Changes in the expression of FcγRs on alveolar macrophages and the negative role of FcγRIIb in cross-protection
(A and B) After immunization with SV plus CpG/alum, clodronate liposomes or control liposomes were administered to mice intranasally. After the 1.2 × 103 TCID50 PR8 challenge, we monitored (A) percentages of initial body weights and (B) survival for the next 15 days. Mice treated with PBS were challenged with 1.2 × 103 TCID50 PR8 and used as control.
(C) After immunization with SV plus CpG/alum, anti-IFN-γ antibody or isotype antibody was injected before the 1.2 × 103 TCID50 PR8 challenge. Five days after PR8 challenge, the expression of FcγRI, FcγRIIb, FcγRIII, and FcγRIV on alveolar macrophages was determined by flow cytometry.
(D and E) A mixture of 1.2 × 10 TCID50 PR8 and 2-fold diluted serum obtained from PBS-treated control mice or SV plus CpG/alum-immunized mice was administered to naive mice intranasally. Anti-FcγRIIb antagonistic monoclonal antibody (AT128) or isotype control antibody was injected into these mice and we monitored (D) percentages of initial body weights and (E) survival for the next 15 days.
(F and G) A mixture of 1.2 × 10 TCID50 PR8 and 2-fold diluted serum obtained from SV plus CpG–ODN-immunized mice was administered to naive mice intranasally. Anti-FcγRIIb agonistic antibody (AT130-2) or isotype control antibody was injected into these mice and we monitored (F) percentages of initial body weights and (G) survival for the next 15 days. (A and B) n = 4–5, (C) n = 5 for FcγRI, FcγRIII, and FcγRIV, or 15 for FcγRIIb, (D and E) n = 8–10, (F and G) n = 5 per group. (A, C, D and F) Data are means ± SD. (B) ∗∗p<0.01 between clodronate-loaded liposomes vs. control liposomes group as indicated by comparing Kaplan–Meier curves using the logrank test. (C) ∗∗∗p<0.001 as indicated by Student's ttest. (E) ∗p<0.05 between AT128 plus serum from SV plus CpG/alum-immunized mice vs. isotype control antibody plus serum obtained from SV plus CpG/alum-immunized mice; (G) ∗∗p<0.01 as indicated by comparing Kaplan–Meier curves using the logrank test. See also Figures S7, S8, and S9.