Splenic CD169+ marginal zone macrophages directly cross-prime CD8+ T cells in vivo using a vacuolar processing pathway

Abstract Cross-priming of CD8+ T cells is of critical importance in triggering CD8+ T cell responses against pathogens and tumors. It is commonly thought that conventional type 1 dendritic cells (cDC1) cross-prime with unequalled efficacy, conferred to them by a proteasome and TAP-dependent pathway. Here we present data challenging the unique cross-priming efficacy of both the cDC1 subset and the proteasome/TAP-dependent pathway. Splenic CD169 macrophages are strategically placed in the marginal zone for capture of blood-borne pathogens. However, they are considered unable to directly cross-present and to depend on antigen transfer to cDC1 for cross-priming. We have developed a protocol for purification of CD169 macrophages. Extensive characterization shows that these cells display morphology, phenotype and gene expression distinguishing them clearly from cDC1, cDC2, plasmacytoid DC and red pulp macrophages. In vitro, CD169 cells cross-present soluble, receptor-targeted and phagocytosed antigen with equal or better efficiency than cDC1. Remarkably, this is achieved through a vacuolar, BFA and epoxomicin-independent pathway. Using intravital biphoton microscopy, 3D light sheet imaging and antigen targeting to CD169, we show that CD169 macrophages establish long-lasting antigen-dependent contacts with cognate CD8+ T cells that result in T cell activation in situ and generation of triple-positive effector as well as memory cells. Therefore, splenic CD169 macrophages represent a second set of myeloid cells equipped for highly efficient cross-priming. The biological role of cross-priming by these cells as well as their cooperation with cDC1 cells are under investigation.