IT 6 : LUNG DEFENSE MECHANISMS AND LUNG IMMUNOLOGY

I. INTRODUCTION II. SPECIALIZEDREGIONAL DEFENSES

Nose and OropharynxConducting AirwaysThe Alveolar SpacesLymphocytes in the Alveolar Space

III. DEFECTS IN HOST DEFENSES THAT CAN BE ASSOCIATED WITH RESPIRATORY INFECTIONS

IV. HOST DEFENSES IN THE APPROACH TO PATIENTS WITH PULMONARY DISEASE

I. INTRODUCTION

The atmosphere that we breathe is more than just “air.” In reality, it is a complex mixture of ambient gases and environmental particulates to which virus- and bacteriacontaining droplets can be added when respiratory secretions are coughed or sneezed out by others.Moreover, normal humans frequently aspirate secretions from the upper respiratory tract, particularly during sleep. The respiratory system must recognize and eliminate these unwanted elements in inspired air to keep pulmonary structures free of infection, yet not overreact inappropriately to every stimulus. This is accomplished by local mechanisms and innate immune defenses spaced along the entire respiratory tract to protect it. The fact that the normal lower respiratory tract is infection free despite its constant exposure to foreign antigens and infectious agents is testimony to the efficiency of these defense mechanisms. The evolving appreciation of direct associations between aging and breakdowns of these host defenses and resultant pulmonary diseases emphasizes the need for all physicians to be familiar with these critical protective processes.

Components of the defense system are spaced along the entire respiratory tract, from the point of air intake at the nose and lips or mouth to the level of oxygen uptake at the alveolar surface. The conducting airways functionally extend from the nares down to the respiratory bronchioles and include the nasal turbinates, epiglottis, larynx, pharyngeal lymphoid tissue (Waldeyer’s ring), and other anatomic barriers. Fourteen generations of dichotomous airway branching of the respiratory tree, as bronchi and bronchioles, in this segment cause the airstream flow to decelerate and deflect the particles it contains onto themucosal surface, trapping them in airway mucus. In this location, inhaled particulates and infectious agents also interact with other locally produced proteins, such as secretory immunoglobulin A (IgA). Resulting ciliary clearance or coughing efficiently removes these particulates from the respiratory tree. Beyond the respiratory

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BLOK 12 : RESPIRATORY SYSTEM

bronchioles, other noncellular host defenses remain important in protecting the alveolar units (Fig. 1). These defenses are in the lining material fluid of the alveoli, which contains

Figure 1 Airway lumen mucosal components. A portion of the conducting airway surface is enlarged (A) and depicts the mucosa and its submucosal structures. The pseudostratified ciliated epithelium has a covering layer of mucus (produced by goblet cells and bronchial glands) and fluid that contains various proteins, including immunoglobulins and secretory component. A few surface cells may be present, such as lymphocytes (from bronchial-associated lymphoid aggregates) and macrophages. Among the epithelial cells are absorptive microvillous brush cells and the dendritic cells, concentrated near lymphoid aggregates or in the respiratory bronchiole area, whose cellular processes interdigitate with the mucosal surface. In addition, the epithelial cells can produce proinflammatory cytokines that influence mucosal swelling and permeability. In the submucosa below the basement membrane, plasma cells and mast cells reside that secrete local immunoglobulins (such as IgA) and mediators (such as histamine). Interacting with all of these glandular and cellular networks are nerves, exerting their control through neuropeptides, and by adrenergic and cholinergic nerve fibers. A rich bronchial arterial vascular supply also exists. (Modified from Reynolds HY: Pulmonary host defenses-state of the art. Chest 95(Suppl):223–230, 1989, with permission.)

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surfactant apoproteins and glycoproteins such as fibronectin, immunoglobulins such as IgG opsonins, and complement (properdin factor B), which are active against aerosolized, inhaled particles or microorganisms.

Alveolar macrophages are the principal phagocytic and scavenger cells on alveolar surfaces. Particulates and microbes that evade other host defense mechanisms and arrive on the alveolar surface are efficiently removed by these roaming cells. When further assistance is required, an inflammatory reaction can be initiated,which attracts olymorphonuclear neutrophils (PMNs) and other vasomediators and humoral immune elements from systemic sources.

At all levels of the respiratory tract, specific and nonspecific defense mechanisms exist to protect respiratory structures. The nonspecific mechanisms, as noted above, include the mechanical barriers, cough, mucociliary elevator, and macrophage phagocytosis, which behave similarly regardless of the inhaled particulate. They also include aspects of the innate immune response, such as the inflammation triggered by Toll-like receptors (TLR). In contrast, prior contact with a microbial agent or a sensitizing substance can induce antigen-specific cellular or humoral immune responses, activating adaptive or acquired immunity. The latter includes the production of secretory IgA antibody in the airway, which preventsmucosal adherence, and IgG opsonins that facilitate phagocytosis. Such responses help the lung deal more efficientlywith these agents and substances on rechallenge in the future.

