Mucous membrane surfaces, respiratory epithelium, and skin are portals of entry for many viruses. Since viral infections depend on host cellular machinery for survival and replication, entry into cells is a critical early step. This is accomplished by a variety of mechanisms, some of which are common to many viruses and others are virus specific. Examples of mechanisms shared by multiple viruses are fusion with cell membrane (enveloped viruses) and/or endocytosis after binding to cell-surface molecules. Many cell-surface molecules are exploited by viruses to gain entry into cells to facilitate viral replication. Expression of such molecules is cell-type specific and can determine tissue tropism for certain viruses. Examples of exploited host cellsurface molecules include CD4, CCR5, and CXCR4 for HIV; sialic acid residues for influenza and other respiratory viruses; heparan sulfate and proteoglycans for herpesviruses; and coxsackie adenovirus receptor (CAR), and B cell receptors CD21 and C3d for Epstein Barr Virus (2). Once in contact with cells, viral replication and the host antiviral immune response begins. Replication of lytic viruses can directly injure mucosal and epithelial protective barriers by causing cell lysis upon release of viral particles (Table 84.1). Via killing of virus-infected cells, expression of inflammatory mediators, and leukocyte infiltration, the antiviral immune response can cause further tissue injury releasing damage-associated molecules (DAMPs) that further activate the immune system (3). The immune response to microbial pathogens consists of several phases. The earliest and least specific is the innate immune response followed by a pathogen-specific adaptive immune response leading to immunologic memory. These phases of immune responses include activation of diverse cellular and humoral mediators—many of which are common to many types of infections. The immune response to viral infection involves mechanisms similar to those induced by bacterial infections as well as unique mechanisms. Recognition of viruses by the innate immune system begins by the host recognition of viral pathogen-associated molecular patterns (PAMPs) expressed during viral replication through their interaction with pattern recognition receptors (PRRs), such as the Toll-like receptors (TLRs). Viral PAMPs include virion proteins, hemagglutinin (HA), double-stranded RNA produced during replication of many viruses, F protein from RSV, single-stranded RNA, and viral DNA (4,5). TLR 2, 3, 4, 7, 8, and 9 have been implicated in responses to viral PAMPs. In fact, the initial cellular reactions to viral PAMPs are very similar to those initiated by bacterial PAMPs and are depicted in Figure 84.2 (4). The innate response is critical for recruitment of effector cells, containment of viral particles, and the initiation of adaptive or antigen-specific immune response necessary for viral eradication and immunologic memory. Binding to TLRs initiates intracellular signal transduction leading to production of proinflammatory cytokines, including type I interferons α and β (IFN-α/β), type III interferons (IFN-λ subtypes), TNF-α, IL-1β, IL-6, and chemokines, such as CCL2, CCL20, and CCR7, which facilitate trafficking of alveolar macrophages, dendritic cells (DCs), and neutrophils to sites of viral invasion (6). Anti-inflammatory cytokines, such as IL-10, soluble TNF-α receptor, and IL-1 receptor antagonist, are also induced. TLR-independent mechanisms for cytoplasmic viral detection also exist including activation of nucleotide-binding and oligomerization domain-like receptors (NLRs) and helicases including retinoic acid-inducible protein (RIG-1). NLRs are a family of multimeric cytosolic structures ultimately capable of converting pro-caspase 1 to its functional form leading to conversion of proforms of IL-1β and IL-18 to their active forms (7). A role for NLRP3 in influenza and RSV has been demonstrated, and more data are emerging to implicate other types of NLRs. The precise activators of NLRP3 are being elucidated; however, it appears that they include conventional PAMPs (dsRNA) as well as reactive oxide species and potassium (K) flux induced during viral replication (7,8,9). In contrast, RIG-1 is activated by intracellular viral dsRNA produced

during the replication of many viruses. Activation of IRF3 and IRF7 are critical for induction of type 1 interferons (IFN-α /β) that are essential antiviral cytokines (10). These pleiotropic cytokines can affect viral replication by reducing “viral receptor” molecule expression on the host cell surface, inhibition of transcription and translation of viral proteins through IFNα /β-stimulated genes such as MxA protein, 2′5′-oligoadenylsynthase, double-stranded RNA kinase (PKR), and eukaryotic initiation factor α (eIF-2α), thereby inhibiting viral replication (5,6). In addition to viral proteins, RNA/DNA-mediated activation of the above pathways and DAMPs released from injured tissue including lysed epithelial cell contents can activate innate immune pathways via their respective TLR, RLR, or inflammasomes (10).

