Involvement of Virus-Induced Interferon Production in IgG Autoantibody-Mediated Anemia

Abstract

Infection with viruses, such as the lactate dehydrogenase-elevating virus (LDV), is known to trigger the onset of autoimmune anemia through the enhancement of the phagocytosis of autoantibody-opsonized erythrocytes by activated macrophages. Type I interferon receptor-deficient mice show enhanced anemia, which suggests a protective effect of these cytokines, partly through the control of type II interferon production. The development of anemia requires the expression of Fcγ receptors (FcγR) I, III, and IV. Whereas LDV infection decreases FcγR III expression, it enhances FcγR I and IV expression in wild-type animals. The LDV-associated increase in the expression of FcγR I and IV is largely reduced in type I interferon receptor-deficient mice, through both type II interferon-dependent and -independent mechanisms. Thus, the regulation of the expression of FcγR I and IV, but not III, by interferons may partly explain the exacerbating effect of LDV infection on anemia that results from the enhanced phagocytosis of IgG autoantibody-opsonized erythrocytes.

Keywords: Fc receptors; autoimmune anemia; interferon; lactate dehydrogenase-elevating virus.

Figure 1

Role of type I and type II IFNs in anemia and ex vivo erythrophagocytosis in LDV-infected mice. (A) Hematocrits in groups of 6 to 8 C57BL/6 mice pooled from 2 independent experiments at different times after administration of 50 μg 34-3C IgG2a mAb or A6202F4M control mAb. LDV infection occurred one day before mAb administration. (B) Survival of 10 129/Sv or IFNAR KO-infected mice after administration of 350 μg 34-3C anti-RBC mAb. (C) Hematocrit in groups of WT 129/Sv or IFNAR KO mice challenged with 50 μg of 34-3C IgG2a mAb or A6202F4M control mAb. Hematocrit were measured 4 days after LDV infection. (D) Hematocrits after administration of 50 μg 34-3C IgG2a mAb to groups of 4 129/Sv or IFNAR KO-infected mice treated with or without IFN-γ and IFN-γR-blocking antibodies (300 μg one day before LDV infection, 1 mg 3 h after infection, followed by a dose of 1 mg every 2 days). (E) Ex vivo erythrophagocytosis of 34-3C-opsonized RBCs by macrophages of 9 to 11 129/Sv or IFNAR KO-infected mice pooled from 2 independent experiments and treated with or without IFN-γ and IFN-γR-blocking antibodies (300 μg one day before LDV infection, 500 μg 3 h after infection). * p < 0.05, ** p < 0.01, *** p < 0.001, * compared to infected 129/Sv; $$$ compared to anti-IFNγ-treated infected 129/Sv; #### compared to IFNAR KO. The results are shown as the means ± SEM.

Figure 2

Involvement of Fcγ receptors in LDV-exacerbated in vivo and ex vivo erythrophagocytosis. (A,B) Hematocrits of C57BL/6, FcγR I KO, FcγR I/III KO, FcγR II/III/IV KO, FcγR I/II/III/IV KO (A), and FcγR III KO (B) mice, 4 days after administration of 50 µg of 34-3C mAb. LDV infection was performed one day before the antibody challenge. The results are representative of at least 2 experiments and shown as the means ± SEM, ). * p < 0.05. (C) Ex vivo erythrophagocytosis of non-opsonized (dashed line) or 34-3C-opsonized (plain line) CMFDA-stained RBCs by macrophages isolated from (left panel) mock-infected C57BL/6, (middle panel) LDV-infected C57BL/6, and (right panel) LDV-infected FcγR I/II/III/IV KO (grey: fluorescence of macrophages without RBC incubation). The results are representative of at least 2 independent experiments.

Figure 3

Modulation of activating Fcγ receptors and of CR3 expression after LDV infection. (A–D) The CD11b+ F4/80+ peritoneal macrophages of 4 mock- or LDV-infected C57BL/6 were stained with either anti-FcγR I (A), anti-FcγR III (B,C), or anti-FcγR IV (D) antibodies. (E–H) Fcγ (E–G) and C3 (H) receptor gene expression in CD11b+ F4/80+ peritoneal macrophages from groups of 4 mock- or LDV-infected C57BL/6 were assessed by RT-qPCR. The results are shown as the means ± SEM. NTC: negative control. * p < 0.05, ** p < 0.01.

Figure 4

Role of type I and type II IFNs in Fcγ receptor expression in peritoneal macrophages of LDV-infected mice. The expressions of FcγR I (A), FcγR III (B), and FcγR IV (C) were measured by RT-qPCR in macrophages isolated from groups of 4 129/Sv or IFNAR KO mice 1 day after LDV infection. * compared to control 129/Sv; # compared to infected 129/Sv; $ compared to infected 129/Sv treated with IFN-γ and IFN-γR-blocking antibodies. & compared to infected IFNAR KO treated with IFN-γ and IFN-γR-blocking antibodies. The results are shown as the means ± SEM.