Chapter 49Nonspecific Defenses

Dianzani F, Baron S.

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General Concepts

Most viral infections are limited by defenses that are antigen nonspecific and/or specific. Nonspecific defenses act sooner than specific defenses. Some are always in place (anatomic barriers, nonspecific inhibitors, and phagocytic cells); others are evoked by the infection (fever, inflammation, and interferon).

Anatomic Barriers

Anatomic barriers are located at body surfaces (skin and mucosa) or within the body (endothelial cells and basement membranes). They are partly effective in preventing virus spread but may be breached by large numbers of virus, by trauma, by increased permeability, by replication of virus in endothelial cells, or by transportation of virus in leukocytes.

Nonspecific Inhibitors

Body fluids and tissues normally contain soluble viral inhibitors. Most prevent viral attachment, some directly inactivate viruses, and others act intracellularly. These inhibitors may be overwhelmed by sufficient virus.

Phagocytosis

Viruses may be phagocytosed to different degrees by polymorphonuclear leukocytes and macrophages. The effect of phagocytosis may be virus inactivation, persistence, or multiplication; consequently, the result may be clearance of virus, transportation to distant sites, or enhanced infection.

Fever

Replication of most viruses is reduced by even a modest rise in temperature. During viral infection, fever can be initiated by several endogenous pyrogens, such as interleukins-1 and -6, interferon, prostaglandin E2, and tumor necrosis factor.

Inflammation

Inflammation inhibits viral replication through (1) elevated local temperature, (2) reduced oxygen tension, (3) metabolic alterations, and (4) acid production. The effects of these mechanisms are often additive.

Viral Interference and Interferon

Viral interference occurs when infection by one virus renders cells resistant to the same or other superinfecting viruses. Interference is usually mediated by newly induced host cell proteins designated as the interferon systems. Secreted interferon binds to cells and induces them to block various stages of viral replication. Interferon also (1) inhibits growth of some normal and tumor cells and of many intracellular parasites, such as rickettsiae and protozoa; (2) modulates the immune response; and (3) affects cell differentiation. There are three main types of interferon, alpha, beta, and gamma interferons. Alpha interferon is produced mainly by certain leukocytes (dendritic cells, macrophages and B cells), beta interferon by epithelial cells and fibroblasts, and gamma interferon by T and natural killer cells. Two other interferon types are related to alpha interferon. Omega interferons share about seventy percent identity with alpha interferons. Tau interferons also are related structurally to alpha interferons but are unusual by (a) being produced for only a few days by normal placental trophoblasts and (b) not being inducible by viruses.

Introduction

Most viral infections are limited by nonspecific defenses, which (1) restrict initial virus multiplication to manageable levels, (2) initiate recovery from established infections that is then completed by a combination of these early nonspecific and subsequent antigen-specific immune defenses, and (3) enable the host to cope with the peak numbers of virus that, if presented as the infecting dose, could be lethal. Although immune and nonimmune (nonspecific) defenses operate together to control viral infections, this chapter considers only nonspecific defenses. Some nonspecific defenses exist independently of infection (e.g., genetic factors, anatomic barriers, nonspecific inhibitors in body fluids, and phagocytosis). Others (e.g., fever, inflammation, and interferon) are produced by the host in response to infection. All nonspecific defenses begin to act before the specific defense responses develop and can potentiate some of the established immune effector mechanisms.

The fact that viruses replicate intracellularly and the ability of some viruses to spread by inducing cell fusion partly protect viruses against such extracellular defenses as neutralizing antibody, phagocytosis, and nonspecific inhibitors. However, because they replicate within the cell, viruses are vulnerable to intracellular alterations caused by host responses to infection. Nonspecific responses that alter the intracellular environment include fever, inflammation, and interferon.

These multiple defenses function with great complexity because of their interactions with one another. This complexity is compounded by the varying effectiveness of the defenses that results from the diversity of viruses, hosts, and sites and stages of infection.