Insummary, the integratedaction of diversepulmonary defensemechanisms along the respiratory tract acts to remove or neutralize microorganisms, particulates, and noxious gases that are inhaled or aspirated into respiratory structures.Many are mechanical barriers and reflex actions that are concentrated in the naso-oropharynx and along the conducting airways. There is also phagocytosis, which occurs in the alveoli and airways. These are surveillance mechanisms that function mechanically and can be activated by nonspecific (nonimmunologic) or immunogenic stimuli. In addition, several augmenting mechanisms exist that enhance the responsiveness of this defense system and make it flexible and adaptable. Crucial in this regard are the pathways of innate immunity and the ability of dendritic-type macrophages in the lung to mount antigen-specific immune responses (humoral and cellular)—adaptive immunity—and a local inflammatory reaction. This allows components in plasma and blood cells to bolster local defenses in the airways and alveoli. A more indepth review of these innate defense mechanisms follows. Also in the “Suggested Reading” section, several reviews by the authors and others providemore details about this actively expanding topic.

II. SPECIALIZED REGIONAL DEFENSES

Nose and Oropharynx

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Inhaled air passes through the nose or mouth back to the glottis and into the extrathoracic portion of the trachea before it enters the thorax. With nasal breathing, air is filtered and conditioned for humidity and body temperature as it flows over the nasal turbinates and mucosa of the posterior pharynx. With nasal obstruction or ventilatory requirements for exertion that exceed about 20 to 30 L/min, mouth breathing occurs. Inhaled air then may pass into the trachea without optimal filtering and climatic conditioning. The nose provides formidable barriers to inhaled particulates. The nasal hairs help to exclude large particles, and materials greater than 10 μm in diameter that bypass the hairs impact upon the nasal mucosa. Sneezing (or blowing) then has the effect of coughing and provides high-velocity ejection from themucosal surface. For substances that attach to the nasal mucosa, production of large quantities of watery secretions helps to wash off the surface (rhinorrhea).Mucociliary clearance is also operant in the nasal cavity. “Downspouts” leading from the ears, lacrimal glands, and sinus cavities provide numerous points for the addition of fluid to the nasal secretions.However, these drainage systems also contain vulnerable points that are prone to blockage. The complex plumbing found in the nose works well if there is good gravitational flow and orifices stay open. If not, sinusitis, otitis media, parotid gland obstruction, and occluded tear ducts result. In some diseases, dryness of secretions (sicca syndrome) is problematic.

Several substances in nasal secretions help control bacteria or viruses. Prominent in this regard are lysozyme and immunoglobulins, especially secretory IgA (SIgA) which bathesmucosal surfaces. The nose and upper airways are contiguous immunologically with the lower airways and have been studied extensively. Nasal secretions, like those from other external or mucosal surfaces, are rich in IgA, which is synthesized locally by submucosal plasma cells. Free secretory component (SC) can also be detected in nasal wash fluid. Of the nasal immunoglobulins, SIgA is the major source of antibody, accounting for approximately 10 percent of the total protein content of nasal washings. IgG is present in smaller amounts. IgE probably is not secreted by normal, nonatopic people. Only in people with allergic rhinitis will IgE antibody be substantial. The usual specificity of IgA antibody is antiviral. After nasalimmunization of normal subjectswith various viral or mycoplasmal vaccines, many experimental studies have shown that appropriate neutralizing IgA antibody can be elicited. Although these antibodies are protective against homologous and live microbial challenge, the duration of protection is often brief, and the antibody titers diminish rapidly unless repeated exposure occurs.

In the oral cavity, the tongue sweeps against many surfaces during chewing and swallowing. This should make it difficult for bacteria to persist in these locations. However, bacteria adhere to buccal squamous cells, and many accumulate in crevices around teeth and gums and colonize dental plaque.Many kinds of bacteria arepresent: aerobes and anaerobes, spirochetes, gram-positive and gram-negative species, and some that specialize in making dental plaque and causing tooth decay. A common

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feature of host defense in the mouth and nose is the plentiful amount of SIgA in secretions that bathe each area. The parotid glands and probably the submandibular salivary glands secrete IgA as their principal humoral immune substance; this immunoglobulin accounts for 12 to 15 percent of the total protein in their secretions. In this fluid, albumin represents about 10 percent of the protein, but IgG is barely detectable (under 1 percent). In parotid fluid, IgA is found in monomeric and dimeric forms, and free secretory component can be detected as well. Thus, normal nasal and parotid (or salivary) secretions have about the same composition of immunoglobulins. As with the nasal immune system, it has been possible to manipulate SIgA in the mouth to produce antibodies against certain cariogenic strains of streptococci that will subsequently prevent bacterial adherence to teeth—the immune exclusionfunction of SIgA antibody. The importance of a vaccine approach for augmenting dental defenses has yet to be fully determined.

Host defenses in the nose and mouth serve as a reminder that the upper portion of the respiratory tract has features in common with the lower part, particularly at the mucosal surfaces. They also demonstrate that infections in the nose, sinuses, ears, teeth, and gums may have ramifications for the diagnosis or successful treatment of illness in the lower respiratory tract. As examples, aspiration of anaerobic bacteria in oral secretions or dental plaque contributes to lung abscess formation; chronic sinusitis can be presentwith cystic fibrosis, dyskinetic ciliary syndromes, and dysgammaglobulinemia; atopic diseases can manifest with rhinitis, sinusitis, and asthma; and control of asthma symptoms often requires vigorous treatment of concomitant sinus infection.

