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Immune complex relay by subcapsular sinus macrophages and noncognate B cells drives antibody affinity maturation

Nature Immunology volume 10, pages 786–793 (2009)Cite this article

Abstract

Subcapsular sinus (SCS) macrophages capture antigens from lymph and present them intact for B cell encounter and follicular delivery. However, the properties of SCS macrophages are poorly defined. Here we show SCS macrophage development depended on lymphotoxin-α1β2, and the cells had low lysosomal enzyme expression and retained opsonized antigens on their surface. Intravital imaging revealed immune complexes moving along macrophage processes into the follicle. Moreover, noncognate B cells relayed antigen opsonized by newly produced antibodies from the subcapsular region to the germinal center, and affinity maturation was impaired when this transport process was disrupted. Thus, we characterize SCS macrophages as specialized antigen-presenting cells functioning at the apex of an antigen transport chain that promotes humoral immunity.

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Figure 1: Isolation and identification of SCS macrophages.
Figure 2: SCS macrophages are poorly endocytic and poorly degradative.
Figure 3: SCS macrophages express low amounts of lysosomal enzymes.
Figure 4: B cell–derived lymphotoxin signaling is required for SCS macrophage differentiation.
Figure 5: Noncognate B cells relay antigen opsonized by newly formed antibodies into the GC.
Figure 6: Immune complex relay into the GC drives affinity maturation.

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References

  1. Nossal, G.J., Abbot, A., Mitchell, J. & Lummus, Z. Antigens in immunity. XV. Ultrastructural features of antigen capture in primary and secondary lymphoid follicles. J. Exp. Med. 127, 277–290 (1968).

    Article  CAS  Google Scholar 

  2. Fossum, S. The architecture of rat lymph nodes. IV. Distribution of ferritin and colloidal carbon in the draining lymph nodes after foot-pad injection. Scand. J. Immunol. 12, 433–441 (1980).

    Article  CAS  Google Scholar 

  3. Szakal, A.K., Holmes, K.L. & Tew, J.G. Transport of immune complexes from the subcapsular sinus to lymph node follicles on the surface of nonphagocytic cells, including cells with dendritic morphology. J. Immunol. 131, 1714–1727 (1983).

    CAS  PubMed  Google Scholar 

  4. Carrasco, Y.R. & Batista, F.D. B cells acquire particulate antigen in a macrophage-rich area at the boundary between the follicle and the subcapsular sinus of the lymph node. Immunity 27, 160–171 (2007).

    Article  CAS  Google Scholar 

  5. Phan, T.G., Grigorova, I., Okada, T. & Cyster, J.G. Subcapsular encounter and complement-dependent transport of immune complexes by lymph node B cells. Nat. Immunol. 8, 992–1000 (2007).

    Article  CAS  Google Scholar 

  6. Junt, T. et al. Subcapsular sinus macrophages in lymph nodes clear lymph-borne viruses and present them to antiviral B cells. Nature 450, 110–114 (2007).

    Article  CAS  Google Scholar 

  7. Steinman, R.M. & Banchereau, J. Taking dendritic cells into medicine. Nature 449, 419–426 (2007).

    Article  CAS  Google Scholar 

  8. MacLennan, I.C.M. Germinal centers. Annu. Rev. Immunol. 12, 117–139 (1994).

    Article  CAS  Google Scholar 

  9. Allen, C.D., Okada, T. & Cyster, J.G. Germinal-center organization and cellular dynamics. Immunity 27, 190–202 (2007).

    Article  CAS  Google Scholar 

  10. Schwickert, T.A. et al. In vivo imaging of germinal centres reveals a dynamic open structure. Nature 446, 83–87 (2007).

    Article  CAS  Google Scholar 

  11. Allen, C.D., Okada, T., Tang, H.L. & Cyster, J.G. Imaging of germinal center selection events during affinity maturation. Science 315, 528–531 (2007).

    Article  CAS  Google Scholar 

  12. Crocker, P.R. & Gordon, S. Properties and distribution of a lectin-like hemagglutinin differentially expressed by murine stromal tissue macrophages. J. Exp. Med. 164, 1862–1875 (1986).