Defense Mechanisms that Precede Infection

Anatomic Barriers

Anatomic barriers to viruses exist at the body surfaces and within the body. At the body surfaces, the dead cells of the epidermis and any live cells that may lack viral receptors resist virus penetration and do not permit virus replication. However, this barrier is easily breached, for example, by animal bites (rabies virus), insect bites (togaviruses), and minor traumas (wart virus). At mucosal surfaces, only the mucus layer stands between invading virus and live cells. The mucus layer forms a physical barrier that entraps foreign particles and carries them out of the body; it also contains nonspecific inhibitors (see following section). The mucus barrier is not absolute, however, since sufficient quantities of many viruses can overwhelm it and infect by this route. In fact, most viruses use mucous surfaces as the portal of entry and initial replication site.

Within the body, anatomic barriers to virus spread are formed by the layer of endothelial cells that separates blood from tissues (e.g., the bloodbrain barrier). Under normal conditions, these barriers have a low permeability for viruses unless the virus can penetrate them by replicating in the capillary endothelial cells or in circulating leukocytes. These internal barriers may explain, in part, the high level of viremia required to infect organs such as the brain, placenta, and lungs.

Nonspecific Inhibitors

A number of viral inhibitors occur naturally in most body fluids and tissues. They vary chemically (lipids, polysaccharides, proteins, lipoproteins, and glycoproteins) and in the degree of viral inhibition and types of viruses affected. Some inhibitors are related to the viral receptors of the cell surface, but most are of unknown origin. Many inhibitors act by preventing virus from attaching to cells, others by directly inactivating virus, and a few by inhibiting virus replication. In the gastrointestinal tract, some susceptible viruses are inactivated by acid, bile salts, and enzymes. Whereas most inhibitors block only one or a few viruses, some have a broad antiviral spectrum. Although the effectiveness of the inhibitors has not been fully established in vivo, their importance as host defenses is suggested by their antiviral activity in tissue culture and in vivo and by the direct correlation between the degree of virulence of some viruses and their degree of resistance to certain inhibitors. Examples are the serum and mucus inhibitors of influenza viruses during experimental infections. However, even sensitive viruses may overwhelm these inhibitors when the infecting dose of virus is sufficiently high. Therefore, the presence of these inhibitors may explain the relatively high dose of virus required to initiate infection in vivo, compared with the dose needed in cell cultures.

Phagocytosis

The limited information available suggests that phagocytosis is less effective against viral infections than against bacterial infections. However, few of the factors that control uptake of virions or infected cells by phagocytes and their digestion by lysosomal enzymes have been studied systemically. Different viruses are affected differently by the various phagocytic cells. Some viruses are not engulfed, whereas others are engulfed but may not be inactivated. In fact, some viruses, such as human immunodeficiency virus (HIV), may even multiply in the phagocytes (e.g., macrophages), which may serve as a persistent reservoir of virus (Fig. 49-1). The virulence of several strains of HIV and herpesviruses correlates with their ability to multiply in macrophages. Infected macrophages may carry virus across the blood-brain barrier. Interestingly, cytomegalovirus has been reported to replicate in granulocytes. Macrophages seem to be more effective against viruses than are granulocytes, and some viruses seem to be more susceptible to phagocytosis than others. Macrophages and polymorphonuclear leukocytes can afford important protection by markedly reducing the viremia caused by virus strains susceptible to phagocytosis.

Figure 49-1. Possible outcomes of phagocytosis of a virus.

Figure 49-1

Possible outcomes of phagocytosis of a virus.

Viruses may stimulate macrophages to produce monokines, which can reduce viral multiplication. For example, macrophage-produced alpha interferon (IFN-α) inhibits viral multiplication both directly and also indirectly by activating natural killer cells. Interleukin 1 (IL-1), produced by macrophages, can interfere with viral multiplication in a number of ways: (1) by inducing T lymphocytes to produce interleukin-2, which in turn induces gamma interferon (IFN-γ), which can induce alpha and beta interferons; (2) by inducing the production of beta interferon (IFN-β) by fibroblasts and epithelial cells; (3) by inducing fever, which inhibits viral replication; (4) by enhancing macrophage-mediated cytolysis of infected cells; and (5) by inducing production of tumor necrosis factor (TNF), which inhibits virus multiplication both directly and indirectly by inducing interferon and other cytokines and augmenting inflammation, phagocytosis and cytotoxic activity. Therefore, depending on the situation, macrophages acting as phagocytes may reduce the number of viruses, help spread the infection, augment or depress immune defenses, or have little effect.

