphingosine confirmed that sphingosine has no adverse effects on the mammalian epithelial cells of the trachea, bronchi, and nose. Thus, sphingosine seems to have a relatively selective antibacterial function. Sphingosine contains an NH2 group and an OH group. At neutral or slightly acidic pH, as found in airways and on many epithelial cell surfaces, the NH2 group will be protonated and, thus, positively charged. We hypothesize that sphingosine, depending on its positive charge, binds to negatively charged phospholipids, mainly cardiolipin. Preliminary studies confirm this hypothesis. Cardiolipin is expressed in bacterial plasma membranes and has very important functions in respiration. In mammalian cells cardiolipin is expressed only in mitochondria, but it is absent from the cell plasma membrane and, thus, is much less accessible to sphingosine applied to cells in the form of micelles. The proposed studies will investigate the hypothesis that sphingosine/sphingosine micelles bind to cardiolipin and induce a clustering of cardiolipin in the bacterial plasma membrane, with the subsequent formation of large gel- or even crystal-like structures that disturb the biophysical properties of the membrane, rapidly resulting in leakiness of the plasma membrane and bacterial cell death.