Severe acute respiratory syndrome (SARS) was first identified in Guangdong Province in China (28). Over the ensuing 9 months, more than 8,000 cases were identified throughout the world, with a ϳ10% case fatality rate. A novel coronavirus, SARS coronavirus (SARS-CoV), was identified as the causative agent (6,17,29,32). Initial investigations indicated that the virus spread to humans from infected exotic animals such as Himalayan palm civets (Paguma larvata) and Chinese ferret badgers (Melogale moschata) (12); more recent work has suggested that the natural reservoirs for the virus are wild bat populations in China (19,24). Although SARS has not recurred in human populations to a significant extent since 2003, the potential severity of such a recurrence has spurred interest in developing an animal model for the human disease.SARS-CoV infects and replicates in mice, ferrets, hamsters, and several species of nonhuman primates (cynomolgus and rhesus macaques, African green monkeys, and common marmosets) (reviewed in reference 37). However, none of these animals develop a clinical disease that is reproducible and equivalent in severity to that observed in SARS patients. A mouse model would be useful for answering many questions about SARS pathogenesis and for testing vaccine efficacy, in part because reagents for the study of the immune response are widely available. However, other than aged or immunocompromised (STAT1 Ϫ/Ϫ ) mice (37), these animals do not develop significant clinical disease, and lethality has not been demonstrated in any murine model of SARS. With the goal of developing a more robust murine model, we generated transgenic (Tg) mice in which expression of hACE2 (human angiotensin-converting enzyme 2, the primary host cell receptor for SARS-CoV [23]) was targeted to epithelial cells. While human ACE2 and murine ACE2 (mACE2) molecules are very homologous, mACE2 does not support SARS-CoV binding as efficiently as hACE2 (22). Here we show that the transgenic expression of hACE2 in epithelia converts a mild SARS-CoV infection into a rapidly fatal disease. MATERIALS AND METHODSMice. All animal studies were approved by the University of Iowa and the Veterans Administration Institutional Animal Care and Use committees. Mice transgenic for expression of hACE2 (K18-hACE2 mice) were generated as follows (see Fig. 1A). The hACE2 coding sequence was PCR amplified from IMAGE consortium clone ID 5243048 (ATCC, Manassas, VA) and cloned into the pCR2.1-TOPO vector (Invitrogen, Carlsbad, CA). The lacZ coding sequence in the previously described pK18mTElacZ-K18i6x7pA construct (16) (a kind gift from Jim Hu, Hospital for Sick Children, Toronto, Canada) was then replaced by the hACE2 coding sequence to create pK18-hACE2. 5Ј of the hACE2 coding sequence, this plasmid contains 2.5 kb of upstream genomic sequence, the promoter, and the first intron (with a mutation in the 3Ј splice acceptor site to reduce exon skipping) of the human cytokeratin 18 (K18) gene as well as a translational enhancer sequence from alfalfa mosaic vi...

Studies of patients with severe acute respiratory syndrome (SARS) demonstrate that the respiratory tract is a major site of SARS-coronavirus (CoV) infection and disease morbidity. We studied host-pathogen interactions using native lung tissue and a model of well-differentiated cultures of primary human airway epithelia. Angiotensin converting enzyme 2 (ACE2), the receptor for both the SARS-CoV and the related human respiratory coronavirus NL63, was expressed in human airway epithelia as well as lung parenchyma. As assessed by immunofluorescence staining and membrane biotinylation, ACE2 protein was more abundantly expressed on the apical than the basolateral surface of polarized airway epithelia. Interestingly, ACE2 expression positively correlated with the differentiation state of epithelia. Undifferentiated cells expressing little ACE2 were poorly infected with SARS-CoV, while well-differentiated cells expressing more ACE2 were readily infected. Expression of ACE2 in poorly differentiated epithelia facilitated SARS spike (S) proteinpseudotyped virus entry. Consistent with the expression pattern of ACE2, the entry of SARS-CoV or a lentivirus pseudotyped with SARS-CoV S protein in differentiated epithelia was more efficient when applied to the apical surface. Furthermore, SARS-CoV replicated in polarized epithelia and preferentially exited via the apical surface. The results indicate that infection of human airway epithelia by SARS coronavirus correlates with the state of cell differentiation and ACE2 expression and localization. These findings have implications for understanding disease pathogenesis associated with SARS-CoV and NL63 infections.

Plasmodium infection of erythrocytes induces clinical malaria. Parasite-specific CD4+ T cells correlate with reduced parasite burdens and severity of human malaria, and are required to control blood-stage infection in mice. However, the characteristics of CD4+ T cells that determine protection or parasite persistence remain unknown. Here we show that P. falciparum infection of humans increased expression of an inhibitory receptor (PD-1) associated with T cell dysfunction. In vivo blockade of PD-L1 and LAG-3 restored CD4+ T cell function, amplified T follicular helper cell and germinal center B cell and plasmablast numbers, enhanced protective antibodies and rapidly cleared blood-stage malaria in mice. Thus, chronic malaria drives specific T cell dysfunction, which can be rescued to enhance parasite control using inhibitory therapies.

A chronic demyelinating disease results from murine infection with the neurotropic strain JHM of mouse hepatitis virus (MHV-JHM). Demyelination is largely immune mediated. In this study, the individual roles of CD4 and CD8 T cells in MHV-induced demyelination were investigated using recombination-activating gene 1−/− (RAG1−/−) mice infected with an attenuated strain of MHV-JHM. These animals develop demyelination only after adoptive transfer of splenocytes from mice previously immunized to MHV. In this study, we show that, following adoptive transfer, virus-specific CD4 and CD8 T cells rapidly infiltrate the CNS of MHV-JHM-infected RAG1−/− mice. Adoptive transfer of CD4 T cell-enriched donors resulted in more severe clinical disease accompanied by less demyelination than was detected in the recipients of undepleted cells. Macrophage infiltration into the gray matter of CD4 T cell-enriched recipients was greater than that observed in mice receiving undepleted splenocytes. In contrast, CD8 T cell-enriched recipients developed delayed disease with extensive demyelination of the spinal cord. MHV-JHM-infected RAG1−/− mice receiving donors depleted of both CD4 and CD8 T cells did not develop demyelination. These results demonstrate that the development of demyelination following MHV infection may be initiated by either CD4 or CD8 T cells. Furthermore, they show that CD4 T cells contribute more prominently than CD8 T cells to the severity of clinical disease, and that this correlates with increased macrophage infiltration into the gray matter.

Summary Inducing memory CD8+ T-cells specific for conserved antigens from Influenza A virus (IAV) is a potential strategy for broadly protective vaccines. Here we show that memory CD8 T-cells in the airways played an important role in early control of IAV. Expression of chemokine receptor CXCR3 was critical for memory CD8 T-cells to populate the airways during the steady state and vaccination approaches were designed to favor the establishment of memory CD8 T-cells in the airways. Specifically, we found that interleukin-12 (IL-12) signaling shortly after immunization limited CXCR3 expression on memory CD8 T-cells. Neutralization of IL-12 or adjuvants that did not induce high amounts of IL-12, enhanced CXCR3 expression, sustained airway localization of memory CD8 T-cells and resulted in superior protection against IAV.