M cell–targeting strategy enhances systemic and mucosal immune responses induced by oral administration of nuclease-producing L. lactis

Abstract

Efficient delivery of antigens to the gut-associated lymphoid tissue (GALT) is the most critical step for the induction of mucosal immunity by oral vaccines. As M cells are the main portal for luminal antigens into the GALT, the M cell-targeting of antigens affords a promising strategy toward the development of effective oral vaccines. Lactococcus lactis is a fascinating recombinant host for oral vaccines, as they survive and produce antigens in the gut and have a particularly safe profile for human use. In this study, we developed and evaluated an M cell–targeting oral immunization system using recombinant L. lactis strains. For the purpose, we generated an L. lactis strain that secretes a model antigen fused with the OmpH β1α1 domain of Yersinia enterocolitica, which has been shown to bind to a complement C5a receptor on the M cell surface. As the model antigen, Staphylococcus aureus nuclease was used for fusion, resulting in L. lactis–expressing Nuc-OmpH (LL/Nuc-OmpH). Ex vivo intestinal loop assays showed that the amount of Nuc-OmpH taken up into Peyer’s patches was more than that of the unfused nuclease (Nuc). In addition, oral administration of the recombinant L. lactis strains to mice demonstrated that LL/Nuc-OmpH-induced nuclease-specific fecal IgA and serum IgG titers were significantly higher than those induced by LL/Nuc. These results indicate that OmpH works as an M cell–targeting molecule when fused with antigens secreted from L. lactis and that the M cell–targeting strategy affords a promising platform for L. lactis–based mucosal immunization.

This is a preview of subscription content, log in to check access.

Fig. 1
Fig. 2
Fig. 3

References

  1. Ahmed B, Loos M, Vanrompay D, Cox E (2014) Oral immunization with Lactococcus lactis-expressing EspB induces protective immune responses against Escherichia coli O157: H7 in a murine model of colonization. Vaccine 32:3909–3916. https://doi.org/10.1016/j.vaccine.2014.05.054

    CAS  Article  PubMed  Google Scholar 

  2. Boyaka PN (2017) Inducing mucosal IgA: a challenge for vaccine adjuvants and delivery systems. J Immunol 199:9–16. https://doi.org/10.4049/jimmunol.1601775

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  3. Chen K, Cerutti A (2010) Vaccination strategies to promote mucosal antibody responses. Immunity 33:479–491. https://doi.org/10.1016/j.immuni.2010.09.013

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  4. Davis A, Moore IB, Parker DSTH (1977) Nuclease B. A possible precursor of nuclease A, an extracellular nuclease of Staphylococcus aureus. J Biol Chem 252:6544–6553

    CAS  PubMed  Google Scholar 

  5. Deenick EK, Hasbold J, Hodgkin PD (2005) Decision criteria for resolving isotype switching conflicts by B cells. Eur J Immunol 35:2949–2955. https://doi.org/10.1002/eji.200425719

    CAS  Article  PubMed  Google Scholar 

  6. Dieye Y, Usai S, Clier F, Gruss A, Piard JC (2001) Design of a protein-targeting system for lactic acid bacteria. J Bacteriol 183:4157–4166. https://doi.org/10.1128/JB.183.14.4157-4166.2001

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  7. Hase K, Kawano K, Nochi T, Pontes GS, Fukuda S, Ebisawa M, Kadokura K, Tobe T, Fujimura Y, Kawano S, Yabashi A, Waguri S, Nakato G, Kimura S, Murakami T, Iimura M, Hamura K, Fukuoka S-I, Lowe AW, Itoh K, Kiyono H, Ohno H (2009) Uptake through glycoprotein 2 of FimH(+) bacteria by M cells initiates mucosal immune response. Nature 462:226–230. https://doi.org/10.1038/nature08529

    CAS  Article  PubMed  Google Scholar 

  8. Kim S-H, Jang Y-S (2014) Antigen targeting to M cells for enhancing the efficacy of mucosal vaccines. Exp Mol Med 46:e85. https://doi.org/10.1038/emm.2013.165

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  9. Kim S-H, Seo K-W, Kim J, Lee K-Y, Jang Y-S (2010) The M cell-targeting ligand promotes antigen delivery and induces antigen-specific immune responses in mucosal vaccination. J Immunol 185:5787–5795. https://doi.org/10.4049/jimmunol.0903184

    CAS  Article  PubMed  Google Scholar 

  10. Kim SH, Jung DI, Yang IY, Kim J, Lee KY, Nochi T, Kiyono H, Jang YS (2011) M cells expressing the complement C5a receptor are efficient targets for mucosal vaccine delivery. Eur J Immunol 41:3219–3229. https://doi.org/10.1002/eji.201141592