Defense Mechanisms Evoked by Infection

Fever

Viral replication is influenced strongly by temperature. Fever can be induced during viral infection by at least three independent endogenous pyrogens: interleukins-1 and 6, interferon, prostaglandin E2, and tumor necrosis factor. Even a modest increase can cause strong inhibition: a temperature rise from 37°C to 38°C drastically decreases the yield of many viruses. This phenomenon has been observed in tissue culture as well as in many experimental (including primate) and natural infections. Artificial induction of fever reduces mortality in mice infected with viruses (Fig. 49-2). Artificial lowering of the temperature during infection may increase mortality, as in suckling mice infected with coxsackieviruses and taken away from the warmth of their mother's nest. Fever also augments the generation of cytotoxic T lymphocytes.

Figure 49-2. Protection of mice by elevated temperature or antibody administered before or after intracerebral infection with the picornavirus EMC type.

Figure 49-2

Protection of mice by elevated temperature or antibody administered before or after intracerebral infection with the picornavirus EMC type.

Several observations suggest strongly that fever reduces virus multiplication during human viral infections. Retrospective studies have shown that the incidence and severity of paralysis among children infected with polioviruses were significantly greater in patients treated with antipyretic drugs (e.g., aspirin) than in untreated children. Also consistent with these findings is the observation that virus strains that replicate best at fever temperature are usually virulent, whereas virus strains that replicate poorly at fever temperature are usually low in virulence and therefore often are used as live virus vaccines.

Temperatures as low as 33°C are normal at body surfaces exposed to air; viruses that infect these sites and replicate optimally at these temperatures establish only local infections that do not spread to deeper tissues, where the body temperature is higher. For example, rhinoviruses that cause common colds replicate optimally at 33°C to 34°C (found in normally ventilated nasal passages); however, they are inhibited at 37°C (found when swelling of the edematous mucosa and secretions interrupt air flow). An interesting question is whether this temperature increase is important for recovery from coryza. The same general considerations of temperature probably apply to other human viral infections such as measles, rubella, and mumps, although, unfortunately, suitable and controlled studies have not been conducted. Nevertheless, available information suggests that antipyretic drugs be used conservatively.

Inflammation

Several antiviral mechanisms are generated by the local inflammatory response to virus-induced cell damage or to virus-stimulated mediators such as activated complement. The major components of the inflammatory process are circulatory alterations, edema, leukocyte accumulation and perhaps prostaglandins A and J. The resulting phenomena are elevated local temperature, reduced oxygen tension in the involved tissues, altered cell metabolism, and increased levels of CO2 and organic acids. All of these alterations, which occur in a cascading and interrelated fashion, drastically reduce the replication of many viruses. For instance, the altered energy metabolism of the infected and surrounding cells, as well as the accumulating lymphocytes, can generate local hyperthermia. At superficial sites where the temperature is normally lower, hyperthermia can also be generated by hyperemia during the early stages of inflammation. As inflammation progresses, hyperemia becomes passive, thereby greatly reducing blood flow and decreasing oxygen tension. Two factors account for this decrease in oxygen tension: limited influx of erythrocytes, and lower diffusion of oxygen through edema fluid. In turn, the decreased oxygen tension causes less ATP production, thus reducing the energy available for viral synthesis and increasing anaerobic glycolysis, which increases the accumulation of CO2 and organic acids in the tissues. These acid catabolites may decrease the local pH to levels that inhibit the replication of many viruses. Local acidity also may increase by accumulation and subsequent degradation of the leukocytes in the affected area. It is possible that other less well-defined factors are also significant .

Therefore, the local inflammation resulting from viral infection clearly activates several metabolic, physicochemical, and physiologic changes; acting individually or together, these changes interfere with virus multiplication. Although further animal and human studies are required, this interpretation is supported by the finding that anti-inflammatory drugs (corticosteroids) often increase the severity of infection in animals. Therefore, these drugs should be used with caution in treating viral diseases.