Conducting Airways

Bridging the upper airway (nose, oropharynx, and larynx) and the alveolar air-exchange area distal to the terminal bronchioles are the conducting airways (Fig.1).Mucociliary clearance and coughing are the principal means of cleansing themucosal surfaces of these airways. SIgA antibodies also prevent epithelial attachment of certain bacteria and viruses to the ciliated and nonciliated airway epithelial cells. The branching structureof the airways also causes airborne particulates to impact against themucosa, enhancing the efficiency of mucociliary clearance. Bronchial-associated lymphoid aggregates are present, especially around branching points. This segment is susceptible to many diseases—e.g., epithelial cell infection with viruses or bacteria such as Bordetella pertussis Chlamydia pneumoniae, orMycoplasma pneumoniae; inflammation, edema, and bronchoconstriction in asthmatic syndromes; chronic infection in bronchiectasis; irritation from noxious gases; and lung cancer. The conducting airways mucosa is coated to a depth of 5 to 100 μm with a mucous gel-aqueous sol complex viscous fluid, which has a low pH (6.6 to 6.9). This is secreted by bronchial glands, goblet cells, and Clara cells (nonciliated bronchiolar secretory cells found in the terminal bronchioles). Airway surface liquid is also derived from transepithelial acid-

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base flux across the bronchial epithelium. Special proteins, such as SIgA and SC, can be added locally along airways by immunoglobulin-secreting plasma cells and epithelial cells. Antimicrobial factors such as lysozyme, lactoferrin, cathelicidin, and defensins are present.

About half of the mucosal epithelial cells have beating cilia that propel secretions up the respiratory tree. Periodic coughing can assist the process. An intactmucosal lining and overlying mucous layer, containing mucin glycoproteins and proteoglycans, provide a protective barrier or blanket that prevents inhaled particulates from penetrating or sticking to the respiratory surface. This seems to be an important component of host defense. Bacteria and other infectious agents may transiently colonize the airways, but mucociliary clearance effectively removes them. Tight junctions between epithelial cells also limit the passage of macromolecules into the submucosa, and microvillous brush cells may help clear fluid. A number of circumstances can alter these protective barriers, making this portion of the respiratory tract susceptible to disease. They include (1) malnutrition, which affects the integrity of mucosal epithelial cells and enhances bacterial adherence; (2) cigarette smoke and noxious fumes, which disrupt the anatomy of epithelial junctions and enhance the passage of airway substances into areas that are usually inaccessible; and (3) some bacteria, which elaborate proteolytic enzymes that may break down IgA, promoting selective colonization and persistence in matrix-enclosed biofilms that help avoid innate immunity and create chronic infections.

Lymphoid tissue is present along the entire respiratory tract, but the level of organization of the lymphoid tissue varies greatly. A ring of lymphoid structures are situated in the naso-oropharynx. Lymphoid nodules may occur in the mucosal surface of large and medium-sized bronchi and are particularly numerous at points of airway branching. On the airway side, these submucosal follicles are covered by a layer of flattened, nonciliated surface epithelium, which is often observed to be infiltratedwith lymphocytes. These bronchialassociated lymphoid tissues (BALT) bear some resemblance to gut-associated lymphoid tissues (Peyer’s patches), and are part of the body’s overall mucosal-associated lymphoid network (known as MALT) that is important inmucosal immunity. Whereas BALT is easily demonstrated in some rodents and rabbits, subhuman primates and humans have decidedly less obvious amounts of this lymphoid tissue, and it may not be as relevant to airway defenses as initially thought, especially in adults.

Loosely organized collections of lymphocytes (lymphoid aggregates) are concentrated in the distal airways, especially at the bronchoalveolar junctions at the interface between the ciliated epithelial cells of the terminal bronchioles and the alveolar lining cells. These aggregates provide an opportunity for close interaction between lymphoid cells and inhaled antigens that have been deposited in the lower respiratory tract. Antigens and microbes may adhere to surface macrophages or dendritic cells imbedded in the mucosa where immune processing or elimination occurs. Bacteria such as Pseudomonas aeruginosa may become enmeshed in a biofilm containing their exopolysaccharides, which can interfere with macrophage or dendritic

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cell elimination of them and contribute to airway colonization and persistence. Also, in the vicinity of the respiratory bronchioles, lymphatic channels begin that might provide these lymphocytes with a route to draining lymph nodes (hilar nodes) where immunologic responses develop.

Respiratory Bronchioles

Anatomically, lung structure changes at the level of the respiratory bronchioles, which are inserted between the distal conducting airways and the acinar units (alveolar ducts and alveoli) of the air exchange surface. They functionally separate the upper and lower respiratory tracts. This segment can be a bottleneck or choke point for airflow, but it is the last surface to capture small airborne particulates and microbial or antigenic debris before entering the alveolar space; adaptive immune responses can begin here. Several structural changes occur: the single-layer cuboidal epithelial surface flattens and differentiates into alveolar type I cells that primarily cover the alveolar lining surface; mucus-secreting cells disappear, although goblet cells can be found in cigarette smokers; and another secretory cell type becomes prominent, the Clara cells. Pulmonary brush cells with a tuft of squat microvilli are found in this area, especially in rodent species, and may be involved with chemosensing or trapping inhaled particles and pollutants, or with regulating fluid and solute absorption. Dendritic macrophage-like cells, which may constitute 1 percent of the cells in the surface of this segment, are present to capture and process antigens. Lymphatic channels form to collect the lymphatic fluid emerging from the interalveolar interstitial spaces. The changeover from the bronchial arterial blood supplying the conducting airways to the pulmonary artery-capillary blood flow structure that surrounds the alveoli also occurs, which is necessary for aeration.