IFN-α/β can activate NK cells and macrophages, induce maturation of DCs, and upregulate proteins important in antigen presentation to T cells. Activated CD8 cells differentiate into cytotoxic T cells (CTLs) that contribute to lysis of virusinfected cells and to viral clearance. Macrophages and DCs can bind de novo synthesized intracellular viral peptides via the major histocompatibility complex I (MHC I) and extracellular viral peptides via MHC II. Once activated by viruses, DCs mature as they migrate to draining lymph nodes where they present viral antigens to CD4 T cells via MHC I and CD8 cells via MHC II (5,6). Antigen-activated CD4 cells differentiate to Th1 (proinflammatory), Th2 (anti-inflammatory), regulatory T cells (Tregs), or Th17 (produces the family of IL-17 proteins [IL-17A-D, IL-17E (IL-25), IL-17F], as well as IL-21 and IL-23) depending on the inflammatory milieu (3). Cytotoxic CD8 T cells can eliminate virus by further induction of another essential antiviral cytokine, IFN-γ, by lysis of virus-infected cells via the release of perforin (a membrane pore-forming protein important for cytotoxicity), and by induction of apoptosis through Fas ligand with Fas (CD95) on the virus-infected cells (3,6,11). A result of this immune cascade ideally is eradication of the virus, production of antibodies specific for multiple viral peptides, and creation of memory T cells and B cells important in protection against subsequent infection with the same virus.

IFN-α/β is a critical antiviral cytokine and in the absence of its signaling viral replication is unchecked, and overwhelming viral infection occurs. In chronic viral infection, a lower but sustained IFN-α/β response continues, and despite this, the virus persists. Very cutting edge data have revealed a paradox in IFN-α/β function regarding the role of IFN-α/β in chronic viral infection (12). Lymphocytic choriomeningitis virus (LCMV) typically causes a self-limited mild illness (aseptic meningitis); however, some strains cause chronic viral infection in mice. Blockade of IFN-α/β after the onset of chronic LCMV infection reversed findings seen in chronic viral infection, such as a hyperimmune state, lymphoid tissue destruction resulting in CD4, and IFN-γ-mediated clearance of virus (12,13,14). Although this is an animal model, its relevance to human disease may be very significant because evidence of IFN-α /β pathway activation in hepatitis C is associated with disease progression and poor response to IFN therapy. Similar findings occur in HIV and latent tuberculosis (12). If these findings hold true in the clinical setting, the use of IFN-α in chronic viral infection may need to be reexamined.

Typically, homeostasis in the lung is maintained by a number of mechanisms, and this homeostasis is important to lung protection as the respiratory tract is continually being challenged by inhaled particulate matter. Immunosuppressive Tregs express the master control transcription factor Forkhead box P3 (FOXP3+) and are critical to this process (10). In RSV and influenza, Tregs are activated and secrete excessive IL-10, thereby dampening Th1 proinflammatory responses. Deletion of these cells in experimental models leads to exaggerated pulmonary inflammation and lung injury (6,10,15). Other cell types that contribute to dampening of the inflammatory response incited during viral infections include some plasmacytoid DCs, neutrophils by facilitating viral clearance, and airway epithelial cells (AECs) via expression of CD200. CD200 receptors are highly upregulated on airway macrophages during influenza, and ligation leads to suppression of alveolar macrophage response to influenza. Latent TGF-β that is constitutively produced by AECs can be activated by influenza neuraminidase (NA), also resulting in dampening of pulmonary inflammatory responses to viral infection (6,15,16,17). When the virus is eradicated and the proinflammatory response is resolving, an anti-inflammatory environment evolves to promote healing. Th2 cytokines IL-4 and IL-13 are important mediators and IL-13 is critical to changing the phenotype of macrophages to “alternatively activated macrophages” (M2) that are anti-inflammatory and important to tissue repair (10).

In summary, in addition to the primary proinflammatory responses to respiratory viral infections, negative regulatory mechanisms are activated to limit immune-mediated tissue injury. In simple viral infection, these mechanisms are key to tissue repair; however, they create vulnerability to bacterial infection because of their immune-dampening effects.

Elegant data have demonstrated that during respiratory viral infection there is disruption of protective epithelial barriers as a result of diversion of cell machinery to intracellular viral replication often with direct lysis of infected epithelial cells (18). When protective epithelial barriers are disrupted, mucociliary dysfunction can occur as well as exaggerated bacterial colonization by adherence to exposed basement membrane elements permitting penetration into deeper tissues (see Table 84.1). This concept was demonstrated in experimental human inoculation with influenza followed by the development of detectable airway colonization with Streptococcus pneumoniae 6 days after inoculation with influenza (23). In addition, several studies have demonstrated significantly increased nasopharyngeal colonization with commensal organisms during other respiratory
viral infections (parainfluenza, RSV, and adenovirus) when compared to baseline colonization in the same children (23). Specifically, bacterial adherence is enhanced in areas where epithelium has been denuded. This has been seen in many experimental settings with cell culture and in vivo animal models. In addition, during the 1957 influenza epidemic at autopsy, direct visualization of Staphylococcus aureus in the lungs adherent to areas of denuded tracheobronchial epithelium was seen (18,24,25).