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. 1997 Jul;65(7):2747–2753. doi: 10.1128/iai.65.7.2747-2753.1997

Pseudomonas aeruginosa lipopolysaccharide binds galectin-3 and other human corneal epithelial proteins.

S K Gupta 1, S Masinick 1, M Garrett 1, L D Hazlett 1
PMCID: PMC175387  PMID: 9199445

Abstract

The aim of this study was to test whether galectin-3 is present in human corneal epithelium and whether lipopolysaccharide (LPS) purified from Pseudomonas aeruginosa ATCC 19660 binds to this animal lectin and/or to another human corneal epithelial protein(s) (HCEP) and to confirm which component of LPS (inner or outer core or lipid A) is important in bacterial binding by using the eye in organ culture. LPS isolated and purified from P. aeruginosa ATCC 19660 and a commercial LPS (serotype 10) differed in polyacrylamide gel analysis but bound similarly to blotted HCEP. Binding was determined to be a receptor-ligand type of interaction by the solid-phase assay, because it was both specific and saturable. Several LPS binding proteins in HCEP were identified by an overlay method. Western blotting with antibody against galectin-3 revealed the presence of this protein in both freshly isolated and cultured transformed human corneal epithelium. Binding inhibition assays showed that antibody specific for the outer core region of LPS and an anti-galectin antibody significantly inhibited bacterial binding in vitro. These data provide further evidence that LPS is an important adhesin of P. aeruginosa, that it binds to protein receptor molecules in HCEP, that one of the LPS binding proteins is galectin-3, and that the outer core portion of the molecule appears to be critical for LPS binding to the eye.