    CAS  Article  PubMed  Google Scholar 

  11. Kuipers OP, De Ruyter PGGA, Kleerebezem M, De Vos WM (1998) Quorum sensing-controlled gene expression in lactic acid bacteria. J Biotechnol 64:15–21

    CAS  Article  Google Scholar 

  12. Liu L, Zhang W, Song Y, Wang W, Zhang Y, Wang T, Li K, Pan Q, Qi X, Gao Y, Gao L, Liu C, Zhang Y, Wang Y, He G, Wang X, Cui H (2018) Recombinant Lactococcus lactis co-expressing OmpH of an M cell-targeting ligand and IBDV-VP2 protein provide immunological protection in chickens. Vaccine 36:729–735. https://doi.org/10.1016/j.vaccine.2017.12.027

    CAS  Article  PubMed  Google Scholar 

  13. Malhotra N, Kang J (2013) SMAD regulatory networks construct a balanced immune system. Immunology 139:1–10. https://doi.org/10.1111/imm.12076

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  14. Mantis NJ, Cheung MC, Chintalacharuvu KR, Rey J, Corthesy B, Neutra MR (2002) Selective adherence of IgA to murine Peyer’s patch M cells: evidence for a novel IgA receptor. J Immunol 169:1844–1851. https://doi.org/10.4049/jimmunol.169.4.1844

    CAS  Article  PubMed  Google Scholar 

  15. Matsumura T, Sugawara Y, Yutani M, Amatsu S, Yagita H, Kohda T, Fukuoka S-I, Nakamura Y, Fukuda S, Hase K, Ohno H, Fujinaga Y (2015) Botulinum toxin A complex exploits intestinal M cells to enter the host and exert neurotoxicity. Nat Commun 6:6255. https://doi.org/10.1038/ncomms7255

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  16. Mierau I, Kleerebezem M (2005) 10 years of the nisin-controlled gene expression system (NICE) in Lactococcus lactis. Appl Microbiol Biotechnol 68:705–717. https://doi.org/10.1007/s00253-005-0107-6

    CAS  Article  PubMed  Google Scholar 

  17. Misumi S, Masuyama M, Takamune N, Nakayama D, Mitsumata R, Matsumoto H, Urata N, Takahashi Y, Muneoka A, Sukamoto T, Fukuzaki K, Shoji S (2009) Targeted delivery of immunogen to primate M cells with tetragalloyl lysine dendrimer. J Immunol 182:6061–6070. https://doi.org/10.4049/jimmunol.0802928

    CAS  Article  PubMed  Google Scholar 

  18. Miyoshi A, Poquet I, Azevedo V, Commissaire J, Domakova E, Loir Y Le, Oliveira SC, Gruss A, Langella P (2002) Controlled production of stable heterologous proteins in Lactococcus lactis. Appl Env Microbiol 68:3141–3146. doi: https://doi.org/10.1128/AEM.68.6.3141

  19. Nakato G, Hase K, Suzuki M, Kimura M, Ato M, Hanazato M, Tobiume M, Horiuchi M, Atarashi R, Nishida N, Watarai M, Imaoka K, Ohno H (2012) Cutting edge: Brucella abortus exploits a cellular prion protein on intestinal M cells as an invasive receptor. J Immunol 189:1540–1544. https://doi.org/10.4049/jimmunol.1103332

    CAS  Article  PubMed  Google Scholar 

  20. Nimmerjahn F, Ravetch J V. (2005) Divergent immunoglobulin G subclass activity through selective fc receptor binding. Science 310:1510–1512. https://doi.org/10.1126/science.1118948

    CAS  Article  Google Scholar 

  21. Ohno H (2015) Intestinal M cells. J Biochem 159:151–160. https://doi.org/10.1093/jb/mvv121

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  22. Pavot V, Rochereau N, Genin C, Verrier B, Paul S (2012) New insights in mucosal vaccine development. Vaccine 30:142–154. https://doi.org/10.1016/j.vaccine.2011.11.003

    CAS  Article  PubMed  Google Scholar 

  23. Poquet I, Saint V, Seznec E, Simoes N, Bolotin A, Gruss A (2000) HtrA is the unique surface housekeeping protease in Lactococcus lactis and is required for natural protein processing. Mol Microbiol 35:1042–1051. https://doi.org/10.1046/j.1365-2958.2000.01757.x