    Article  CAS  Google Scholar 

  13. Hume, D.A., Robinson, A.P., MacPherson, G.G. & Gordon, S. The mononuclear phagocyte system of the mouse defined by immunohistochemical localization of antigen F4/80. Relationship between macrophages, Langerhans cells, reticular cells, and dendritic cells in lymphoid and hematopoietic organs. J. Exp. Med. 158, 1522–1536 (1983).

    Article  CAS  Google Scholar 

  14. Fu, Y.-X. & Chaplin, D.D. Development and maturation of secondary lymphoid tissues. Annu. Rev. Immunol. 17, 399–433 (1999).

    Article  CAS  Google Scholar 

  15. Browning, J.L. Inhibition of the lymphotoxin pathway as a therapy for autoimmune disease. Immunol. Rev. 223, 202–220 (2008).

    Article  CAS  Google Scholar 

  16. Ngo, V.N., Cornall, R.J. & Cyster, J.G. Splenic T zone development is B cell dependent. J. Exp. Med. 194, 1649–1660 (2001).

    Article  CAS  Google Scholar 

  17. Cascalho, M., Ma, A., Lee, S., Masat, L. & Wabl, M. A quasi-monoclonal mouse. Science 272, 1649–1652 (1996).

    Article  CAS  Google Scholar 

  18. Germain, R.N., Miller, M.J., Dustin, M.L. & Nussenzweig, M.C. Dynamic imaging of the immune system: progress, pitfalls and promise. Nat. Rev. Immunol. 6, 497–507 (2006).

    Article  CAS  Google Scholar 

  19. Roozendaal, R. & Carroll, M.C. Complement receptors CD21 and CD35 in humoral immunity. Immunol. Rev. 219, 157–166 (2007).

    Article  CAS  Google Scholar 

  20. Chtanova, T. et al. Dynamics of neutrophil migration in lymph nodes during infection. Immunity 29, 487–496 (2008).

    Article  CAS  Google Scholar 

  21. Farr, A.G., Cho, Y. & De Bruyn, P.P. The structure of the sinus wall of the lymph node relative to its endocytic properties and transmural cell passage. Am. J. Anat. 157, 265–284 (1980).

    Article  CAS  Google Scholar 

  22. Prigozhina, N.L. & Waterman-Storer, C.M. Protein kinase D-mediated anterograde membrane trafficking is required for fibroblast motility. Curr. Biol. 14, 88–98 (2004).

    Article  CAS  Google Scholar 

  23. Wulfing, C. & Davis, M.M. A receptor/cytoskeletal movement triggered by costimulation during T cell activation. Science 282, 2266–2269 (1998).

    Article  CAS  Google Scholar 

  24. Moss, W.C., Irvine, D.J., Davis, M.M. & Krummel, M.F. Quantifying signaling-induced reorientation of T cell receptors during immunological synapse formation. Proc. Natl. Acad. Sci. USA 99, 15024–15029 (2002).

    Article  CAS  Google Scholar 

  25. Engstler, M. et al. Hydrodynamic flow-mediated protein sorting on the cell surface of trypanosomes. Cell 131, 505–515 (2007).

    Article  CAS  Google Scholar 

  26. Kabashima, K. et al. Intrinsic lymphotoxin-beta receptor requirement for homeostasis of lymphoid tissue dendritic cells. Immunity 22, 439–450 (2005).

    Article  CAS  Google Scholar 

  27. Wang, Y. et al. Antigen persistence is required for somatic mutation and affinity maturation of immunoglobulin. Eur. J. Immunol. 30, 2226–2234 (2000).

    Article  CAS  Google Scholar 

  28. Hannum, L.G., Haberman, A.M., Anderson, S.M. & Shlomchik, M.J. Germinal center initiation, variable gene region hypermutation, and mutant B cell selection without detectable immune complexes on follicular dendritic cells. J. Exp. Med. 192, 931–942 (2000).

