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The immunogenicity of the liposome-associated outer membrane proteins (OMPs) of Moraxella catarrhalis

Abstract

The outer membrane proteins (OMPs) are the most immunogenic and attractive of the Moraxella catarrhalis vaccine antigens that may induce the protective immune response. The aim of this study was to determine the effectiveness of two types of OMP-associated phosphatidylcholine (PC) liposomal formulations (OMPs-PC, PC-OMPs) and of Zwittergent-based proteomicelles (OMPs-Z) in potentiating an anti-OMP systemic immune response in mice. The immunogenicities of the above preparations were evaluated by assessing serum anti-OMP IgG and IgA reactivity in the post-immunized mouse antisera using ELISA and Western blotting. Additionally, the cross-reactivity of the most effective anti-OMP response was determined using heterologous sera from both humans and mice. Both the proteoliposomes and the proteomicelles showed high immunogenic properties and did not elicit any distinct quantitative differences in the antibody titer or qualitative differences in the pattern of the mouse antisera. The post-immunized mouse antisera predominantly recognized a 60-kDa OMP of M. catarrhalis. That protein was also found to be a highly cross-reactive antigen interacting with a panel of pooled mouse antisera produced by immunization either with whole cells or the purified OMPs of heterologous M. catarrhalis strains. Furthermore, normal sera collected from healthy children were found to be preferentially reactive with the 60-kDa OMP. The serum-specific IgG, IgA and IgM were respectively detected via immunoblotting in 90%, 85% and 30% of heterologous human sera. This similar immunogenic effectiveness of both OMP-associated liposomal formulations could contribute to the practical use of such formulations in the future in human vaccination. Moreover, the highly cross-reactive 60-kDa OMP seems to be an important antigenic marker of M. catarrhalis, and, as it is responsible for the induction of an antibody-mediated and long-lasting immune response, studying it may partially aid us in understanding the relatively low degree of pathogenicity of the bacterium in immunocompetent individuals.

Abbreviations

AFM:

atomic force microscopy

BSA:

bovine serum albumin

EDTA:

ethylenediaminetetracetic acid

ELISA:

enzyme-linked immunosorbent assay

HRP:

horseradish peroxidase

LOS:

lipooligosaccharide

OMPs:

outer membrane proteins

OMPs-PC and PC-OMPs:

the studied proteoliposomes

OMPs-Z:

Zwittergent-based micelles of OMPs

OPD:

o-phenylenediamine

PBS:

phosphate buffered saline

PC:

phosphatidylcholine

TPBS:

PBS supplemented with Tween 20

References

  1. Karaulus, R. and Campagnari, A. Moraxella catarrhalis: a review of an important human mucosal pathogen. Microb. Infect. 2 (2000) 547–559.

    Article  Google Scholar 

  2. Murphy, T.F., Brauer, A.L., Aebi, Ch. and Sethi, S. Identification of surface antigens of Moraxella catarrhalis as targets of human serum antibody responses in chronic obstructive pulmonary disease. Infect. Immun. 73 (2005) 3471–3478.

    Article  PubMed  CAS  Google Scholar 

  3. Murphy, T.F., Brauer, A.L., Grant, B.J. and Sethi, S. Moraxella catarrhalis in chronic obstructive pulmonary disease: burden of disease and immune response. Am. J. Respir. Crit. Care. Med. 172 (2005) 195–199.

    Article  PubMed  Google Scholar 

  4. Sethi, S., Sethi, R., Eschberger, K., Lobbins, P., Cai, X., Grant, B.J. and Murphy, T.F. Airway bacterial concentrations and exacerbations of chronic obstructive pulmonary disease. Am. J. Respir. Crit. Care Med. 176 (2007) 356–361.

    Article  PubMed  Google Scholar 

  5. Verduin, C.M. Hol, C. Fleer, A. van Dijk, H. and van Belkum, A. Moraxella catarrhalis: from emerging to established pathogen. Clin. Microb. Rev. 15 (2002) 125–144.

    Article  Google Scholar 

  6. Bandak, S.M., Turnak, M.R., Allen, B.S., Bolzon, L.D., Preston, D.A., Bouchillon, S.K. and Hoban, D.J. Antibiotic susceptibility among recent clinical isolates of Haemophilus influenza and Moraxella catarrhalis from fifteen countries. Eur. J. Clin. Microbiol. Infect. Dis. 20 (2001) 55–60.

