Further insights into the anti-PF4/heparin IgM immune response

Summary

Anti-platelet factor 4 (PF4)/heparin antibodies are not only the cause of heparin-induced thrombocytopenia but might also play a role in the antibacterial host defence. Recently, marginal zone (MZ) B cells were identified to be crucial for anti-PF4/heparin IgG antibody production in mice. Combining human studies and a murine model of polymicrobial sepsis we further characterised the far less investigated anti-PF4/heparin IgM immune response. We detected anti-PF4/heparin IgM antibodies in the sera of paediatric patients < 6 months of age after cardiac surgery and in sera of splenectomised mice subjected to polymicrobial sepsis. In addition, PF4/heparin-specific IgM B cells were not only found in murine spleen, but also in peritoneum and bone marrow upon in vitro stimulation. Together, this indicates involvement of additional B cell populations, as MZ B cells are not fully developed in humans until the second year of life and are restricted to the spleen in mice. Moreover, PF4/heparin-specific B cells were detected in human cord blood upon in vitro stimulation and PF4^-/- mice produced anti-PF4/heparin IgM antibodies after polymicrobial sepsis. In conclusion, the anti-PF4/heparin IgM response is a potential innate immune reaction driven by a B cell population distinct from MZ B cells.

Supplementary Material to this article is available online at www.thrombosis-online.com.

Keywords

HIT - anti-PF4/heparin antibodies - B cells - immune response

^* Authors contributed equally to this work.