The Alveolar Spaces

Defenses in the airways (Fig.2) eliminate most particles and microbes inspired into the lungs. As a result, the airways distal to the major bronchi are probably sterile in normal subjects. However, some particles of small size and special geometry can elude the airwaymucosal mechanisms and reach the air-exchange surface of the alveoli.When this occurs, another set of host defense mechanisms must take over. Microbial clearance and the removal of other antigenic material from alveoli depend on cellular and humoral factors such as the lipoproteins, immunoglobulins, and complement factors in the alveolar lining fluid and phagocytic cells such as alveolar macrophages and PMNs.

Inhaled microbes are an appropriate example. If a bacterium of critical size (0.5 to 3 μm in diameter) is deposited in an alveolus, it is likely to make contact with the alveolar wall and roll along in about 0.2 μm of alveolar lining fluid, pH 6.9, which is a ombination of a watery subphase with an overlying film of surfactant secreted by type II pneumocytes. In the process, a microbe encounters several substances that can inactivate it and assist in its eventual

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phagocytosis. These substances include a variety of soluble lipoprotein substances, IgG, complement factor (C3b), and nonimmune opsonins, such as high-molecular-weight fibronectin fragments. The lipoproteins in the form of surfactant are secreted by type II pneumocytes, and surfactant proteins A and D have opsonic effects through binding of surface carbohydrates, which promotes antibacterial activity against staphylococci and rough colony strains of some gram-negative rod bacteria. The immunoglobulins are principally of the IgG class. They account for 5 percent of the total protein in alveolar fluid, with subclasses IgG1 and IgG3 being the most important and lesser concentrations of monomeric and secretory forms of IgA being noted. These immunoglobulins can develop specific opsonic antibody activity for the bacterium. The complement components, especially properdin factor B, interact with the bacterium and can trigger the alternative complement pathway, thereby lysing the microbe directly. One or all of these interactions can prepare the bacterium for ingestion by an alveolar macrophage. Although alveolar macrophages avidly phagocytose some inert particles, they ingest viable bacteria with considerably less enthusiasm. Coating or opsonizing the organisms will enhance phagocytosis appreciably as studied in an in vitro culture system. The nonimmune opsonins nonspecifically enhance this process. The immunoglobulins are capable of enhancing alveolar macrophage phagocytosis in an antigen-specific fashion, and the C3b complement fragment can function in concert with IgG to enhance or amplify this process.

Phagocytosis, the ingestion of particulate matter by cells, is divided into two phases: receptor attachment of the particle to the cell surface and internalization. Attachment of the particle to the surface of the phagocytic cell is essential before ingestion occurs. Although binding occurs randomly, it is greatly enhanced by opsonization of the particle by antibody (especially IgG) or a component of the complement system, C3b. Opsonin-dependent phagocytosis is mediated by receptors on the cell surface for the Fc component of the opsonizing immunoglobulin or complement. Specific receptors for the Fc portion of IgG(Fcγ) (IgG3 and IgG1 primarily) and for the third component of complement (C3b) are present on human monocytes and alveolar macrophages. Receptors for IgAare also found on alveolar macrophages.There is evidence that the number and function of these receptors can be modulated by lymphocyte-derived cytokines such as interferon-γ (IFN-γ). Ingestion of membrane-bound particles occurs via a process that is energy-dependent as the plasma membrane of the ingesting cell surrounds the bound particle, enclosing it in an endocytic vesicle. This is followed by the activation of a number of well-developed mechanisms that operate to kill internalized pathogens.

Following internalization of bacteria, the fate of alveolar macrophages is not certain. They are long-lived tissue cells that can survive at least for several months and presumably are capable of handling repeated bacterial and other microbial challenges (reusable phagocytes). Because they are mobile cells, they can migrate quickly to other alveoli through the pores of Kohn, or move to more proximal areas of the respiratory tract (to the region of the respiratory bronchioles) for elimination from the lungs by the mucociliary

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escalator. In addition, macrophages may gain entry into lung lymphatics at the same location and be carried to regional lymph nodes. This exit gives them access to systemic lymphoid tissue and is important in initiating cellular immune responses. Undoubtedly, macrophages are also instrumental in degrading antigenic material and presenting it in an appropriate manner to local T lymphocytes as part of innate and adaptive immunity in the lung.

Increasingly, attention is being given to the immune effector role of macrophages. The alveolar macrophage has a dual role in the respiratory tract—one as a phagocyte to dispose of debris, process foreign antigens, and kill ingested microorganisms and a second as an effector cell to initiate immuneandinflammatory responses. Alveolarmacrophages are usually successful in inactivating inhaled microorganisms. As a result, clinical disease and pneumonitis rarely develop after day-to-day exposures.However, if a sufficiently large bacterial inoculum reaches the lower respiratory tract, or if particularly virulent microorganisms are inhaled, the macrophage system can be overwhelmed. By the secretion of proinflammatory chemotactic factors such as the chemokine family cytokines, alveolar macrophages then recruit PMNs and other cells to the lung, and pneumonitis develops. Also, airway epithelial cells can generate proinflammatory cytokines to assist with PMN attraction.