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Selected References

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  1. Aderem A. A., Cohen D. S., Wright S. D., Cohn Z. A. Bacterial lipopolysaccharides prime macrophages for enhanced release of arachidonic acid metabolites. J Exp Med. 1986 Jul 1;164(1):165–179. doi: 10.1084/jem.164.1.165. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Apicella M. A., Griffiss J. M., Schneider H. Isolation and characterization of lipopolysaccharides, lipooligosaccharides, and lipid A. Methods Enzymol. 1994;235:242–252. doi: 10.1016/0076-6879(94)35145-7. [DOI] [PubMed] [Google Scholar]
  3. Barondes S. H., Castronovo V., Cooper D. N., Cummings R. D., Drickamer K., Feizi T., Gitt M. A., Hirabayashi J., Hughes C., Kasai K. Galectins: a family of animal beta-galactoside-binding lectins. Cell. 1994 Feb 25;76(4):597–598. doi: 10.1016/0092-8674(94)90498-7. [DOI] [PubMed] [Google Scholar]
  4. Barondes S. H., Cooper D. N., Gitt M. A., Leffler H. Galectins. Structure and function of a large family of animal lectins. J Biol Chem. 1994 Aug 19;269(33):20807–20810. [PubMed] [Google Scholar]
  5. Barondes S. H. Soluble lectins: a new class of extracellular proteins. Science. 1984 Mar 23;223(4642):1259–1264. doi: 10.1126/science.6367039. [DOI] [PubMed] [Google Scholar]
  6. Beachey E. H. Bacterial adherence: adhesin-receptor interactions mediating the attachment of bacteria to mucosal surface. J Infect Dis. 1981 Mar;143(3):325–345. doi: 10.1093/infdis/143.3.325. [DOI] [PubMed] [Google Scholar]
  7. Beutler B., Cerami A. Cachectin and tumour necrosis factor as two sides of the same biological coin. Nature. 1986 Apr 17;320(6063):584–588. doi: 10.1038/320584a0. [DOI] [PubMed] [Google Scholar]
  8. Cohen E. J., Laibson P. R., Arentsen J. J., Clemons C. S. Corneal ulcers associated with cosmetic extended wear soft contact lenses. Ophthalmology. 1987 Feb;94(2):109–114. doi: 10.1016/s0161-6420(87)33491-8. [DOI] [PubMed] [Google Scholar]
  9. Cryz S. J., Jr, Pitt T. L., Fürer E., Germanier R. Role of lipopolysaccharide in virulence of Pseudomonas aeruginosa. Infect Immun. 1984 May;44(2):508–513. doi: 10.1128/iai.44.2.508-513.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Darveau R. P., Hancock R. E. Procedure for isolation of bacterial lipopolysaccharides from both smooth and rough Pseudomonas aeruginosa and Salmonella typhimurium strains. J Bacteriol. 1983 Aug;155(2):831–838. doi: 10.1128/jb.155.2.831-838.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Dziarski R. Cell-bound albumin is the 70-kDa peptidoglycan-, lipopolysaccharide-, and lipoteichoic acid-binding protein on lymphocytes and macrophages. J Biol Chem. 1994 Aug 12;269(32):20431–20436. [PubMed] [Google Scholar]
  12. Dziarski R. Peptidoglycan and lipopolysaccharide bind to the same binding site on lymphocytes. J Biol Chem. 1991 Mar 15;266(8):4719–4725. [PubMed] [Google Scholar]
  13. Fukuse S., Maeda T., Webb D. R., Devens B. H. A 69-kDa membrane protein associated with lipopolysaccharide (LPS)-induced signal transduction in the human monocytic cell line THP-1. Cell Immunol. 1995 Sep;164(2):248–254. doi: 10.1006/cimm.1995.1168. [DOI] [PubMed] [Google Scholar]
  14. Gillette T. E., Chandler J. W., Greiner J. V. Langerhans cells of the ocular surface. Ophthalmology. 1982 Jun;89(6):700–711. doi: 10.1016/s0161-6420(82)34737-5. [DOI] [PubMed] [Google Scholar]
  15. Gupta S. K., Berk R. S., Masinick S., Hazlett L. D. Pili and lipopolysaccharide of Pseudomonas aeruginosa bind to the glycolipid asialo GM1. Infect Immun. 1994 Oct;62(10):4572–4579. doi: 10.1128/iai.62.10.4572-4579.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Gupta S. K., Masinick S. A., Hobden J. A., Berk R. S., Hazlett L. D. Bacterial proteases and adherence of Pseudomonas aeruginosa to mouse cornea. Exp Eye Res. 1996 Jun;62(6):641–650. doi: 10.1006/exer.1996.0075. [DOI] [PubMed] [Google Scholar]
  17. Halling J. L., Hamill D. R., Lei M. G., Morrison D. C. Identification and characterization of lipopolysaccharide-binding proteins on human peripheral blood cell populations. Infect Immun. 1992 Mar;60(3):845–852. doi: 10.1128/iai.60.3.845-852.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hampton R. Y., Golenbock D. T., Penman M., Krieger M., Raetz C. R. Recognition and plasma clearance of endotoxin by scavenger receptors. Nature. 1991 Jul 25;352(6333):342–344. doi: 10.1038/352342a0. [DOI] [PubMed] [Google Scholar]
  19. Hassman G., Sugar J. Pseudomonas corneal ulcer with extended-wear soft contact lenses for myopia. Arch Ophthalmol. 1983 Oct;101(10):1549–1550. doi: 10.1001/archopht.1983.01040020551008. [DOI] [PubMed] [Google Scholar]
