LBP and CD14 secreted in tears by the lacrimal glands modulate the LPS response of corneal epithelial cells.
OBJECTIVE
Lipopolysaccharide (LPS) is one of the most powerful bacterial virulence factors in terms of proinflammatory properties and is likely to contribute to corneal bacterial keratitis. Better understanding of the spatial expression of the LPS receptor components at the tear-corneal interface might facilitate enhanced functions of the LPS receptor complex in ocular defense against Gram-negative infections.


METHODS
The expression of LPS-binding protein (LBP), CD14, toll-like receptor (TLR)-4, and MD-2 in human lacrimal glands, reflex tears, and corneal epithelia was examined by ELISA, RT-PCR, Western blot analysis, and immunofluorescence. The release of proinflammatory cytokines after the activation of primary and immortalized corneal epithelial cells with LPS and human tears was measured by ELISA.


RESULTS
LBP and CD14 proteins were detected in reflex human tears. Human lacrimal glands and corneal epithelia expressed LBP, CD14, TLR4, and MD-2 mRNAs and proteins. In the corneal epithelium, LBP was mainly expressed by superficial and basal epithelial cells, whereas CD14, TLR4, and MD-2 expression were limited to the wing and basal epithelial cells. In a dose-dependant manner, tear CD14 and LBP mediated the secretion of interleukin (IL)-6 and IL-8 by corneal epithelia cells when challenged with LPS.


CONCLUSIONS
Tear CD14 and LBP complemented the LPS receptor complex expressed by the corneal epithelia to trigger an immune response in the presence of LPS. The complementation of these tear and corneal immune proteins could play an important role in LPS recognition and signaling and, therefore, could modulate ocular innate immunity.
Blais
David R
DR
Department of Biochemistry, Microbiology and Immunology, University of Ottawa, Ontario, Canada.
Vascotto
Sandy G
SG
Griffith
May
M
Altosaar
Illimar
I
D000209:Acute-Phase Proteins / Q000235:genetics / Q000378:metabolism*; D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D019573:Epithelium, Corneal / Q000187:drug effects / Q000378:metabolism*; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism*; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D015850:Interleukin-6 / Q000378:metabolism; D016209:Interleukin-8 / Q000378:metabolism; D007765:Lacrimal Apparatus / Q000378:metabolism*; D018950:Lipopolysaccharide Receptors / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology*; D051218:Lymphocyte Antigen 96 / Q000235:genetics / Q000378:metabolism; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism*; D008875:Middle Aged; D011549:Pseudomonas; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013666:Tears / Q000378:metabolism*; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000378:metabolism; D000099231:Lipopolysaccharide-Binding Protein
D000209:Acute-Phase Proteins; D002352:Carrier Proteins; D005136:Eye Proteins; D015850:Interleukin-6; D016209:Interleukin-8; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D051218:Lymphocyte Antigen 96; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D051197:Toll-Like Receptor 4; D000099231:Lipopolysaccharide-Binding Protein; C495377:LY96 protein, human; C495345:TLR4 protein, human; C042434:tear proteins