Gram-negative rod bacteria provide an interesting example. Some complement components, particularly factor B, are present in small amounts in bronchoalveolar fluids. The bacterial endotoxin in the gram-negative rod bacteria can directly activate the alternative complement pathway—leading to the formation of C5a, which is a potent stimulus for PMN chemotaxis. Also, the inflammatory responsemay activate the kinin system; this results in generation of kallikrein, which has chemotactic activity, and bradykinin, which is capable of increasing vascular permeability. The latter allows for the seepage of fluid and other humoral and bioactive substances from the intravascular compartment into the alveoli. Another mechanism of inflammation emanates from the alveolar macrophage itself. Following phagocytosis of opsonized bacteria or other forms of activation, proinflammatory chemokines are synthesized and secretedbymacrophages that will attract PMNs and other cells. Several substances with chemotactic activity have been found to be produced by human alveolar macrophages. These include interleukin-8 (IL-8), macrophage inflammatory protein-2 (MIP), monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor (TNF), and lipoxygenase pathway metabolites of arachidonic acid, namely leukotrienes. Leukotriene B4 (LTB4) is one of the most important of these.

Inflammation is the ultimate host response to contain common bacteria that reach the alveolar space. This response can be activated in several ways: (1) directly by microbes or substances such as lipopolysaccharide (endotoxin) that can activate the complement cascade, probably via the alternate complement pathway; (2) through the generation of phlogistic factors from the kallikrein and bradykinin pathways; and (3) from the effector cell function of macrophages. It is also known that other airway cells, such as epithelial

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cells, elsewhere in the respiratory tract, can produce chemokines like IL-8 and that this can stimulate inflammation in other sites (bronchitis).

Special interest has focused on the macrophagesecreted proinflammatory chemokines, a family of cytokines that can stimulate cellular motion (chemokinesis) and promote directed migration of different populations of responder cells (chemotaxis). These populations are primarily PMNs in the acute inflammatory responses. Lymphocytes, monocytes, and eosinophils are also recruited in the chronic phase of pneumonia, chronic inflammatory disorders such as hypersensitivity pneumonitis and sarcoidosis, and atopic and eosinophilic syndromes. Investigation has elucidated the cellular mechanisms whereby CXC chemokines activate and initiate the migratory process of PMNs. An extensive review of the literature is summarized to say that this process involves a number of cell surface adhesion molecules, found on endothelial cells (adhesion molecule ICAM-1, L- andP-selectins and integrins) and granulocytes, that bind to one another. At sites of inflammation mediators such as IL-1, TNF, and IFN-γ induce or augment the expression of these adhesion molecules. As a result, intravascular PMNs slow down, roll along, deform, and then anchor on the endothelium. They then enter the interstitium via traversing capillary endothelial cells, which contract or pull apart to allow a gap through which PMNs pass, and plasma fluid can leak, and the cells emerge through the alveolar type I pneumocyte lining barrier into the alveoli.Microvillous brush cells may also absorb fluid or regulate ion-solute flux.

Eventually, all pneumonic responses run their course. If the host is successful in containing the infective microbes or particles that initially incited the host response, resolution usually occurs. Resolution can be passive, resulting from the removal of the initiating agent. Resolution can have an active phase as well. In the active phase, signals must go out to begin the healing and resorption phases that will restore the lung to normal respiratory function and architecture. Less is known about active resolution of inflame mation. A plateletderivedsubstance, sphingosine 1-phosphate (S 1-p) may help restore the endothelial barrier by reducing PMN infiltration and vascular leak, as found with endotoxin injury.Moreover, cytokines such as transforming growth factor-β, IL-6, IL-10, and the IL-1 receptor antagonist released by macrophages and possibly other cells are believed to be important mediators of this process. As such, they provide a view of potential anti-inflammatory therapies of the future.

Lymphocytes in the Alveolar Space

When cells are retrieved from the alveolar surface by bronchoalveolar lavage (BAL), approximately 7 to 10 percent of the respiratory cells are lymphocytes. Some characteristics of these cells are given in Table 19-1. Two major populations of lymphoid cells are recognized, those that depend on the thymus gland for differentiation (T cells) and those that differentiate independently of the thymus in the bone marrow (B cells).

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TheTandBlymphocytes are indistinguishable by usual morphologic criteria but can be differentiated by membrane surface markers. They are also functionally distinct, with T cells playing an important role in cell-mediated immunity and cell-mediated cytotoxicity while the B cells serve as precursors for cells that synthesizeimmunoglobulins and, hence, antibody molecules that are the basis of the humoral immune response.

As shown in Table 19-1, approximately 70 percent of the lymphocytes in lavage fluid are T cells and approximately 5 percent are B cells. The ratio of T to B cells in lavage fluid is roughly that of peripheral blood, although in blood more circulating B cells are usually identified (approximately15 percent). Approximately 1 to 5 percent of lung lymphocytes seem to be able to release or secrete class-specific immunoglobulin. Enumeration of these cells has found that IgG- and IgA-secreting cells are much more numerous than IgM-producing cells. Natural killer lymphocytes make up about 5 to 8 percent of lung lymphocytes. As they do not express T-cell receptors or surface immunoglobulins for specific antigens, they can respond in an antigen-independent way to help contain viral infections and are thus important in innate immunity.With phenotypic markers, T cells can be divided into two principal groups. The CD8 cells usually have a suppressor-cytotoxic phenotype. TheCD4cells usually have a helper-inducer phenotype and thus are also called T-helper (TH) cells. In the BAL fluid from normal subjects there is a greater percentage of CD4 cells, with approximately 45 percent of the total T cells expressing this surface marker. In contrast, approximately 25 percent of lungTcells express the CD8 phenotype. In lung lavage fluid, the ratio of these subtypes of T cells is approximately 1.5 to 2:1, which is approximately the same ratio found among peripheral blood lymphocytes. As noted, most of the T lymphocytes in the alveoli are CD4-positive. When activated, these TH cells are capable of producing regulatory cytokines that in turn modulate the function of other immune and structural cells. Recent studies suggest that there are at least two subsets of CD4 TH cells, T-helper–1 (TH1) and T-helper–2 (TH2) cells.