    Article  CAS  Google Scholar 

  29. Brady, L.J. Antibody-mediated immunomodulation: a strategy to improve host responses against microbial antigens. Infect. Immun. 73, 671–678 (2005).

    Article  CAS  Google Scholar 

  30. Getahun, A. & Heyman, B. How antibodies act as natural adjuvants. Immunol. Lett. 104, 38–45 (2006).

    Article  CAS  Google Scholar 

  31. Song, H., Nie, X., Basu, S. & Cerny, J. Antibody feedback and somatic mutation in B cells: regulation of mutation by immune complexes with IgG antibody. Immunol. Rev. 162, 211–218 (1998).

    Article  CAS  Google Scholar 

  32. Ehrenstein, M.R., O'Keefe, T.L., Davies, S.L. & Neuberger, M.S. Targeted gene disruption reveals a role for natural secretory IgM in the maturation of the primary immune response. Proc. Natl. Acad. Sci. USA 95, 10089–10093 (1998).

    Article  CAS  Google Scholar 

  33. Boes, M. et al. Enhanced B-1 cell development, but impaired IgG antibody responses in mice deficient in secreted IgM. J. Immunol. 160, 4776–4787 (1998).

    CAS  PubMed  Google Scholar 

  34. Hadjantonakis, A.K., Macmaster, S. & Nagy, A. Embryonic stem cells and mice expressing different GFP variants for multiple non-invasive reporter usage within a single animal. BMC Biotechnol. 2, 11 (2002).

    Article  Google Scholar 

  35. Schaefer, B.C., Schaefer, M.L., Kappler, J.W., Marrack, P. & Kedl, R.M. Observation of antigen-dependent CD8+ T-cell/ dendritic cell interactions in vivo. Cell. Immunol. 214, 110–122 (2001).

    Article  CAS  Google Scholar 

  36. Kitamura, D., Roes, J., Kuhn, R. & Rajewsky, K. A B cell-deficient mouse by targeted disruption of the membrane exon of the immunoglobulin μ chain gene. Nature 350, 423–426 (1991).

    Article  CAS  Google Scholar 

  37. De Togni, P. et al. Abnormal development of peripheral lymphoid organs in mice deficient in lymphotoxin. Science 264, 703–707 (1994).

    Article  CAS  Google Scholar 

  38. Futterer, A., Mink, K., Luz, A., Kosco-Vilbois, M.H. & Pfeffer, K. The lymphotoxin beta receptor controls organogenesis and affinity maturation in peripheral lymphoid tissues. Immunity 9, 59–70 (1998).

    Article  CAS  Google Scholar 

  39. Molina, H. et al. Markedly impaired humoral immune response in mice deficient in complement receptors 1 and 2. Proc. Natl. Acad. Sci. USA 93, 3357–3361 (1996).

    Article  CAS  Google Scholar 

  40. Barnden, M.J., Allison, J., Heath, W.R. & Carbone, F.R. Defective TCR expression in transgenic mice constructed using cDNA-based alpha- and beta-chain genes under the control of heterologous regulatory elements. Immunol. Cell Biol. 76, 34–40 (1998).

    Article  CAS  Google Scholar 

  41. Crocker, P.R. & Gordon, S. Mouse macrophage hemagglutinin (sheep erythrocyte receptor) with specificity for sialylated glycoconjugates characterized by a monoclonal antibody. J. Exp. Med. 169, 1333–1346 (1989).

    Article  CAS  Google Scholar 

  42. Paus, D. et al. Antigen recognition strength regulates the choice between extrafollicular plasma cell and germinal center B cell differentiation. J. Exp. Med. 203, 1081–1091 (2006).

    Article  CAS  Google Scholar 

  43. Carroll, R.C. et al. Dynamin-dependent endocytosis of ionotropic glutamate receptors. Proc. Natl. Acad. Sci. USA 96, 14112–14117 (1999).

    Article  CAS  Google Scholar 

  44. Vandesompele, J. et al. Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol. 3, RESEARCH0034 (2002).

    Article  Google Scholar 

  45. Valledor, A.F., Borras, F.E., Cullell-Young, M. & Celada, A. Transcription factors that regulate monocyte/macrophage differentiation. J. Leukoc. Biol. 63, 405–417 (1998).