    CAS  Article  PubMed  Google Scholar 

  24. Pouwels PH, Leer RJ, Shaw M, Heijne den Bak-Glashouwer M-J, Tielen FD, Smit E, Martinez B, Jore J, Conway PL (1998) Lactic acid bacteria as antigen delivery vehicles for oral immunization purposes. Int J Food Microbiol 41:155–167. https://doi.org/10.1016/S0168-1605(98)00048-8

    CAS  Article  PubMed  Google Scholar 

  25. Reboldi A, Cyster JG (2016) Peyer’s patches: organizing B cell responses at the intestinal frontier. Immunol Rev 271:230–245. https://doi.org/10.1111/imr.12400

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  26. Rescigno M, Urbano M, Valzasina B, Francolini M, Rotta G, Bonasio R, Granucci F, Kraehenbuhl J, Ricciardi-castagnoli P (2001) Dendritic cells express tight junction proteins and penetrate gut epithelial monolayers to sample bacteria. Nat Immunol 2:361–367. https://doi.org/10.1038/86373

    CAS  Article  PubMed  Google Scholar 

  27. Rey J, Garin N, Spertini F, Corthesy B (2004) Targeting of secretory IgA to Peyer’s patch dendritic and T cells after transport by intestinal M cells. J Immunol 172:3026–3033. https://doi.org/10.4049/jimmunol.172.5.3026

    CAS  Article  PubMed  Google Scholar 

  28. Schilcher K, Andreoni F, Uchiyama S, Ogawa T, Schuepbach RA, Zinkernagel AS (2014) Increased neutrophil extracellular trap-mediated Staphylococcus aureus clearance through inhibition of nuclease activity by clindamycin and immunoglobulin. J Infect Dis 210:473–482. https://doi.org/10.1093/infdis/jiu091

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  29. Shima H, Watanabe T, Fukuda S, Fukuoka S-I, Ohara O, Ohno H (2014) A novel mucosal vaccine targeting Peyer’s patch M cells induces protective antigen-specific IgA responses. Int Immunol 26:619–625. https://doi.org/10.1093/intimm/dxu061

    CAS  Article  PubMed  Google Scholar 

  30. Song AA-L, In LLA, Lim SHE, Rahim RA (2017) A review on Lactococcus lactis: from food to factory. Microb Cell Factories 16:55. https://doi.org/10.1186/s12934-017-0669-x

    CAS  Article  Google Scholar 

  31. Stavnezer J (1995) Regulation of antibody production and class switching by TGF-beta. J Immunol 155:1647–1651

    CAS  PubMed  Google Scholar 

  32. Wyszyńska A, Kobierecka P, Bardowski J, Jagusztyn-Krynicka EK (2015) Lactic acid bacteria-20 years exploring their potential as live vectors for mucosal vaccination. Appl Microbiol Biotechnol 99:2967–2977. https://doi.org/10.1007/s00253-015-6498-0

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  33. Yamamoto M, Pascual DW, Kiyono H (2012) M cell-targeted mucosal vaccine strategies. Curr Top Microbiol Immunol 354:39–52

    CAS  PubMed  Google Scholar 

  34. Yano A, Takahashi K, Mori Y, Watanabe S, Hanamura Y, Sugiyama T, Inoue N (2018) Peyer’s patches as a portal for DNA delivery by Lactococcus lactis in vivo. Biol Pharm Bull 41:190–197. https://doi.org/10.1248/bpb.b17-00657

    CAS  Article  PubMed  Google Scholar 

  35. Ye T, Yue Y, Fan X, Dong C, Xu W, Xiong S (2014) M cell-targeting strategy facilitates mucosal immune response and enhances protection against CVB3-induced viral myocarditis elicited by chitosan-DNA vaccine. Vaccine 32:4457–4465. https://doi.org/10.1016/j.vaccine.2014.06.050

    CAS  Article  PubMed  Google Scholar 

Download references

Funding

This study was supported by a Grant-in-Aid for Young Scientists B (JP 16K19127) from the Japan Society for the Promotion of Science to KT and by a scholarship fund from the Lactic Acid Bacteria Foundation to NI.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Keita Takahashi.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical statement

All animal experiments were approved by the institutional Animal Care and Use Committee (approval number for ligated intestinal loop assay: 2017-236, and for immunization of mice: 2017-242).

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Takahashi, K., Yano, A., Watanabe, S. et al. M cell–targeting strategy enhances systemic and mucosal immune responses induced by oral administration of nuclease-producing L. lactis. Appl Microbiol Biotechnol 102, 10703–10711 (2018). https://doi.org/10.1007/s00253-018-9427-1

Download citation

Keywords

  • Lactococcus lactis
  • Oral immunization
  • OmpH
  • M cell
  • Peyer’s patch
  • NICE system