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These cell populations have different functions based on the different array of cytokines that they produce. TheTH1cells secrete IFN-γand IL-2, which activate macrophages and play a major role in cell-mediated immunity. The TH2 cells produce IL-4, IL-5, and IL-6, which stimulateBlymphocytes to produce immunoglobulins and, by their production of IL-10 and IL-13, suppress monocyte/macrophage activity and cell-mediated immune responses. Thus, TH2 cells play a particularly important role in generating tissue eosinophilia and stimulating IgE production, processes that are extremely important in atopy, allergic asthma, and other inflammatory pulmonary disorders. IL-2, formerly called T-cell growth factor (TCGF), is among the most important T-cell–regulating cytokines. It is produced by activated T cells and acts in an autocrine or paracrine fashion to stimulate TH1 cells and TH2-cell precursors.

IL-2 can also activate killer T cells. A few killer lymphocytes can be identified among alveolar T cells, but these cells seem to be dormant in normal subjects until stimulated. Lastly, IL-2 can stimulate B lymphocytes to differentiate into plasma cells that synthesize various classes of immunoglobulins. This is a mechanism by which local production of immunoglobulin in the lung can occur. In all cases, the effects of IL-2 are mediated by the multimeric IL-2 receptor, a component of which is the Tac-surface ligand. The expression of the IL-2 receptor is highly regulatable, and the expression of the Tac antigen can be used as a marker of T-cell activation.Most T cells have T-cell receptors with alpha and beta subunits (α/β T-cell receptors). In the normal lung, a lesser number of T cells have gamma and delta T-cell receptors.

The function of these cells is poorly understood. They may, however, play an important role in mucosal immunity, since they are increased in atopic allergic subsets. Alveolar macrophages and lymphocytes have the capacity to produce many cellular mediators (cytokines) that in turn affect each other as well as other inflammatory, structural, and immune effector cells. This dynamic and complex interaction is illustrated in Fig. 19-2, which reviews dendritic cell, alveolar macrophage, and lymphocyte interactions in the airways and alveolar milieu. Monocyte precursors from the blood differentiate into mature macrophages under the influence of vitamin D metabolites and undoubtedly other stimuli and become long-lived, aerobically metabolizing alveolar phagocytes. Their principal activity is to cleanse the alveolar surface and ingest debris that accumulates or microbes aerosolized into the lungs. In the process, the macrophages may become “activated” and are then capable of secreting an enormousarrayof enzymesandcytokines. These moieties can affect the function of resident cells of the lung such as lymphocytes or epithelial cells. In addition, the release of proinflammatory chemokines attractsPMNs,lymphocytes,monocytes, and other cells into the alveoli. Of particular note are LTB4, IL-8, TNF-α, MIP-1α, MCP-1, and IL-1. When secreted by activated macrophages (especially in active lung forms of sarcoidosis) IL-1 may attract T lymphocytes to the lungs. In the other direction, activated TH cells can produce several monokines that affect macrophage function. Such a substance is migration inhibition factor, which immobilizes macrophages

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engaged in phagocytosis. Of special interest is IFN-γ, which activates macrophages, increasing their expression of membrane receptors, which in turn enhances macrophage phagocytic uptake. IFN-γ also has other functions that promote cellular immunity.

Figure 2 Host defenses in the alveolar space. Bacteria (B) that escape clearance mechanisms in the upper respiratory tract (URT) can reach the alveolus (represented by an enlargement of one). Most of the alveolar surface is lined by type I epithelial cells with pulmonary microvillous brush cells interspersed, and type II cells positioned in the corners that secrete surfactant. A variable amount of interstitial space separates the epithelium from the capillary endothelium where sequestered PMNs and platelets reside. A bacterium deposited in an alveolus may encounter at least three different but coordinated sets of innateimmunity immunologic materials and cells that can destroy it: opsonins, both IgG and surfactant proteins A and D, or complement factors that facilitate phagocytosis or create a lysis of the microbe; activated macrophages stimulated by cytokines produced by nearby lymphocytes; and other inflammatory phagocytic cells, usually PMNs attracted into the alveolar space by proinflammatory chemokines produced locally by macrophages and epithelial cells. (Modified from Reynolds HY: Respiratory infections may reflect deficiencies in host defense mechanisms. Dis Mon 31:1–98, 1985, with permission.)

Almost mutually exclusive sets of chemokines can be induced by TH1 immune responses (IL-12 and IFN-γ) and by TH2 cells (IL-4 and IL-13) toward infectious challenges. The scheme shown in Fig. 3 may help to explain certain derangements found in a number of lung diseases that have excessive or deficient secretion of cytokines and feature changes in the relative proportions of macrophages and lymphocytes. Examples of

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Figure 3 Major immunity pathways. Within the body, mucosal surfaces are positioned at initial intake and contact points to ‘ ‘meet” external substances that enter with inhaled air, ingested food and liquids, or reproductive secretions.Mucosae in the nose, airways, and gastrointestinal and genital tracts must discriminate between pathogens and harmless microbes or possible toxins and essential nutrients, and then respond quickly to exclude, tolerate, or initiate immune responses.