    Article  CAS  Google Scholar 

  46. Phan, T.G. et al. B cell receptor-independent stimuli trigger immunoglobulin (Ig) class switch recombination and production of IgG autoantibodies by anergic self-reactive B cells. J. Exp. Med. 197, 845–860 (2003).

    Article  CAS  Google Scholar 

  47. Smith-Gill, S.J., Mainhart, C.R., Lavoie, T.B., Rudikoff, S. & Potter, M. VL-VH expression by monoclonal antibodies recognizing avian lysozyme. J. Immunol. 132, 963–967 (1984).

    CAS  PubMed  Google Scholar 

  48. Smith-Gill, S.J., Lavoie, T.B. & Mainhart, C.R. Antigenic regions defined by monoclonal antibodies correspond to structural domains of avian lysozyme. J. Immunol. 133, 384–393 (1984).

    CAS  PubMed  Google Scholar 

  49. Okada, T. et al. Antigen-engaged B cells undergo chemotaxis toward the T zone and form motile conjugates with helper T cells. PLoS Biol. 3, e150 (2005).

    Article  Google Scholar 

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Acknowledgements

We thank L. Shiow for cell sorting and discussions, K. Suzuki for preparation of HEL-PE, C. Allen for help with single cell sorting, I. Girgorova for help with video editing, J. An for mouse screening, J. Atkinson (Washington University) for CR2-deficient mice, M. Wabl (University of California, San Francisco) for QM mice, J. Browning (Biogen Idec) for LTβR-Fc, P. Crocker (University of Glasgow) for Ser-4 antibody, A. Bullen and M. Krummel for help with the two-photon microscope, A. Bankovich for advice, the UCSF Hybridoma Core, the Gladstone Genomics Core and the Diabetes and Endocrinology Research Center Microscopy Core and Biological Imaging Development Center. Supported by an Australian National Health and Medical Research Council CJ Martin fellowship (T.G.P.), a US National Science Foundation graduate research fellowship (J.A.G.), a Howard Hughes Medical Institute investigator award (J.G.C.), the US National Institutes of Health (AI45073 and AI40098) and a Sandler New Technology Award.

Author information

Author notes

  1. Jesse A Green and Elizabeth E Gray: These authors contributed equally to this work.

Authors and Affiliations

  1. Howard Hughes Medical Institute and Department of Microbiology and Immunology, University of California San Francisco (UCSF), California, USA

    Tri Giang Phan, Jesse A Green, Elizabeth E Gray, Ying Xu & Jason G Cyster

  2. Garvan Institute of Medical Research, Darlinghurst, Sydney, New South Wales, Australia

    Tri Giang Phan

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Contributions

T.G.P. and J.G.C. designed and conceptualized the research; T.G.P., J.A.G. and E.E.G. did the experiments; Y.X. performed the microarray and Q-PCR analysis; T.G.P., J.A.G., E.E.G. and J.G.C. analyzed the data; T.G.P., J.A.G. and E.E.G. prepared figures; T.G.P. and J.G.C. wrote the manuscript.

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Correspondence to Tri Giang Phan or Jason G Cyster.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1 and 2 (PDF 401 kb)

Supplementary Video 1

SCS macrophages transport immune complexes along cellular processes into the follicle. (MPG 2404 kb)

Supplementary Video 2

Noncognate B cells carry immune complex from subcapsular region into the GC. (MPG 5312 kb)

Supplementary Video 3

Noncognate B cell carrying immune complex in the GC. (MPG 5212 kb)

Supplementary Video 4

Noncognate B cell migrating in GC light zone with immune complex. (MPG 7836 kb)

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Phan, T., Green, J., Gray, E. et al. Immune complex relay by subcapsular sinus macrophages and noncognate B cells drives antibody affinity maturation. Nat Immunol 10, 786–793 (2009). https://doi.org/10.1038/ni.1745

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