III. ABSTRACTS AND POSTERS / ORAL PRESENTATIONS
6. Mice deficient in interferon regulatory factor 4 (IRF4) are more susceptible to infection with mouse-adapted influenza A/Aichi/2/68
2.1.2 Severe Acute Respiratory Syndrome coronavirus (SARS-CoV)
The severe acute respiratory syndrome (SARS) was unprecedented in the rapidity and extent of its spread, in the magnitude of its impact on health systems and economies. A total of 774 deaths were reported out of 8,096 SARS-CoV infection cases in 29 countries.
The cellular entry of the SARS-CoV involves the S protein and ACE2 receptor.
The spike (S) protein of SARS-CoV is a type 1 transmembrane glycoprotein(Leth- Larsen, Zhong et al. 2007). The S1 subunit is responsible for virus binding to the Angiotensin-Converting Enzyme 2 (ACE2) receptor (Li, Moore et al. 2003, Prabakaran, Xiao et al. 2004) where the ACE2 receptor-binding domain lie within amino acids (aa) 318-510 of the S1 subunit (Dimitrov 2003, Xiao, Chakraborti et al.
2003, Wong, Li et al. 2004). The S2 subunit contains a putative fusion peptide and two heptad repeats HR1 and HR2. The structure is also responsible for the fusion of the viral and target cell membranes. Four regions with highly immune reactivity are located at 67-119 aa, 265-345 aa, 588-645 aa and 1120-1234 aa (Wang, Chen et al.
2004, Wang, Sin et al. 2004). SARS-CoV may also gain entry into cell through pH- dependent endocytosis, mediated by the S protein (Yang, Huang et al. 2004).
Additionally, it has been reported that the S protein-driven cell-to-cell fusion can also occur in the absence of low pH (Dimitrov 2003). It is possible that the S protein may be able to mediate membrane fusion in both pH-dependent and –independent manner.
The ACE2 metallopeptidase was originally thought to have direct effects on cardiac function (Boehm and Nabel 2002), but was discovered to be a functional receptor for SARS-CoV (Li, Moore et al. 2003, Wang, Chen et al. 2004) by binding to the S protein. Besides being expressed predominantly in vascular endothelial cells of the heart and the kidneys, it is present in human lung alveolar Type I and Type II
14 epithelial cells, enterocytes of the small intestine, the brush border of the proximal tubular cells of the kidney and endothelial cells of arteries, veins and arterial smooth muscle cells in several other organs (Hamming, Timens et al. 2004). ACE2 is not expressed on B or T cells, macrophages in spleen of lymphoid organs. The localization of ACE2 may explain the tissue tropism of SARS-CoV in the lung, small intestine and kidney (Ding, He et al. 2004).
2.1.2.1 Innate immune responses to SARS-CoV infection
In clinical cases of SARS infections, the host immune response undergoes several unique events: lymphopenia (occurring progressively in the early course of the infection, reaching the minimum during the second week of the infection), production of specific antibodies (IgG persisted indefinitely while IgM expressed transiently) together with a distinct cytokine profile (increased in levels of IFN-γ and IL-10 whereas decreased in IL-4 levels) (Zhu 2004). Innate immunity functions as the first host defense against viral infections. The key componetns include natural killer (NK) cells, the IFN response, chemokines and cytokines.
NK cells in peripheral blood form clinical SARS cases correlated with the severity of disease and the presence of antibodies against the virus (SARS 2004).
Antiviral IFN response is mediated by IFN production and signaling or by direct inactivation of effector molecules. In SARS-CoV infection, there is an unusual lack of an antiviral IFN response (Chen and Subbarao 2007). SARS-CoV is thought to have developed mechanisms necessary to block activation of IFN regulatory pathways at the initial stages following the nuclear transport of the IFN regulatory factor-3 (IRF-3) (Spiegel, Pichlmair et al. 2005). The IRF family of transcriptional factors has diverse roles, among them the regulation of host defense in the innate and adaptive immune
15 responses. IRF-3 is primarily responsible for the activation of Type I IFNs (IFN-α and IFN-β). Clinical data suggests that these IFNs were undetectable in patients and neither was it induced in infected cells in vitro. The SARS-CoV infection upregulates the expression of chemokines IP-10, MCP-1, MIP-1α and RANTES in macrophages and dentritic cells (Cheung, Poon et al. 2005, Law, Cheung et al. 2005, Yilla, Harcourt et al. 2005). Levels of IP-10, IL-8 and MCP-1 were found to be elevated in lungs and peripheral blood of patients where IL-6, IL-8 and MCP-1 were found in lungs of fatal cases (Jiang, Xu et al. 2005). There was not change in TNF-α level in clinical samples (Wong, Lam et al. 2004).
2.1.2.2 Adaptive cellular responses to SARS-CoV infection
A rapid development of lymphopenia is characteristics of the adverse outcome of the disease, with CD4+ T cells being more severely reduced than CD8+ T cells during acute infection (He, Zhao et al. 2005). Progressive lymphopenia occurred in the early course of the disease and reached its lowest point in the second week in most cases. It has been hypothesized that the cause may be related to virus-induced infection and destruction of lymphocytes, chemokine-mediated lymphocyte trafficking/redistribution and sequestration of lymphocytes in the lung, bone marrow suppression or apoptosis (O'Donnell, Tasker et al. 2003, Panesar 2003, Wong, To et al.
2003).
IFN-γ is known to increase over the progression of a SARS-CoV infection (Zhu 2004). This Th1 cytokine is associated with potent cell-mediated immunity and resistance to intracellular pathogens. However, IL-4 levels were found to be decreased after the onset of the infection. This indicated that a Th1 dominant response is responsible for the elimination of the virus from the body as IL-4 is a dominant Th2
16 cytokine which also promotes humoral immunity. On the other hand, IL-10 was found to be elevated. As IL-10 is produced by Th2 cells and has dual effect on T lymphocytes in suppressing Th1 cells from producing IL-2, IFN-γ and TNF while promoting proliferation and cytotoxicity of CD8+ T cells and NK cells. IL-10 expression increase may be associated with the susceptibility of the infection (Zhu 2004)
2.1.2.3 Humoral immune response to SARS-CoV infection
Clinical cases reveal that humoral responses towards the virus became apparent during the convalescent phase. Serum IgG, IgM and IgA occurred around with same time with most cases seroconverting by day 14 after illness onset (Hsueh, Huang et al. 2004). IgG levels persisted for a long time but IgM was detected to be present for a shorter time. This suggested that IgG antibody against the virus represents the primary humoral immune response for protection. Among the structural proteins, only the S protein elicits neutralizing antibody (Buchholz, Bukreyev et al.
2004), with the major S protein immunodominant epitope existing between amino acids 441 and 700 (Lu, Manopo et al. 2004).