Pegylated interferon-alpha protects type 1 pneumocytes against SARS coronavirus infection in macaques

Abstract

The primary cause of severe acute respiratory syndrome (SARS) is a newly discovered coronavirus. Replication of this SARS coronavirus (SCV) occurs mainly in the lower respiratory tract, and causes diffuse alveolar damage. Lack of understanding of the pathogenesis of SARS has prevented the rational development of a therapy against this disease. Here we show extensive SCV antigen expression in type 1 pneumocytes of experimentally infected cynomolgus macaques (Macaca fascicularis) at 4 d postinfection (d.p.i.), indicating that this cell type is the primary target for SCV infection early in the disease, and explaining the subsequent pulmonary damage. We also show that prophylactic treatment of SCV-infected macaques with the antiviral agent pegylated interferon-alpha (IFN-alpha) significantly reduces viral replication and excretion, viral antigen expression by type 1 pneumocytes and pulmonary damage, compared with untreated macaques. Postexposure treatment with pegylated IFN-alpha yielded intermediate results. We therefore suggest that pegylated IFN-alpha protects type 1 pneumocytes from SCV infection, and should be considered a candidate drug for SARS therapy.

Figure 1. Histological lesions and immunohistochemical and ultrastructural detection of SCV in lungs of experimentally infected macaques.

(a) DAD in pulmonary alveoli (control group). (b) Normal pulmonary alveoli (prophylactic group). (c,d) SCV antigen expression in type 1 pneumocytes (control group). (e) Lack of SCV antigen expression in pulmonary alveoli (prophylactic group). (f) Viral nucleocapsids in cytoplasmic vesicle of type 1 pneumocyte (control group), showing pinocytotic vesicles (arrowheads) and basement membrane (arrows). Scale bar, 1 μm. (g) Higher magnification of viral nucleocapsids shown in f. Scale bar, 200 nm. (h) SCV antigen expression in type 2 pneumocytes (control group). Sections were stained with H&E (a,b), for SCV antigen (c–e,h), or with uranyl acetate and lead citrate (f,g). Original magnification, ×25 (a left, b left), ×100 (a right, b right, c,e) or ×250 (d,h).

Figure 2. Antiviral activity of pegylated IFN-α against SCV in vitro and its biological activity in macaques.

(a) Effect of pegylated IFN-α against SCV infection in vitro. Similar results were obtained in three separate experiments. (b) Pharmacokinetic analysis of pegylated IFN-α (b) and induction of neopterin (c) in macaques treated with PBS (□, control group; n = 4) or pegylated IFN-α (▪, prophylactic group; n = 4) on days −3 and −1. Data are expressed as mean ± s.d. ^**, P < 0.01 compared with control.

Figure 3. Effect of pegylated IFN-α on SCV excretion in macaques.

SCV detection in pharyngeal swabs taken at 0, 2 or 4 d.p.i. from macaques treated with PBS (control group; n = 4) or pegylated IFN-α prophylactically (n = 6) or postexposure (n = 4). Eq, equivalents. Data are expressed as mean ± s.d. ^*, P < 0.05; ^**, P < 0.01 compared with control group at 2 d.p.i.

Figure 4. Effect of pegylated IFN-α on SCV replication, viral antigen expression and histological lesions in lungs of SCV-infected macaques at 4 d.p.i.

(a) SCV titration of lung homogenates. TCID₅₀, median tissue culture infectious dose. (b) Immunohistochemical detection of SCV-infected cells in lung sections. (c) Histopathological scoring of lung sections. SCV-infected macaques were treated with pegylated IFN-α prophylactically (n = 4) or postexposure (n = 4), or with PBS (control group; n = 4). Data are expressed as mean ± s.d. ^*, P < 0.05; ^**, P < 0.01 compared with control group.