Complement‐mediated follicular localization of T‐independent type‐2 antigens: the role of marginal zone macrophages revisited

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Abstract
In this study we demonstrate a hitherto undescribed phenomenon, namely that thymus-independent type-2 antigens (TI-2 Ag) localize in splenic follicles within 1 h after administration. The follicular localization of 2,4,6-trinitrophenyl (TNP)-Ficoll was not antibody mediated. In addition in case of high-dose administration we observed a relatively large amount of TI-2 Ag in marginal zone macrophages. However, after low-dose administration we observed a preferential localization of TNP-Ficoll in the splenic follicles. Detection of TNP-haptenated Ag in cryostat sections of murine spleens was performed with a high-affinity TNP-specific monoclonal antibody conjugated to β-galactosidase. Within minutes after injection the TI-2 Ag localized in the marginal zone, attached to marginal zone macrophages and B cells. Twenty minutes after injection the Ag was also detected in the follicles and gradually accumulated there until 7 h after injection. Thereafter, the amount of follicular Ag gradually decreased but was still detectable up to 14 days after immunization. The follicular localization of TNP-Ficoll was complement dependent in contrast to the binding to and uptake by marginal zone macrophages. Double staining revealed that Ag was bound by macrophages, B cells and follicular dendritic cells. Haptenated thymus-dependent (TD) Ag localized exclusively in the red pulp macrophages. In vivo macrophage elimination drastically increased the amount of TNP-Ficoll in the follicles, and enhanced the humoral immune response at low doses of Ag. Moreover, complement deprivation of mice abrogated the localization of TI-2 Ag in the follicles, and led to a decreased humoral TI-2 immune response. In conclusion, we demonstrate for the first time that TI-2 Ag localize in follicles. Moreover, the presented results provide further evidence that B cells and follicular localized Ag play an important role in the induction of humoral TI-2 immune responses.