Panjwani N, Zhao Z, Raizman M B, Jungalwala F
New England Eye Center, Tufts University School of Medicine, Boston, MA 02111, USA.
Infect Immun. 1996 May;64(5):1819-25. doi: 10.1128/iai.64.5.1819-1825.1996.
Clinical isolates of Pseudomonas aeruginosa were examined for binding interactions with phospholipids of corneal epithelium. Thin-layer chromatography (TLC) of lipids extracted from corneal epithelia followed by staining with an ammonium molybdate spray reagent revealed three phospholipid components, PL1, PL2, and PL3. The chromatographic mobility of PL1 was similar to that of the phospholipid standards phosphatidylinositol (PI) and phosphatidylserine (PS), which were not well resolved from one other; PL2 and PL3 comigrated with the standards phosphatidylcholine and phosphatidylethanolamine, respectively. By use of a TLC-bacterial overlay procedure, 35S-labeled P. aeruginosa organisms were shown to bind to PL1 but not to PL2 or PL3. P. aeruginosa binding to PL1 was concentration dependent. Alkaline methanolysis abolished the binding. PL1 was separated into two components, PL1-I and PL1-S, by chromatography on borate-treated TLC plates. Both PL1-I and PL1-S contained binding sites for P. aeruginosa. Mass spectral analysis identified PL1-I and PL1-S as PI and PS, respectively. Radiolabeled P. aeruginosa organisms were subsequently shown to bind to commercially available bovine PI and PS and synthetic dipalmitoyl-PS but not to other phospholipid standards, including bovine SM and PC or synthetic dioleoyl- and distearoyl-PC. A control Escherichia coli strain did not bind to either PS or PI. Tetramethylurea, a disrupter of hydrophobic associations, did not influence the binding of P. aeruginosa to PS or PI. P. aeruginosa bound to the monolayers of corneal epithelial cells. P. aeruginosa binding to the monolayer cultures as well as to rabbit corneas pretreated with exogenous PS and PI was significantly higher than that to those preincubated with PC or medium alone. The data suggest that phospholipids PS and PI present in mucus or on the cell surface may function as P. aeruginosa receptors and contribute to selective bacterium-host interactions responsible for initial colonization.
对铜绿假单胞菌的临床分离株进行了与角膜上皮细胞磷脂结合相互作用的检测。从角膜上皮细胞中提取脂质，然后用钼酸铵喷雾试剂染色进行薄层色谱（TLC）分析，结果显示有三种磷脂成分，即PL1、PL2和PL3。PL1的色谱迁移率与磷脂标准品磷脂酰肌醇（PI）和磷脂酰丝氨酸（PS）相似，二者未能很好地分离；PL2和PL3分别与标准品磷脂酰胆碱和磷脂酰乙醇胺迁移至同一位置。通过TLC-细菌覆盖法，发现35S标记的铜绿假单胞菌与PL1结合，但不与PL2或PL3结合。铜绿假单胞菌与PL1的结合具有浓度依赖性。碱性甲醇解可消除这种结合。通过在硼酸盐处理的TLC板上进行色谱分析，PL1被分离为两个成分，即PL1-I和PL1-S。PL1-I和PL1-S均含有铜绿假单胞菌的结合位点。质谱分析确定PL1-I和PL1-S分别为PI和PS。随后发现放射性标记的铜绿假单胞菌与市售牛PI和PS以及合成二棕榈酰-PS结合，但不与其他磷脂标准品结合，包括牛鞘磷脂和磷脂酰胆碱或合成二油酰-和二硬脂酰-磷脂酰胆碱。对照大肠杆菌菌株不与PS或PI结合。疏水缔合破坏剂四甲基脲不影响铜绿假单胞菌与PS或PI的结合。铜绿假单胞菌与角膜上皮细胞单层结合。铜绿假单胞菌与单层培养物以及与用外源性PS和PI预处理的兔角膜的结合显著高于与单独用PC或培养基预孵育的角膜的结合。数据表明，存在于黏液或细胞表面的磷脂PS和PI可能作为铜绿假单胞菌的受体，并有助于介导负责初始定植的选择性细菌-宿主相互作用。
