A major pathway for B cell acquisition of lymph-borne particulate antigens relies on antigen capture by subcapsular sinus macrophages of the lymph node. Here we tested whether this mechanism is also important for humoral immunity to inactivated influenza virus. By multiple approaches, including multiphoton intravital imaging, we found that antigen capture by sinus-lining macrophages was important for limiting the systemic spread of virus but not for the generation of influenza-specific humoral immunity. Instead, we found that dendritic cells residing in the lymph node medulla use the lectin receptor SIGN-R1 to capture lymph-borne influenza virus and promote humoral immunity. Thus, our results have important implications for the generation of durable humoral immunity to viral pathogens through vaccination.
Protein-carbohydrate interactions play a critical role in many biological recognition events. Multivalent therapeutic agents that utilize protein-carbohydrate interactions have proven difficult to design, primarily because the fundamental requirements of protein-carbohydrate interactions are not well understood. Here, we report a systematic study of the effect on lectin binding of varying the loading of mannose surface residues on generations three through six PAMAM dendrimers. The degree of mannose functionalization was controlled by stoichiometric addition, and dendrimers were characterized using NMR and MALDI-TOF MS. Hemagglutination assays and quantitative precipitation assays were performed to determine the relative activity of the dendrimers. Using the mannose/hydroxyl-functionalized dendrimers reported here, we could systematically control both the degree of lectin clustering and the overall activity of the lectin with the dendrimer.
G4-, G5-, and G6-PAMAM dendrimers were functionalized with mixtures of mannose and glucose in varying ratios, and the relative affinities of these compounds for Concanavalin A (Con A) were evaluated using the hemagglutination assay. As the ratio of mannose to glucose increases, the relative activity in the hemagglutination assay (on a per sugar basis) increases linearly. Methyl mannose binds to Con A with an affinity 4-fold higher than that of methyl glucose; multivalency amplifies this trend. The mannose/glucose-functionalized dendrimer results reported here suggest that the affinity of multivalent associations can be attenuated in predictable, reliable ways based on monovalent affinities of the ligands.
[structure: see text] First- through sixth-generation PAMAM dendrimers have been functionalized with mannose residues. Characterization with MALDI-TOF MS and (1)H NMR is reported. Different binding enhancements consistent with monovalent interaction, glycoside clustering, and multivalent binding are observed for different generations of dendrimers.
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