  20. Hazlett L. D., Masinick S., Berk R. S., Zheng Z. Proteinase K decreases Pseudomonas aeruginosa adhesion to wounded cornea. Exp Eye Res. 1992 Oct;55(4):579–587. doi: 10.1016/s0014-4835(05)80171-x. [DOI] [PubMed] [Google Scholar]
  21. Hazlett L. D., Rosen D. D., Berk R. S. Age-related susceptibility to Pseudomonas aeruginosa ocular infections in mice. Infect Immun. 1978 Apr;20(1):25–29. doi: 10.1128/iai.20.1.25-29.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hazlett L., Barrett R., Klettner C., Berk R., Singh A. Pseudomonas aeruginosa: a probe to detail change with age in corneal receptor(s). Ophthalmic Res. 1991;23(3):141–146. doi: 10.1159/000267113. [DOI] [PubMed] [Google Scholar]
  23. Hobden J. A., Gupta S. K., Masinick S. A., Wu X., Kernacki K. A., Berk R. S., Hazlett L. D. Anti-receptor antibodies inhibit Pseudomonas aeruginosa binding to the cornea and prevent corneal perforation. Immunol Cell Biol. 1996 Jun;74(3):258–264. doi: 10.1038/icb.1996.46. [DOI] [PubMed] [Google Scholar]
  24. Homma J. Y. Roles of exoenzymes and exotoxin in the pathogenicity of Pseudomonas aeruginosa and the development of a new vaccine. Jpn J Exp Med. 1980 Jun;50(3):149–165. [PubMed] [Google Scholar]
  25. Kaufmann S. H., Ewing C. M., Shaper J. H. The erasable Western blot. Anal Biochem. 1987 Feb 15;161(1):89–95. doi: 10.1016/0003-2697(87)90656-7. [DOI] [PubMed] [Google Scholar]
  26. Koide S., Steinman R. M. Induction of murine interleukin 1: stimuli and responsive primary cells. Proc Natl Acad Sci U S A. 1987 Jun;84(11):3802–3806. doi: 10.1073/pnas.84.11.3802. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Krivan H. C., Roberts D. D., Ginsburg V. Many pulmonary pathogenic bacteria bind specifically to the carbohydrate sequence GalNAc beta 1-4Gal found in some glycolipids. Proc Natl Acad Sci U S A. 1988 Aug;85(16):6157–6161. doi: 10.1073/pnas.85.16.6157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Kurland J. I., Bockman R. Prostaglandin E production by human blood monocytes and mouse peritoneal macrophages. J Exp Med. 1978 Mar 1;147(3):952–957. doi: 10.1084/jem.147.3.952. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  30. Laibson P. R. Cornea and sclera. Arch Ophthalmol. 1972 Nov;88(5):553–574. doi: 10.1001/archopht.1972.01000030555018. [DOI] [PubMed] [Google Scholar]
  31. Lei M. G., Stimpson S. A., Morrison D. C. Specific endotoxic lipopolysaccharide-binding receptors on murine splenocytes. III. Binding specificity and characterization. J Immunol. 1991 Sep 15;147(6):1925–1932. [PubMed] [Google Scholar]
  32. Liu P. V. Extracellular toxins of Pseudomonas aeruginosa. J Infect Dis. 1974 Nov;130 (Suppl)(0):S94–S99. doi: 10.1093/infdis/130.supplement.s94. [DOI] [PubMed] [Google Scholar]
  33. Lynn W. A., Golenbock D. T. Lipopolysaccharide antagonists. Immunol Today. 1992 Jul;13(7):271–276. doi: 10.1016/0167-5699(92)90009-V. [DOI] [PubMed] [Google Scholar]
  34. Massa S. M., Cooper D. N., Leffler H., Barondes S. H. L-29, an endogenous lectin, binds to glycoconjugate ligands with positive cooperativity. Biochemistry. 1993 Jan 12;32(1):260–267. doi: 10.1021/bi00052a033. [DOI] [PubMed] [Google Scholar]
  35. McEachran D. W., Irvin R. T. Adhesion of Pseudomonas aeruginosa to human buccal epithelial cells: evidence for two classes of receptors. Can J Microbiol. 1985 Jun;31(6):563–569. doi: 10.1139/m85-105. [DOI] [PubMed] [Google Scholar]
  36. Mey A., Leffler H., Hmama Z., Normier G., Revillard J. P. The animal lectin galectin-3 interacts with bacterial lipopolysaccharides via two independent sites. J Immunol. 1996 Feb 15;156(4):1572–1577. [PubMed] [Google Scholar]
  37. Morrison D. C., Lei M. G., Kirikae T., Chen T. Y. Endotoxin receptors on mammalian cells. Immunobiology. 1993 Apr;187(3-5):212–226. doi: 10.1016/S0171-2985(11)80340-2. [DOI] [PubMed] [Google Scholar]
  38. Ofek I., Beachey E. H. Bacterial adherence. Adv Intern Med. 1980;25:503–532. [PubMed] [Google Scholar]
  39. Prasadarao N. V., Wass C. A., Hacker J., Jann K., Kim K. S. Adhesion of S-fimbriated Escherichia coli to brain glycolipids mediated by sfaA gene-encoded protein of S-fimbriae. J Biol Chem. 1993 May 15;268(14):10356–10363. [PubMed] [Google Scholar]
  40. Prpic V., Weiel J. E., Somers S. D., DiGuiseppi J., Gonias S. L., Pizzo S. V., Hamilton T. A., Herman B., Adams D. O. Effects of bacterial lipopolysaccharide on the hydrolysis of phosphatidylinositol-4,5-bisphosphate in murine peritoneal macrophages. J Immunol. 1987 Jul 15;139(2):526–533. [PubMed] [Google Scholar]
  41. Raetz C. R. Biochemistry of endotoxins. Annu Rev Biochem. 1990;59:129–170. doi: 10.1146/annurev.bi.59.070190.001021. [DOI] [PubMed] [Google Scholar]