• References

• 1 Arepally GM, Ortel TL. Clinical practice. Heparin-induced thrombocytopenia. N Engl J Med 2006; 355: 809-817.
• 2 Greinacher A, Juhl D, Strobel U. et al. Heparin-induced thrombocytopenia: a prospective study on the incidence, platelet-activating capacity and clinical significance of antiplatelet factor 4/heparin antibodies of the IgG, IgM, and IgA classes. J Thromb Haemost 2007; 05: 1666-1673.
• 3 Pouplard C, May MA, Iochmann S. et al. Antibodies to platelet factor 4-heparin after cardiopulmonary bypass in patients anticoagulated with unfractionated heparin or a low-molecular-weight heparin : clinical implications for heparin-induced thrombocytopenia. Circulation 1999; 99: 2530-2536.
• 4 Suh JS, Malik MI, Aster RH. et al. Characterisation of the humoral immune response in heparin-induced thrombocytopenia. Am J Hematol 1997; 54: 196-201.
• 5 Warkentin TE. Heparin-induced thrombocytopenia: pathogenesis and management. Br J Haematol 2003; 121: 535-555.
• 6 Krauel K, Potschke C, Weber C. et al. Platelet factor 4 binds to bacteria, [corrected] inducing antibodies cross-reacting with the major antigen in heparin-induced thrombocytopenia. Blood 2011; 117: 1370-1378.
• 7 Krauel K, Weber C, Brandt S. et al. Platelet factor 4 binding to lipid A of Gramnegative bacteria exposes PF4/heparin-like epitopes. Blood 2012; 120: 3345-3352.
• 8 Brandt S, Krauel K, Jaax M. et al. Polyphosphates form antigenic complexes with platelet factor 4 (PF4) and enhance PF4-binding to bacteria. Thromb Haemost 2015; 114: 1189-1198.
• 9 Zheng Y, Yu M, Podd A. et al. Critical role for mouse marginal zone B cells in PF4/heparin antibody production. Blood 2013; 121: 3484-3492.
• 10 Suvarna S, Rauova L, McCracken EK. et al. PF4/heparin complexes are T cell-dependent antigens. Blood 2005; 106: 929-931.
• 11 Zheng Y, Yu M, Padmanabhan A. et al. Critical role of CD4 T cells in PF4/hepa-rin antibody production in mice. Blood 2015; 125: 1826-1829.
• 12 Selleng K, Schutt A, Selleng S. et al. Studies of the anti-platelet factor 4/heparin immune response: adapting the enzyme-linked immunosorbent spot assay for detection of memory B cells against complex antigens. Transfusion 2010; 50: 32-39.
• 13 Martin F, Oliver AM, Kearney JF. Marginal zone and B1 B cells unite in the early response against T-independent blood-borne particulate antigens. Immunity 2001; 14: 617-629.
• 14 Cerutti A, Cols M, Puga I. Marginal zone B cells: virtues of innate-like antibody-producing lymphocytes. Nat Rev Immunol 2013; 13: 118-132.
• 15 Yamanishi S, Iizumi T, Watanabe E. et al. Implications for induction of autoimmunity via activation of B-1 cells by Helicobacter pylori urease. Infect Immun 2006; 74: 248-256.
• 16 Parra D, Takizawa F, Sunyer JO. Evolution of B cell immunity. Annu Rev Anim Biosci 2013; 01: 65-67.
• 17 Pillai S, Cariappa A, Moran ST. Marginal zone B cells. Annu Rev Immunol 2005; 23: 161-196.
• 18 Kantor A. A new nomenclature for B cells. Immunol Today 1991; 12: 388.
• 19 Choi YS, Dieter JA, Rothaeusler K. et al. B-1 cells in the bone marrow are a significant source of natural IgM. Eur J Immunol 2012; 42: 120-129.
• 20 Kantor AB, Stall AM, Adams S. et al. De novo development and self-replenishment of B cells. Int Immunol 1995; 07: 55-68.
• 21 Wardemann H, Boehm T, Dear N. et al. B-1a B cells that link the innate and adaptive immune responses are lacking in the absence of the spleen. J Exp Med 2002; 195: 771-780.
• 22 Griffin DO, Holodick NE, Rothstein TL. Human B1 cells in umbilical cord and adult peripheral blood express the novel phenotype CD20+ CD27+ CD43+ CD70. J Exp Med 2011; 208: 67-80.
• 23 Timens W, Boes A, Rozeboom-Uiterwijk T. et al. Immaturity of the human splenic marginal zone in infancy. Possible contribution to the deficient infant immune response. J Immunol 1989; 143: 3200-3206.
• 24 Montecino-Rodriguez E, Dorshkind K. B-1 B cell development in the fetus and adult. Immunity 2012; 36: 13-21.
• 25 Newman PM, Swanson RL, Chong BH. Heparin-induced thrombocytopenia: IgG binding to PF4-heparin complexes in the fluid phase and cross-reactivity with low molecular weight heparin and heparinoid. Thromb Haemost 1998; 80: 292-297.
• 26 Selleng S, Malowsky B, Strobel U. et al. Early-onset and persisting thrombocytopenia in post-cardiac surgery patients is rarely due to heparin-induced thrombocytopenia, even when antibody tests are positive. J Thromb Haemost 2010; 08: 30-36.
• 27 Juhl D, Eichler P, Lubenow N. et al. Incidence and clinical significance of anti-PF4/heparin antibodies of the IgG, IgM, and IgA class in 755 consecutive patient samples referred for diagnostic testing for heparin-induced thrombocytopenia. Eur J Haematol 2006; 76: 420-426.
• 28 Eslin DE, Zhang C, Samuels KJ. et al. Transgenic mice studies demonstrate a role for platelet factor 4 in thrombosis: dissociation between anticoagulant and antithrombotic effect of heparin. Blood 2004; 104: 3173-3180.
• 29 Schulze A, Jensch I, Krauel K. et al. New insights in heparin-induced thrombocytopenia by the use of fluid-phase assays to detect specifically platelet factor 4/heparin complex antibodies and antibody-secreting cells. Thromb Res 2014; 134: 174-181.
• 30 Bakchoul T, Zollner H, Amiral J. et al. Anti-protamine-heparin antibodies: incidence, clinical relevance, and pathogenesis. Blood 2013; 121: 2821-2827.
• 31 Zhang X. Regulatory functions of innate-like B cells. Cell Mol Immunol 2013; 10: 113-121.
• 32 Chovancova Z, Vlkova M, Litzman J. et al. Antibody forming cells and plasmab-lasts in peripheral blood in CVID patients after vaccination. Vaccine 2011; 29: 4142-4150.
• 33 Greinacher A, Holtfreter B, Krauel K. et al. Association of natural anti-platelet factor 4/heparin antibodies with periodontal disease. Blood 2011; 118: 1395-1401.
• 34 Jones DD, DeIulio GA, Winslow GM. Antigen-driven induction of polyreactive IgM during intracellular bacterial infection. J Immunol 2012; 189: 1440-1447.
• 35 Choi YS, Baumgarth N. Dual role for B-1a cells in immunity to influenza virus infection. J Exp Med 2008; 205: 3053-3064.
• 36 Suvarna S, Qi R, Arepally GM. Optimisation of a murine immunisation model for study of PF4/heparin antibodies. J Thromb Haemost 2009; 07: 857-864.