Viral Interference and Interferon

Viral Interference

Generally, infection by one virus renders host cells resistant to other, superinfecting viruses. This phenomenon, called viral interference, occurs frequently in cell cultures and in animals (including humans). Although interference occurs between most viruses, it may be limited to homologous viruses under certain conditions. Some types of interference are caused by competition among different viruses for critical replicative pathways (extracellular competition for cell surface receptors, intracellular competition for biosynthetic machinery and genetic control). Similar interference may result from competition between defective (nonmultiplying) and infective viruses that may be produced concurrently. Another type of interference—the most important type in natural infections—is directed by the host cells themselves. These infected cells may respond to viral infection by producing interferon proteins, which can react with uninfected cells to render them resistant to infection by a wide variety of viruses.

Interferon

The important role played by interferon as a defense mechanism is clearly documented by three types of experimental and clinical observations: (1) for many viral infections, a strong correlation has been established between interferon production and natural recovery; (2) inhibition of interferon production or action enhances the severity of infection; and (3) treatment with interferon protects against infection. In addition, the interferon system is one of the earliest appearing of known host defenses, becoming operative within hours of infection.Figure 49-3 compares the early production of interferon with the level of antibody during experimental infection of humans with influenza virus. Clinical studies of interferon and its inducers have shown protection against certain viruses, including hepatitis B and C viruses, papovaviruses, rhinoviruses, and herpes simplex virus.

Figure 49-3. Production of virus, interferon, and antibody during experimental infection of humans with influenza wild-type virus.

Figure 49-3

Production of virus, interferon, and antibody during experimental infection of humans with influenza wild-type virus. Nonspecific defenses include anatomic barriers, inhibitors, phagocytosis, fever, inflammation, and IFN. Specific defenses include antibody (more...)

Although interferon was first recognized as an extraordinarily potent antiviral agent, it was found subsequently to affect other vital cell and body functions. For example, it may enhance killing by granulocytes, macrophages, natural killer (NK) cells, and cytotoxic lymphocytes and affect the humoral immune response and the expression of cell membrane antigens and receptors. It may also lyse or inhibit the division of certain cells, influence cell differentiation, and cross-activate hormone functions such as those of epinephrine and adrenocorticotropin (ACTH). The effect of these modulations may influence many viral infections.

Interferon Production and Types

Interferon is produced de novo by cellular protein synthesis. The three types (alpha, beta, and gamma) differ both structurally and antigenically and have molecular weights ranging from 16,000 to 45,000. Interferons are secreted by the cell into the extracellular fluids (Fig. 49-4). Usually, virus-induced interferon is produced at about the same time as the viral progeny are released by the infected cell, thus protecting neighboring cells from the spreading virus.

Figure 49-4. Induction of beta interferon, alpha interferon, and gamma interferon, respectively, by foreign nucleic acids, foreign cells, and foreign antigens.

Figure 49-4

Induction of beta interferon, alpha interferon, and gamma interferon, respectively, by foreign nucleic acids, foreign cells, and foreign antigens.

The three known types of interferon are induced by different stimuli. Beta interferon is induced by viral and other foreign nucleic acids in most body cells (fibroblasts, epithelial cells, and macrophages). This induction mechanism is illustrated inFigure 49-4 and the top portion ofFigure 49-5.

Figure 49-5. Cellular events of the induction, production, and action of interferon.

Figure 49-5

Cellular events of the induction, production, and action of interferon. Inducers of interferon react with cells to depress the interferon gene(s) (A). This leads to the production of mRNA for interferon (B). The mRNA is translated into the interferon (more...)

Alpha interferon can be induced by foreign cells, virus-infected cells, tumor cells, bacterial cells, and viral envelopes that stimulate mostly circulating dendritic cells and to a lesser degree monocytes and B lymphocytes to produce it (Fig. 49-4, middle).

Gamma interferon is produced (along with other lymphokines) by T lymphocytes induced by foreign antigens to which the T lymphocytes have been presensitized (Fig. 49-4). Mitogens for T cells may mimic this induction. Gamma interferon has several unusual properties: (1 ) it exerts greater immunomodulatory activity, including activation of macrophages, than the other interferons; (2) it exerts greater lytic effects than the other interferons; (3) it potentiates the actions of other interferons; (4) it activates cells by a mechanism significantly different from that of the other interferons; and (5) it inhibits intracellular microorganisms other than viruses (e.g., rickettsia).