  42. Raetz C. R., Ulevitch R. J., Wright S. D., Sibley C. H., Ding A., Nathan C. F. Gram-negative endotoxin: an extraordinary lipid with profound effects on eukaryotic signal transduction. FASEB J. 1991 Sep;5(12):2652–2660. doi: 10.1096/fasebj.5.12.1916089. [DOI] [PubMed] [Google Scholar]
  43. Ramphal R., McNiece M. T., Polack F. M. Adherence of Pseudomonas aeruginosa to the injured cornea: a step in the pathogenesis of corneal infections. Ann Ophthalmol. 1981 Apr;13(4):421–425. [PubMed] [Google Scholar]
  44. Reed W. P., Williams R. C., Jr Bacterial adherence: first step in pathogenesis of certain infections. J Chronic Dis. 1978 Feb;31(2):67–72. doi: 10.1016/0021-9681(78)90091-7. [DOI] [PubMed] [Google Scholar]
  45. Rietschel E. T., Kirikae T., Schade F. U., Mamat U., Schmidt G., Loppnow H., Ulmer A. J., Zähringer U., Seydel U., Di Padova F. Bacterial endotoxin: molecular relationships of structure to activity and function. FASEB J. 1994 Feb;8(2):217–225. doi: 10.1096/fasebj.8.2.8119492. [DOI] [PubMed] [Google Scholar]
  46. Rudner X. L., Berk R. S., Hazlett L. D. Immunization with homologous Pseudomonas aeruginosa pili protects against ocular disease. Reg Immunol. 1993 Sep-Oct;5(5):245–252. [PubMed] [Google Scholar]
  47. Rudner X. L., Zheng Z., Berk R. S., Irvin R. T., Hazlett L. D. Corneal epithelial glycoproteins exhibit Pseudomonas aeruginosa pilus binding activity. Invest Ophthalmol Vis Sci. 1992 Jun;33(7):2185–2193. [PubMed] [Google Scholar]
  48. Singh A., Hazlett L., Berk R. S. Characterization of pseudomonal adherence to unwounded cornea. Invest Ophthalmol Vis Sci. 1991 Jun;32(7):2096–2104. [PubMed] [Google Scholar]
  49. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Tsai C. M., Frasch C. E. A sensitive silver stain for detecting lipopolysaccharides in polyacrylamide gels. Anal Biochem. 1982 Jan 1;119(1):115–119. doi: 10.1016/0003-2697(82)90673-x. [DOI] [PubMed] [Google Scholar]
  51. Vasil M. L. Pseudomonas aeruginosa: biology, mechanisms of virulence, epidemiology. J Pediatr. 1986 May;108(5 Pt 2):800–805. doi: 10.1016/s0022-3476(86)80748-x. [DOI] [PubMed] [Google Scholar]
  52. Wilkinson S. G. Composition and structure of lipopolysaccharides from Pseudomonas aeruginosa. Rev Infect Dis. 1983 Nov-Dec;5 (Suppl 5):S941–S949. doi: 10.1093/clinids/5.supplement_5.s941. [DOI] [PubMed] [Google Scholar]
  53. Woods D. E., Iglewski B. H. Toxins of Pseudomonas aeruginosa: new perspectives. Rev Infect Dis. 1983 Sep-Oct;5 (Suppl 4):S715–S722. doi: 10.1093/clinids/5.supplement_4.s715. [DOI] [PubMed] [Google Scholar]
  54. Wright S. D., Kolesnick R. N. Does endotoxin stimulate cells by mimicking ceramide? Immunol Today. 1995 Jun;16(6):297–302. doi: 10.1016/0167-5699(95)80185-5. [DOI] [PubMed] [Google Scholar]
  55. Wright S. D., Levin S. M., Jong M. T., Chad Z., Kabbash L. G. CR3 (CD11b/CD18) expresses one binding site for Arg-Gly-Asp-containing peptides and a second site for bacterial lipopolysaccharide. J Exp Med. 1989 Jan 1;169(1):175–183. doi: 10.1084/jem.169.1.175. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Wright S. D., Ramos R. A., Tobias P. S., Ulevitch R. J., Mathison J. C. CD14, a receptor for complexes of lipopolysaccharide (LPS) and LPS binding protein. Science. 1990 Sep 21;249(4975):1431–1433. doi: 10.1126/science.1698311. [DOI] [PubMed] [Google Scholar]
  57. Wu X., Gupta S. K., Hazlett L. D. Characterization of P. aeruginosa pili binding human corneal epithelial proteins. Curr Eye Res. 1995 Oct;14(10):969–977. doi: 10.3109/02713689508995137. [DOI] [PubMed] [Google Scholar]
  58. Wu X., Kurpakus M., Hazlett L. D. Some P. aeruginosa pilus-binding proteins of human corneal epithelium are cytokeratins. Curr Eye Res. 1996 Jul;15(7):782–791. doi: 10.3109/02713689609003463. [DOI] [PubMed] [Google Scholar]
  59. Zaidi T. S., Fleiszig S. M., Preston M. J., Goldberg J. B., Pier G. B. Lipopolysaccharide outer core is a ligand for corneal cell binding and ingestion of Pseudomonas aeruginosa. Invest Ophthalmol Vis Sci. 1996 May;37(6):976–986. [PubMed] [Google Scholar]
  60. Zoeller R. A., Wightman P. D., Anderson M. S., Raetz C. R. Accumulation of lysophosphatidylinositol in RAW 264.7 macrophage tumor cells stimulated by lipid A precursors. J Biol Chem. 1987 Dec 15;262(35):17212–17220. [PubMed] [Google Scholar]
  61. de Kievit T. R., Lam J. S. Monoclonal antibodies that distinguish inner core, outer core, and lipid A regions of Pseudomonas aeruginosa lipopolysaccharide. J Bacteriol. 1994 Dec;176(23):7129–7139. doi: 10.1128/jb.176.23.7129-7139.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]

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