    Article  PubMed  CAS  Google Scholar 

  7. Klingman, K.L., Pye, A., Murphy, T.F. and Hill, S.L. Dynamics of respiratory tract colonization by Branhamella catarrhalis in bronchiectasis. Am. J. Respir. Crit. Care. Med. 152 (1995) 1072–1078.

    PubMed  CAS  Google Scholar 

  8. Faden, H., Duffy, L., Wasielewski, R., Wolf, J., Krystofik, D. and Tung, Y. Relationship between nasopharyngeal colonization and the development of otitis media in children. J. Infect. Dis. 175 (1997) 1440–1445.

    Article  PubMed  CAS  Google Scholar 

  9. Yokota, S., Harimaya, A., Sato, K., Somekawa, Y., Himi, T. and Fujii, N. Colonization and turnover of Streptococcus pneumonia, Haemophilus influenzae, and Moraxella catarrhalis in otitis-prone children. Microbiol. Immunol. 51 (2007) 223–230.

    PubMed  CAS  Google Scholar 

  10. Heiniger, N., Spaniol, V., Troller, R., Vischer, M. and Aebi, C. A reservoir of Moraxella catarrhalis in human pharyngeal lymphoid tissue. J. Infect. Dis. 196 (2007) 1080–1087.

    Article  PubMed  CAS  Google Scholar 

  11. Peng, D., Hong, W., Choudhury, B., Carlson, R.W. and Gu, X.X. Moraxella catarrhalis bacterium without endotoxin, a potential vaccine candidate. Infect. Immun. 73 (2005) 7569–7577.

    Article  PubMed  CAS  Google Scholar 

  12. Troncoso, G., Sánchez, S., Criado, M.T. and Ferreirós, C. Analysis of Moraxella catarrhalis outer membrane antigens cross-reactive with Neisseria meningitidis and Neisseria lactamica. FEMS Immunol. Med. Microbiol. 40 (2004) 89–94.

    Article  PubMed  CAS  Google Scholar 

  13. Mleczko, J., Augustyniak, D. and Jankowski, A. Efficiency of oral immunization of mice with Candida albicans and Moraxella catarrhalis heat-killed cells and cross reactivity of induced antibodies. Centr. Eur. J. Immunol. 32 (2007) 185–188.

    CAS  Google Scholar 

  14. Steeghs, L., Kuipers, B., Hamstra, H.J., Kersten, G., Van Alphen, L. and Van der Ley, P. Immunogenicity of outer membrane proteins in a lipopolysaccharide —deficient mutant of Neisseria meningitidis: influence of adjuvants on the immune response. Infect. Immun. 67 (1999) 4988–4993.

    PubMed  CAS  Google Scholar 

  15. Schweizer, M., Hindennach, I., Garten, W. and Henning, U. Major proteins of the Escherichia coli outer cell envelope membrane. Interaction of protein II* with lipopolysaccharide. Eur. J. Biochem. 82 (1978) 211–217.

    Article  PubMed  CAS  Google Scholar 

  16. Bogdanov, M. and Dowhan, W. Lipid-assisted protein folding. J. Biol. Chem. 274 (1999) 36827–36830.

    Article  PubMed  CAS  Google Scholar 

  17. Qi, H.L., Tai, J.Y. and Blake, M.S. Expression of large amounts of neisserial porin proteins in Escherichia coli and refolding of the proteins into native trimers. Infect. Immun. 62 (1994) 2432–2439.

    PubMed  CAS  Google Scholar 

  18. Wetzler, L.M., Blake, M.S. and Gotschlich, E.C. Characterization and specificity of antibodies to protein I of Neisseria gonorrhoeae produced by injection with various protein I-adjuvant preparation. J. Exp. Med. 168 (1988) 1883–1897.

    Article  PubMed  CAS  Google Scholar 

  19. Jolley, K.A., Appleby, L., Wright, J.C., Christodoulides, M. and Heckels, J. Immunization with recombinant Opc outer membrane protein from Neisseria meningitidis: Influence of sequence variation and levels of expression on the bactericidal immune response against meningococci. Infect. Immun. 69 (2001) 3809–3916.

    Article  PubMed  CAS  Google Scholar 

  20. Frézard, F. Liposomes: from biophysics to the design of peptide vaccines. Braz. J. Med. Biol. Res. 32 (1999)181–189.

    Article  PubMed  Google Scholar 

  21. Altin, J.G. and Parish, Ch.R. Liposomal vaccines-targeting the delivery of antigen. Methods 40 (2006) 39–52.

    Article  PubMed  CAS  Google Scholar 

  22. Uchida, T. and Taneichi, M. Clinical application of surface-linked liposomal antigens. Mini Rev. Med. Chem. 8 (2008) 184–192.