Respiratory host defenses balance two important immune mechanisms created for dealing with airway microbes or other entering antigens: (1) an innate or quick reaction response producing inflammation as an end point (bronchitis or pneumonitis), and (2) a more deliberate approach through stimulation of lymphocytic pathways that creates a versatile and adaptive response involving specific T-cell activity and/or production of immunoglobulins (antibodies). Foreign substances or microbes or their exoproducts (lipopolysaccharide from gram-negative rod bacteria) that enter the airway lumen and adhere to the mucosa will be picked up by macrophages (M) or dendritic cells (DCs). Toll receptor recognition and attachment are important, and dealt with in a variety of ways. Phagocytic uptake and intracellular killing of bacteria might suffice, or recruitment of PMNs may be needed through secretion of proinflammatory cytokines by macrophages, creating pneumonitis for example. Later, active resolution of inflammation requires inhibition of PMN influx (suppress chemotaxis) and

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cellular cleanup (apoptosis). Alternatively, DCs (or macrophages) can process antigens, present these to major histocompatibility complex (MHC) compatible but na¨ıve CD4+ cells, facilitated with the stimulatory cytokine IL-12; IL-2 produced by CD4+ T cells can direct TH1 lymphocytes to develop and proliferate. In turn, TH1 cells can produce IL-1 and IFN- γ that can stimulate macrophages for the inflammatory pathway, or induce clonal expansion of CD4+lymphocytes that contribute to building granulomata for containment of certain microbes or particles. Returning to the DC-antigen–presenting cell process involving the CD4 cells, another subset of DCs (or macrophages) can produce IL-10, an inhibitory cytokine that promotes the TH1 response preferentially in normal subjects and suppresses the TH2 cellular pathway. However, pending the allergic status of the host (atopy) and/or the particular antigen present, TH2 lymphocytes can be stimulated and in turn produce IL-4, 5, 6, and 13 cytokines that culminate in allergy (asthma and allergic rhinitis) with stimulation of mast cells and then eosinophils and production of reaginic antibodies (IgE, IgG 4). The TH2 immune response is also effective against certain parasites. (From Reynolds HY: Modulating airway defenses against microbes. Curr Opin Pulm Med 8:154–165, 2002.) such cellular imbalances include sarcoidosis, hypersensitivity pneumonitis, and acquired immunodeficiency syndrome (AIDS).

III. DEFECTS IN HOST DEFENSES THAT CAN BE ASSOCIATED WITH RESPIRATORY INFECTIONS

Infection can occur everywhere along the respiratory tract-upper airways (nose, sinuses, ears, and oropharynx), conducting airways (trachea and bronchi down to the respiratory bronchioles), or the alveolar area. Although exposure to a virulent microorganism or to a large inoculum, if inhaled or aspirated into the lungs, may cause illness in a normal person, recurrent or chronic infections may point to deficiency or malfunction of a particular component of the host defense system (Table 19-2). A number of situations associated with frequent respiratory infections serve as examples. Endotracheal tubes give direct access to the lung but, in so doing, bypass the larynx and the other upper-airway protective structures. Patients with depressed consciousness or with postoperative chest or abdominal pain become infected because of their inability to cough and clear airway secretions. In addition, patients with viral infections have an increased incidence of bacterial superinfection. The cause of this association appears to be multifactorial, including the ability of these infectious agents to damage ciliated epithelial cells and diminish the clearance of airway secretions; also viruses and other microbes can infect alveolar macrophages, diminishing their bactericidal activity. In combination, these host defense defects are believed to contribute to the frequent association of influenza infection and staphylococcal superinfection.Ultrastructural defects in the cilia located on the apical edge of the airway epithelial lining cells cause mucociliary dysfunction. As a result, the removal of mucus and respiratory secretions is depressed, and recurrent infections

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and bronchiectasis occur. The constellation ofmultiple upper and lower respiratory infections and bronchiectasis should raise the possibility of a ciliary dyskinesis syndrome. Infertility, especially in males,may be associated, and the evaluation of this problem may bring the respiratory symptoms to the physician’s attention.With age, ciliary beat frequency decreases and might be a factor in greater susceptibility to lung infections in the elderly.A variety of γ-globulin abnormalities are associated with recurrent infection. In patients with hypogammaglobulinemia, the lack of opsonic antibody can promote infections with encapsulated bacteria. Several common bacteria that colonize the airways of patients with chronic bronchitis and chronic obstructive pulmonary disease (Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria) can also produce a specific IgA protease that cleaves the IgA heavy chain in its hinge region adjacent to the Fc portion. By this mechanism, these bacteria could inactivate a substantial portion

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of the secretory IgA coating the conducting airways and gain better access to the ciliated epithelial cells for attachment. While this mechanism is somewhat theoretical, associations between deficiencies in IgG and recurrent infection are well documented. Particularly important are the associations between deficiencies of IgG subclasses IgG2 and IgG4, alone and in combination with IgA deficiencies and chronic inflammation and bronchiectasis. Presumably, an absence of these subclasses denies phagocytic cells potential opsonic antibody, thereby diminishing membrane receptor attachment of opsonized particles or bacteria and subsequent phagocyte ingestion. Clinically, establishing the diagnosis of an IgG deficiency is quite important because, in contrast to many other immunodeficiencies, replacement preparations of IgGare often available for these patients. Cytotoxic antineoplastic chemotherapy and other forms of immunosuppression also compromise host

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defenses in a major way. A major side effect of these therapies is granulocytopenia, which prevents the mobilization of PMNs and creates a poor inflammatory reaction.

