IgM is Unable to Enhance Antibody Responses in Mice Lacking C1q or C3

To the Editor: Lack of complement factors C1q, C2, C3 or C4, but not of factor B in the alternative pathway or of mannan-binding lectin in the lectin pathway, leads to severe impairment of primary antibody responses (reviewed in 1). Because C1q is only used in classical pathway activation, these observations suggest that only this pathway is involved in antibody responses. An efficient classical pathway activator is IgM. Specific IgM, passively administered together with suboptimal doses of the antigen it recognizes, induces a stronger antibody response than does the antigen alone 2. This effect has been attributed to the ability of IgM to activate complement because IgM with a point mutation in the heavy chain, rendering it unable to activate complement 3, 4, and monomeric IgM which cannot activate complement 5 loose the ability to enhance antibody responses. Moreover, IgM-mediated enhancement is severely impaired in mice lacking complement receptors 1 and 2 (CR1/2) 6, 7 and in mice partially depleted of complement factor C3 by treatment with cobra venom factor 3. The requirement for C1q in primary antibody responses is paradoxical because naïve mice have very low titres of specific antibody available to bind the antigen and to form the immune complexes presumably required to initiate the classical pathway. Reports that also natural IgM may upregulate immune responses (reviewed in 8, 9) led us and others to postulate that natural IgM in naïve animals could bind with low affinity to antigen, activate complement and enhance the antibody response in a similar way as specific IgM has been shown to do. Knock-in mice (Cμ13) with the same point mutation shown to inhibit the ability of specific IgM to enhance antibody responses 3, 4 were constructed and immunized with SRBC alone. Surprisingly, their antibody responses were largely normal and never as impaired as those observed in mice lacking C1q or CR1/2 10. This unexpected observation made us re-investigate the requirement for complement, in particular the classical pathway, for the ability of specific IgM to enhance antibody responses. In a previously not used approach, wild-type mice, C3 knockout mice (in which all three complement pathways are blocked) and C1qA knockout mice (lacking the entire C1q molecule and thereby classical pathway activation) were immunized with a low dose of sheep red blood cells (SRBC) ± IgM anti-SRBC. The IgG anti-SRBC response was followed during four weeks (Fig. 1). IgM enhanced the antibody responses in wild-type, but not in C1q- or C3-deficient mice. This clearly demonstrates that activation via the classical pathway and the presence of C3 are crucial for the ability of specific IgM to feedback enhance antibody responses. The observation strengthens the conclusion (based on the unperturbed antibody responses seen in Cμ13 mice 10) that feedback enhancement by complement-activating natural IgM cannot explain the role of C1q in primary antibody responses. Therefore, this remains a paradox to be resolved.