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. 2007 Feb 1;109(3):1131-7.
doi: 10.1182/blood-2006-05-023770. Epub 2006 Sep 19.

Control of coronavirus infection through plasmacytoid dendritic-cell-derived type I interferon

Luisa Cervantes-Barragan  1 , Roland Züst, Friedemann Weber, Martin Spiegel, Karl S Lang, Shizuo Akira, Volker Thiel, Burkhard Ludewig
Affiliations

Control of coronavirus infection through plasmacytoid dendritic-cell-derived type I interferon

Luisa Cervantes-Barragan et al. Blood. .

Abstract

This study demonstrates a unique and crucial role of plasmacytoid dendritic cells (pDCs) and pDC-derived type I interferons (IFNs) in the pathogenesis of mouse coronavirus infection. pDCs controlled the fast replicating mouse hepatitis virus (MHV) through the immediate production of type I IFNs. Recognition of MHV by pDCs was mediated via TLR7 ensuring a swift IFN-alpha production following encounter with this cytopathic RNA virus. Furthermore, the particular type I IFN response pattern was not restricted to the murine coronavirus, but was also found in infection with the highly cytopathic human severe acute respiratory syndrome (SARS) coronavirus. Taken together, our results suggest that rapid production of type I IFNs by pDCs is essential for the control of potentially lethal coronavirus infections.

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Figures

Figure 1
Figure 1
Type I IFN production and viral replication in MHV-infected splenic cDCs and pDCs. (A) Flow cytometric analysis of splenic cDCs (CD11c+PDCA-1−) and (B) splenic pDCs CD11clow B220+ PDCA-1− before FACS sorting. Gates for sorting are indicated. (C) Primary FACS-purified murine splenic cDCs or pDCs were infected with MHV at a multiplicity of infection (moi) of 1. IFN-α secretion to culture supernatants was determined by ELISA at the indicated time points. (D) Virus titers in culture supernatants were determined by plaque assay. (C-D) Data represent mean values ± SD pooled from 2 experiments. Statistical analysis was performed using Student t test (*P < .05; **P < .01).
Figure 2
Figure 2
Type I IFN production and viral replication in MHV-infected in vitro–derived cDCs and pDCs. Infection of bone marrow–derived pDCs and cDCs with MHV. (A,C) IFN-α and virus titers in tissue culture supernatants were determined at the indicated times after MHV infection (moi = 1), or (B,D) at 24 hours after MHV infection with different moi as indicated. Values in panels A-D represent means ± SD from triplicate measurements. Experiments in panels A-D were repeated 3 times with comparable results. Expression of IFN-β, IP-10, IFN-α4, IFN-α, GAPDH, or MHV nucleoprotein (MHV-N) mRNAs was determined by RT-PCR using total RNA from bone marrow–derived (E) cDCs or (F) pDCs infected with MHV (moi = 1) or treated with PBS (mock). (G-H) Viral replication in MHV-infected wt or IFNAR−/− pDCs. Cells were infected with MHV A59 at (G) an moi of 1 or (H) at different moi as indicated. Virus titers in culture supernatants were determined by plaque assay at (G) the indicated times after MHV A59 infection (moi = 1) or (H) 24 hours after MHV infection. (A-D,G-H) Data represent mean values ± SD pooled from 2 experiments. Statistical analysis was performed using Student t test (*P < .05; **P < .01, ***P < .001).
Figure 3
Figure 3
Impact of type I IFN signaling during MHV infection. IFNAR−/− or wt mice were injected intraperitoneally with 5 pfu MHV A59. (A) Health status of IFNAR−/− and wt mice was monitored twice daily after infection (n = 6). (B) ALT values were measured at the indicated time points after infection. (C) Liver pathology in IFNAR−/− and wt mice before or 48 hours after MHV A59 infection. Hematoxylin-eosin staining of 4% formaldehyde-fixed sections. Images were acquired using a Leica DMRA microscope (Leica, Heerbrugg, Switzerland) with a 25×/0.65 NA objective (total magnification, ×162). Images were processed using Adobe Photoshop (Adobe Systems, San Jose, CA). (D) Viral titers in liver, spleen, brain, and lung of MHV A59–infected IFNAR−/− or wt mice were determined at different time points after infection. Results represent the mean of 6 individual mice per time point. Solid horizontal lines in panel D represent limit of detection in the plaque assay. Data in panels B and D represent means ± SD from 2 experiments with a total of 3 or 6 mice evaluated per time point. Statistical analysis was performed using Student t test (ns, P > .05; *P < .05; **P < .01; ***P < .001).
Figure 4
Figure 4
pDCs sense MHV via TLR7. Bone marrow–derived pDCs from TLR3−/−, TLR7−/−, TLR3−/−/TLR7−/−, MyD88−/−, or wt mice were infected with MHV A59 (moi = 1) or treated with CpG oligonucleotides. IFN-α in tissue culture supernatants was determined 24 hours after infection by ELISA. Bars represent means with values from individual mice shown as open circles. Statistical analysis was performed using Student t test (**P < .01).
Figure 5
Figure 5
Effect of antibody-mediated pDC depletion on MHV infection. 129Sv mice were treated with rat IgG2b (wt isotype control) or α–mPDCA-1 (wt PDCA-1) and infected intraperitoneally with 5 pfu MHV A59 (n = 6). IFNAR−/− mice (n = 3) were used to demonstrate uncontrolled MHV spread in the absence of a functional IFN system. (A) IFN-α concentration in serum (means ± SD), (B) viral titers (means ± SD) in liver, spleen, brain, and lung, and (C) serum ALT values (means ± SD) were assessed at 48 hours after infection. (A-C) Statistical analysis was performed using Student t test (*P < .05; **P < .01).
Figure 6
Figure 6
Infection of human pDCs and cDCs with SARS-CoV. Primary pDCs or cDCs were isolated from peripheral blood of healthy donors and infected with either SARS-CoV (moi = 1) or Newcastle disease virus (NDV) as positive control (moi = 5), or were left uninfected (mock). (A) IFN-α in culture supernatants was determined at the indicated time points. Results represent pooled data (means ± SD) using pDCs and cDCs from 4 healthy donors. Statistical analysis was performed using Student t test (*P < .05). (B-C) Expression of IFN-β, ISG56, MxA, SARS-CoV N protein, and γ-actin mRNAs in (B) cDCs and (C) pDCs was determined by RT-PCR. One representative result of 4 individual experiments is shown.
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