Surfactant protein D is present in human tear fluid and the cornea and inhibits epithelial cell invasion by Pseudomonas aeruginosa
Surfactant protein D is present in human tear fluid and the cornea and inhibits epithelial cell invasion by Pseudomonas aeruginosa
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DOI：
10.1128/iai.73.4.2147-2156.2005
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发表时间：
2005-04-01
影响因子：
3.1
通讯作者：
Fleiszig, SMJ
中科院分区：
文献类型：
--
作者：
Ni, M;Evans, DJ;Fleiszig, SMJ
我们先前已经证明，人泪液在体外和体内保护角膜上皮细胞免受铜绿假单胞菌的侵害，并且这种保护并不依赖于泪液的抑菌活性。我们试图确定导致撕裂的成分。检验的假设是胶原凝集素（胶原钙依赖性凝集素）参与。从健康人受试者中收集反射性泪液，并通过酶联免疫吸附测定（ELISA）和蛋白质印迹法（Western blot）用表面活性蛋白D（SP-D）、SP-A或甘露糖结合凝集素（MBL）抗体检测聚集素含量。用ELISA法检测人反射泪液中的SP-D，而未检测到SP-A和MBL。全泪液和高效液相色谱泪液组分的Western印迹分析证实了SP-D的存在，其中大部分与免疫球蛋白A在同一组分中洗脱。SP-D泪液浓度计算值为2 - 5 μ g/ml。用甘露聚糖缀合的琼脂糖凝胶或抗SP-D抗体耗尽SP-D降低了泪液对铜绿假单胞菌侵袭的保护作用。单独使用重组人或小鼠SP-D减少了铜绿假单胞菌对上皮细胞的侵袭，而没有可检测到的抑菌活性或细菌聚集。免疫荧光显微镜显示SP-D抗体标记在正常的角膜上皮，但不是基因靶向SP-D敲除小鼠。在体外培养的人和小鼠角膜上皮细胞中也检测到SP-D。总之，SP-D存在于人类泪液以及人类和小鼠角膜上皮中。SP-D参与人类泪液对铜绿假单胞菌侵袭的保护。SP-D是否在眼表的其他先天性或适应性免疫应答的调节中发挥其他作用，如在气道中所起的作用，仍有待探索。
We have previously shown that human tear fluid protects corneal epithelial cells against Pseudomonas aeruginosa in vitro and in vivo and that protection does not depend upon tear bacteriostatic activity. We sought to identify the responsible tear component(s). The hypothesis tested was that collectins (collagenous calcium-dependent lectins) were involved. Reflex tear fluid was collected from healthy human subjects and examined for collectin content by enzyme-linked immunosorbent assay (ELISA) and Western blot with antibody against surfactant protein D (SP-D), SP-A, or mannose-binding lectin (MBL). SP-D, but not SP-A or MBL, was detected by ELISA of human reflex tear fluid. Western blot analysis of whole tears and of high-performance liquid chromatography tear fractions confirmed the presence of SP-D, most of which eluted in the same fraction as immunoglobulin A. SP-D tear concentrations were calculated at similar to 2 to 5 mu g/ml. Depletion of SP-D with mannan-conjugated Sepharose or anti-SP-D antibody reduced the protective effect of tears against P. aeruginosa invasion. Recombinant human or mouse SP-D used alone reduced P. aeruginosa invasion of epithelial cells without detectable bacteriostatic activity or bacterial aggregation. Immunofluorescence microscopy revealed SP-D antibody labeling throughout the corneal epithelium of normal, but not gene-targeted SP-D knockout mice. SP-D was also detected in vitro in cultured human and mouse corneal epithelial cells. In conclusion, SP-D is present in human tear fluid and in human and mouse corneal epithelia. SP-D is involved in human tear fluid protection against P. aeruginosa invasion. Whether SP-D plays other roles in the regulation of other innate or adaptive immune responses at the ocular surface, as it does in the airways, remains to be explored.