The 24 genes that code for interferons alpha and the single gene for beta in humans, are located in adjacent positions on chromosome 9. The only gene for interferon gamma is found on chromosome 12. The genes for interferons alpha and beta exhibit significant homology but not with interferon gamma.

Genes for interferon alpha may be differentiated into two distinct clusters on the basis of the degree of homology. As a consequence, interferon alpha comprises two families of proteins, at least 14 of which belong to the alpha-1 type and two to the alpha-2 type (omega and tau).

Also, interferon occurs without apparent stimulation in the plasma of patients with autoimmune diseases (such as rheumatoid arthritis, disseminated lupus erythematosus and pemphigus) and in patients with advanced HIV infection. In these cases, an interferon antigenically identical to interferon alpha is present but which, unlike the latter, is partially inactivated at pH 2 (acid-labile interferon alpha). This interferon is a synergistic combination of interferons alpha (acid stable) and gamma (acid labile). Consequently, acid treatment reduces the interferon activity by inactivating the synergistic interferon gamma.

Mechanism of Action

Interferon does not inactivate viruses directly. Instead, it prevents viral replication in surrounding cells by reacting with specific receptors on the cell membranes to derepress cellular genes that encode intracellular effector antiviral proteins, which must be synthesized before virus replication can be inhibited (Figs. 49-5 and49-6). Alpha and beta interferons both bind to the same type of membrane receptor; gamma interferon binds to a different receptor. The antiviral proteins probably inhibit viral multiplication by inhibiting the synthesis of essential viral proteins, but alternative or additional inhibitory mechanisms (e.g, inhibition of transcription and viral release) also occur. Viral protein synthesis may be inhibited by several biochemical alterations of cells, which may, in theory, inhibit viral replication at the different steps shown inFigure 49-6.

Figure 49-6. Molecular mechanisms of interferon antiviral actions.

Figure 49-6

Molecular mechanisms of interferon antiviral actions.

It has been shown that the antiviral state may be transferred from interferon-treated cells to adjacent untreated cells without the continued presence of interferon (Fig. 49-4); this transfer mechanism may further amplify and spread the activity of the interferon system.

The interferon system is nonspecific in two ways: (1) various viral stimuli induce the same type of interferon, and (2) the same type of interferon inhibits various viruses. On the other hand, the interferon molecule is mostly specific in its action for the animal species in which it was induced: interferon produced by animals or humans generally stimulates antiviral activity only in cells of the same or closely related families (e.g., human interferon protects human and monkey cells, but not chicken cells).

Interferon During Natural Infection

The importance of interferon in the response to certain natural virus infections varies. Much depends on the effectiveness of the virus in stimulating interferon production and on its susceptibility to the antiviral action of interferon. Interferon protects solid tissues during virus infection; it is also disseminated through the bloodstream during viremia, thereby protecting distant organs against the spreading infection. Cells protected against viral replication may eliminate virus by degrading the virus genome (Fig. 49-7).

Figure 49-7. Nonspecific elimination of viruses by cells.

Figure 49-7

Nonspecific elimination of viruses by cells.

Medical Applications

Interferons have been approved in several nations for treatment of viral infections (papillomas and condylomata, herpes simplex, and hepatitis B and C) and cancers (hairy cell leukemia, chronic myelogenous leukemia, non-Hodgkin's lymphomas, and Kaposi's sarcoma in AIDS patients). Clinical trials also have shown effectiveness against cryoglobulinemia and thrombocytosis and maintenance of remission in multiple myeloma. Interferon beta has received governmental approval for treatment of relapsing multiple sclerosis and interferon gamma for chronic granulomatous disease. Studies of effectiveness in other viral infections and cancers are continuing, as are studies with substances capable of inducing endogenous interferon.

Conclusion

In conclusion, individual defense mechanisms assume roles of varying importance during different viral infections; in most cases, the recovery process is probably carried out by the simultaneous or sequential action of several mechanisms. The presence of multiple defenses helps explain why suppression of one or several mechanisms does not entirely abrogate host resistance to viral infections; however, impairment of host defenses by medications used for symptomatic relief of viral infections may lead to more severe illness. For example, aspirin and corticosteroids reduce the nonspecific defenses. Therefore, the well-established principle of the ancient physician—“primum non nocere” (primarily do not harm)—is still valid.

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