    Article  PubMed  CAS  Google Scholar 

  23. Parmar, M.M., Edwards, K. and Madden, T.D. Incorporation of bacterial membrane proteins into liposomes: factors influencing protein reconstitution. Biochim. Biophys. Acta 1421 (1999) 77–90.

    Article  PubMed  CAS  Google Scholar 

  24. Stebelska, K., Wyrozumska, P., Gubernator, J. and Sikorski A.F. Highly fusogenic cationic liposomes transiently permeabilize the plasma membrane of HeLa cells. Cell. Mol. Biol. Lett. 12 (2007) 35–39.

    Article  CAS  Google Scholar 

  25. Gutowicz, J. and Terlecki, G. The association of glycolytic enzymes with cellular and model membranes. Cell. Mol. Biol. Lett. 8 (2003) 667–680.

    PubMed  CAS  Google Scholar 

  26. Murphy, T.F. and Bartos, L.C. Surface-exposed and antigenically conserved determinants of outer membrane proteins of Branhamella catarrhalis. Infect. Immun. 57 (1989) 2938–2941.

    PubMed  CAS  Google Scholar 

  27. Rigaud, J-L. and Lévy, D. Reconstitution of membrane proteins into liposomes. in: Methods in Enzymol. Liposomes, Part B. 372 Elsevier Academic Press, Co. Elseviere Inc., 2003,65–86.

    Article  CAS  Google Scholar 

  28. Rigaud, J.L., Paternoster, M.T. and Bluza, A. Mechanisms of membrane protein insertion into liposomes during reconstitution procedures involving the use of detergents.2. Incorporation of the light-driven proton pump bacteriorhodopsin. Biochemistry 27 (1988) 2677–2688.

    Article  PubMed  CAS  Google Scholar 

  29. Sarwar, J., Campagnari, A.A., Kirkham, C. and Murphy, T. Characterization of an antigenically conserved heat-modifiable major outer membrane protein of Branhamella catarrhalis. Infect. Immun. 60 (1992) 804–809.

    PubMed  CAS  Google Scholar 

  30. Mandrell, R.E. and Zollinger, W.D. Use of a zwitterionic detergent for the restoration of the antibody-binding capacity of electroblotted meningococcal outer membrane proteins. J. Immunol. Met. 67 (1984) 1–11.

    Article  CAS  Google Scholar 

  31. Idänpään-Heikkilä, I., Wahlström, E., Muttilainen, S., Nurminen, M., Käyhty, H., Sarvas, M. and Mäkelä, P.H. Immunization with meningococcal class 1 outer membrane protein produced in Bacillus subtilis and reconstituted in the presence of Zwittergent or Triton X-100. Vaccine 14 (1996) 886–891.

    Article  PubMed  Google Scholar 

  32. Shahum, E. and Thérien, H.M. Effect of liposomal antigens on the priming and activation of the immune system by dendritic cells. Int. Immunopharmacol. 2 (2002) 591–601.

    Article  PubMed  CAS  Google Scholar 

  33. Ignatius, R., Mahnke, K., Rivera, M., Hong, K., Isdell, F., Steinman, R.M., Pope, M. and Stamatatos, L. Presentation of proteins encapsulated in sterically stabilized lioposomes by dendritic cells initiates CD8(+) T-cell response in vivo. Blood 96 (2000) 3505–3513.

    PubMed  CAS  Google Scholar 

  34. Alving, C.R. Immunologic aspects of liposomes: presentation and processing of liposomal protein and phospholipid antigens. Biochim. Biophys. Acta 1113 (1992) 307–322.

    PubMed  CAS  Google Scholar 

  35. Wright, J.C., Wiliams, J.N., Christodoulides, M. and Haeckels, J.E. Immunization with the recombinant PorB membrane prorein induces bactericidal immune response against Neisseria meningitidis. Infect. Immune. 70 (2002) 4028–4034.

    Article  CAS  Google Scholar 

  36. Sprott, G.D., Dicaire, C.J., Gurnani, K., Deschatelets, L.A. and Krishnan, L. Liposome adjuvants prepared from the total polar lipids of Haloferax volcanii, Planococcus spp. and Bacillus firmus differ in ability to elicit and sustain immune responses. Vaccine 22 (2004) 2154–2162.

