Fig. 5.

Reconstitution of CD4 T cell priming restores GC B cell responses in SIGN-R1 macrophage-depleted mice. (A–C) CD45.2⁺ mice were preprimed with OVA at 2 wk before CLL or PBS-L treatment. Two weeks later, mice were transferred with CD45.1⁺ MD4 B cells, followed by immunization with HEL-OVA. Eight days later, spleens were analyzed by IHC and flow cytometry. (A) Graphic representation of the experimental protocol. (B) IHC analysis at day 8 postimmunization. Sections (from two mice per group) were stained for GL7 (to identify GCs; brown) and IgD (to define follicular B cells; blue). Selective ablation of SIGN-R1, but not of CD169⁺, cells was confirmed by staining consecutive sections for IgD (blue) and SIGN-R1 or CD169 (brown). (C) Frequencies of CD45.1⁺ B cells determined by flow cytometry analysis. A representative example is shown out of three independent experiments performed. (D) Mice were preprimed with SRBCs 2 wk before CLL or PBS-L treatment, then immunized with PE 2 wk later. Shown are frequencies of PE⁺ GC B cells in splenocytes analyzed by cytometry at 8 d postimmunization, from a representative experiment out of three experiments performed. (E) Mice (CD45.2⁺) treated with CLL or PBS-L 3 wk earlier were transferred with CD45.1⁺ MD4 B cells with or without cotransfer of CD4 T cells (CD45.2⁺) that had been purified from mice immunized with OVA 2 wk earlier. Recipients were immunized with HEL-OVA, and spleens were analyzed 8 d later to determine the frequencies of CD45.1⁺ GC B cells. (F and G) Hosts (CD45.2⁺) were treated with CLLs or PBS-Ls. After 2 to 3 wk, the mice were transferred with CD45.1⁺ MD4 B cells and immunized with HEL-OVA. Then, 3.5 d later, the mice were i.v. infused with or without chimeric anti-DEC205 (F) or anti-33D1 (G) fused to an OVA-derived peptide. The frequencies of CD45.1⁺ GC B cells were determined 8 d later by flow cytometry. Shown are the results of a representative experiment out of three experiments performed.