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Infection and Immunity, April 2005, p. 2147-2156, Vol. 73, No. 4
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.4.2147-2156.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Surfactant Protein D Is Present in Human Tear Fluid and the Cornea and Inhibits Epithelial Cell Invasion by Pseudomonas aeruginosa

Minjian Ni,1 David J. Evans,1 Samuel Hawgood,2 E. Margot Anders,3 Robert A. Sack,4 and Suzanne M. J. Fleiszig1*

School of Optometry, University of California, Berkeley,1 Department of Pediatrics, Cardiovascular Research Institute, University of California, San Francisco, California,2 Department of Microbiology and Immunology, University of Melbourne, Victoria, Australia,3 College of Optometry, State University of New York, New York, New York4

Received 21 May 2004/ Returned for modification 31 July 2004/ Accepted 22 November 2004

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 ~2 to 5 µg/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.

* Corresponding author. Mailing address: School of Optometry, University of California, Berkeley, CA 94720-2020. Phone: (510) 643-0990. Fax: (510) 643-5109. E-mail: fleiszig{at}socrates.berkeley.edu.

Editor: V. J. DiRita

Infection and Immunity, April 2005, p. 2147-2156, Vol. 73, No. 4
0019-9567/05/$08.00+0     doi:10.1128/IAI.73.4.2147-2156.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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