    Article  PubMed  CAS  Google Scholar 

  37. Witkowska, D., Masłowska, E., Staniszewska, M., Szosto, B., Jankowski, A. and Gamian, A. Enterobacterial 38-kDa outer membrane protein is an age-dependent molecular marker of innate immunity and immunoglobulin deficiency as results from its reactivity with IgG and IgA antibody. FEMS Immunol. Med. Microbiol. 48 (2006) 205–214.

    Article  PubMed  CAS  Google Scholar 

  38. Christiensen, J.J. Moraxella (Branhamella) catarrhalis: clinical, microbiological and immunological features in lower respiratory tract infections. APMIS 107S (1999) 1–36.

    Google Scholar 

  39. Mathers, K., Leinonen, M. and Goldblatt, D. Anibody response to outer membrane proteins of Moraxella catarrhalis in children with otitis media. Pediatr. Infect. Dis. J. 18 (1999) 982–988.

    Article  PubMed  CAS  Google Scholar 

  40. Murphy, T.F., Kirkham, C., Liu, D.F. and Sethi, S. Human immune response to outer membrane protein CD of Moraxella catarrhalis in adults with chronic obstructive pulmonary disease. Infect. Immun. 71 (2003) 1288–1294.

    Article  PubMed  CAS  Google Scholar 

  41. Murphy, T.F., Kirkham, C. and Lesse, A.J. The major heat modifiable outer membrane protein CD is highly conserved among strains of Branhamella catarrhalis. Mol. Microbiol. 10 (1993) 87–97.

    Article  PubMed  CAS  Google Scholar 

  42. Hsiao, C.B., Sethi, S. and Murphy, T.F. Outer membrane protein CD of Branhamella catarrhalis: sequence conservation in strains recovered from the human respiratory tract. Microb. Pathog. 19 (1995) 215–225.

    Article  PubMed  CAS  Google Scholar 

  43. McMichael, J. Vaccines for Moraxella catarrhalis. Vaccine 19 (2001) 101–107.

    Article  Google Scholar 

  44. Yang-Ping, Y., Myers, L.E., McGuinnes, U., Chong, P., Kwok, Y., Klein, M.H. and Harkness, R.E. The major outer membrane protein, CD, extracted from Moraxella (Branhamella) catarrhalis is a potential vaccine antigen that induces bactericidal antibodies. FEMS Immunol. Med. Microbiol. 17 (1997) 187–199.

    Article  Google Scholar 

  45. Meier, P.S., Freiburghaus, S., Martin, A., Heiniger, N., Troller, R. and Aebi, C. Mucosal immune response to specific outer membrane proteins of Moraxella catarrhalis in young children. Pediatr. Infect. Dis. J. 22 (2003) 256–262.

    Article  PubMed  Google Scholar 

  46. Meier, P.S., Heiniger, N., Troller, R. and Aebi, C. Salivary antibodies directed against outer membrane proteins of Moraxella catarrhalis in healthy adults. Infect. Immun. 71 (2003) 6793–6798.

    Article  CAS  Google Scholar 

  47. Holm, M.M., Vanlerberg, S.L., Foley, I.M., Sledjeski, D.D. and Lafontaine, E.R. The Moraxella catarrhalis porin-like outer membrane protein CD is an adhesion for human lung cells. Infect. Immun. 72 (2004)1906–1913.

    Article  PubMed  CAS  Google Scholar 

  48. Liu, D.F., McMichael, J.C. and Baker, S.M. Moraxella catarrhalis outer membrane protein CD elicits antibodies that inhibit CD binding to human mucin and enhance pulmonary clearance of M.catarrhalis in a mouse model. Infect. Immun. 75 (2007) 2818–2825.

    Article  PubMed  CAS  Google Scholar 

  49. Hu, W.G., Berry, J., Chen, J. and Gum, X-X. Exploration of Moraxella catarrhalis outer membrane proteins, CD and UspA, as new carriers for lipooligosaccharide-based conjugates. FEMS Immunol. Med. Microbiol. 41 (2004)109–115.

    Article  PubMed  CAS  Google Scholar 

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Correspondence to Daria Augustyniak.

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Augustyniak, D., Mleczko, J. & Gutowicz, J. The immunogenicity of the liposome-associated outer membrane proteins (OMPs) of Moraxella catarrhalis . Cell Mol Biol Lett 15, 70 (2010). https://doi.org/10.2478/s11658-009-0035-z

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  • DOI: https://doi.org/10.2478/s11658-009-0035-z

Key words

  • Moraxella catarrhalis
  • Outer membrane proteins
  • Proteoliposomes
  • Proteomicelles
  • Anti-OMP antibodies
  • Cross-reactivity
  • Zwittergent