IV. HOST DEFENSES IN THE APPROACH TO PATIENTS WITH PULMONARY DISEASE

As noted above, normal hosts can develop respiratory infections or inflammation as a result of exposure to particularly virulent agents or a large inoculum of aerosolized particulates. In others, respiratory infections are associated with obvious clinical features that compromise pulmonary defenses (Table 19-2). Occasionally, however, the physician is confronted with a relatively young person who has an unexpected number of respiratory problems that seem inappropriate. The illness can manifest as recurrent infection or poorly controlled allergic rhinitis, asthma, frequent sinusitis, recurrent nasal polyps, and/or bouts of otitis media. Because the severity of these respiratory problems may not seem great, the physician may not initially suspect that something unusual is present. The propensity for infection may not have been obvious in childhood but became apparent as the patient reached adolescence or adulthood. Although genetic defects usually are manifested in infancy, minor forms of host deficiency, creating antibody deficiency diseases, may not be recognized until later in life. Cystic fibrosis (adult onset), selective absence of IgG subclassimmunoglobulins, structural ciliary defects, and IgAdeficiency are the principal diseases that should be considered in this differential diagnosis. Recurrent sinopulmonary infections are an important clue to all these syndromes.

The physician should be prepared to examine such a patient thoroughly. A detailed history will immediately provide important information about affected siblings, infertility, or a striking change in respiratory health that makes an acquired abnormality likely. Preliminary screening tests are a complete blood count and quantitative serum immunoglobulins, and perhaps pulmonary function tests, even if the chest radiograph is normal in appearance; also indicated may be microbial cultures of respiratory secretions and analysis of the electrolytes contained in a sample of sweat or nasal potential difference measurements. Mucoid strains of Pseudomonas aeruginosa and elevated sweat chloride values can be noted in cystic fibrosis. Other useful secondary-level screening tests are quantitation of subclasses of IgG; secretory IgA as sampled in parotid fluid or nasal wash samples; subtyping of blood lymphocytes; measurement of antibody responses to protein and/or polysaccharide antigens; search for genetic mutations of the cystic fibrosis transmembrane conductance regulator (CFTR); assessment of ciliary clearance with an aerosolized, isotopic tracer; nasalmucosal biopsy for electron-microscopic ultrastructural analysis of cilia; sperm motility in males of appropriate age; and documentation of bronchiectasis by high-resolution computed tomographic scans of the chest. A thorough evaluation by an

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otolaryngologist is also often helpful because of the recurrent sinusitis, otitis media, and nasal polyps that might be present.

Alternatively, certain forms of pneumonia point to possible deficiencies in lung cells such as alveolar macrophages, lymphocytes, or PMNs. As opsonization of certain encapsulated bacteria is necessary for optimal phagocytosis by macrophages and PMNs, the lack of appropriate IgG antibodies against pneumococci, Haemophilus species, Klebsiella pneumoniae, and staphylococci may contribute to infections with these common bacteria.However, other causes of pneumonia may reflect abnormal lymphocyte function and cellmediated immunity. Infection with Legionella bacteria is an example. After an infection with L. pneumophila, the host develops specific IgM and IgG serum antibodies. These antibodies, in the presence of complement, do not create a lytic state that is sufficient to kill the bacteria.However, they do behave as opsonins to ensure that the Legionella organisms can attach and be ingested by various phagocytic cells, including PMNs, blood monocytes, and alveolar macrophages. Once inside the phagocytes, Legionellamultiply and eventually can kill and disrupt the host cells.When alveolar macrophages are activated with IFN-γ these stimulated phagocytes will inhibit the growth of the bacteria. This may be the result of the ability of IFN-γ to down-regulate the transferrin receptors on these cells, thereby limiting the accumulation of intracellular iron which is an essential metabolite for Legionella. Support for this concept comes from experiments with an experimental Legionella pulmonary infection rat model, in which administration of intratracheal IFN-γ reduced intrapulmonary replication of the bacteria, improving host defenses. Another example of defects at the level of the lymphocyte is AIDS, in which the human host is infected with human immunodeficiency virus (HIV) that destroys CD4 TH lymphocytes. These patients experience recurrent respiratory infections with diverse organisms, including viruses (cytomegalovirus or herpes simplex), Pneumocystis carinii, Mycobacterium tuberculosis, M. avium-intracellulare, fungi such asCryptococcus species, andToxoplasma gondii andLegionella.These infectious agents have a common feature of residing in macrophages or similar cells as facultative intracellular organisms.

One reason why a patient with AIDS has trouble with this group of infections relates to the relative imbalance of lymphocytes found in the alveoli, as sampled by BAL of the lung.Normal values for T lymphocytes have been given in Table 19-1. From subjects with AIDS, the recoverable alveolar lymphocytes reflect a decrease in the CD4 TH cells from HIV infection, offset by an increase in the suppressor-cytotoxic species of T lymphocytes. Although alveolar macrophages normally exist in an environment where they can be activated sufficiently to kill or control microbes of this sort, the CD4 deficiency in lungs of patients with AIDS compromises this activation process. This causes an impressive defect in cell-mediated immunity and the ability of macrophages to contain or kill organisms such as Pneumocystis or mycobacterial species.

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