The Immune System

2. The Immune System

Function

The immune system, in all its complexity, has evolved for the defense against infectious organisms from viruses, bacteria, and fungi, to large multicellular parasites. Immune responses range from non-specific barrier-type functions, to phylogenetically advanced, complex, adaptive responses that may involve destruction or elimination of the pathogen (Figure 3). A perfect response to infection would result in elimination without self damage. Immune responses are never perfect however, and damage to the host always occurs, ranging from undetectable to disproportionate, and at its most extreme, fatal.

Remembering this basic concept is essential in interpreting the effect of nutrition on immunity.

Figure 3. Features and functions of innate and adaptive immunity. Key points of nutritional modulation.

General Aspects of Immune Responses

Innate

Anatomical and physiological mechanisms that contribute to immunity and that are in place regardless of previous exposure, are referred to as "innate". Many of these mechanisms are phylogenetically ancient (e.g., lysozyme, phagocytes), whilst others are complex and have only evolved in vertebrates, becoming refined in mammalian species (e.g., natural killer cells) (Table 1).

In mammals, the initial role of innate immunity is to exclude micro-organisms where possible. When infection occurs, the innate responses to the pathogen result in any, or a combination of:

1. Elimination of infection
2. Limiting the initial progression of infection (the "speed-bump" for initial infectious agents)
3. Stimulation of adaptive immunity through the production of the early inflammatory response to infection. Thus innate immunity provides the "danger signals" that alert and activate adaptive immune responses.

Recognition of Microbes

Cells of innate immunity have evolved receptors that recognize phylogenetically conserved molecules. These molecular patterns have been termed pathogen associated molecular patterns or "PAMPS". Examples of PAMPS are lipopolysaccharide (LPS) from gram negative bacterial cell walls, lipoteichoic acid from gram positive bacterial cell walls, and double-stranded RNA from viruses. The PAMP receptors include scavenger receptors, mannose receptors, and the family of Toll-like receptors (TLR) (Akira, 2003). To date there are 10 known mammalian TLRs, although the expression of all 10 types has not yet been described in cats. Most TLRs are membrane proteins, although the TLR 9 binds to its ligand intracellularly (bacterial DNA). Binding of a TLR with its ligand results in the generation of the nuclear transcription factor NF-κB, which diffuses into the nucleus and binds to specific sites on the DNA of the host cell, leading to the transcription of a variety of pro-inflammatory genes. In macrophages and neutrophils these genes include cytokines (TNF-α, IL-1, and IL-12), adhesion molecules (E-selectin), cycloxogenase (COX), nitric oxide synthase (iNOS), and on macrophages the costimulatory molecules CD80 and CD86.

The net effect of TLR signaling in leukocytes is migration into inflamed tissues, enhanced killing of microbes or infected cells, and the production of inflammatory cytokines and chemokines to signal and activate the cells of the adaptive immune response (Figure 4).
Figure 4. Ligands and effects of toll like receptors (TLR) signaling.

Killing of Phagocytosed Microbes

Phagocytosed microbes remain within the membrane bound phagosome in the cytoplasm. Once internalized, these phagosomes then fuse with preformed lysosomes, which contain several proteases (e.g., elastase). In addition, activation of the phagocyte (e.g., by signaling through TLRs) results in assembly of the multi-subunit machinery of the NADPH-oxidase in the phagosome membrane, and within the plasma membrane. This enzyme complex catalyses the reduction of diatomic oxygen (O2) to the superoxide radical (O2 • –). The O2 • – is then enzymatically dismutated to produce hydrogen peroxide, a potent oxidant that may be partially responsible for microbial killing. However, the presence of myeloperoxidase within the phagosome utilizes the peroxide to produce a more potent antibacterial, hypochlorous acid (HOCl). This process of producing powerful oxidants following activation and phagocytosis by neutrophils and macrophages rapidly utilizes large amounts of available oxygen and is termed the respiratory burst (Figure 5) (DeLeo et al, 1999).
Figure 5. Respiratory burst and HOCl production.

Following activation of the phagocyte, the inducible form of nitric oxide synthetase (iNOS) is also expressed, resulting in the production of the free radical nitric oxide (•NO), which reacts with superoxide to form the toxic metabolite peroxynitrite (Eiserich et al, 1998). These various oxidants are not only confined to the phagosome, but are also released extracellularly to contribute to microbial killing in the immediate vicinity. Inevitably, this results in collateral oxidative damage to surrounding tissues.

To protect themselves from massive autogenously derived oxidative damage, phagocytes require greater concentrations of cytosolic (aqueous) and membrane (lipophilic) antioxidants, which are degraded and rapidly replenished during the respiratory burst. The most important antioxidants in this regard appear to be glutathione, ascorbate, tocopherol, and taurine. Feline neutrophils contain high intracellular concentrations of taurine. In fact, taurine constitutes 76% of the free amino acid cytosolic pool, compared with 44% in lymphocytes (Fukuda et al, 1982). Elimination of HOCl by the conversion of taurine to taurine chloramine protects cells against endogenously created oxidants. It has been also suggested that the taurine chloramine may also act as an intracellular signaling molecule that limits further O2• – and •NO production.

However, in cats maintained on taurine deficient diets, suppression of both phagocytosis and respiratory burst activity by neutrophils occurs, consistent with its role primarily as an antioxidant (Schuller-Levis et al, 1990).

Natural Killer Cells

Natural killer cells (NK cells) are large granular lymphocytes, distinct from T and B lymphocytes. NK cells are responsible for the identification and killing of virally infected and neoplastic cells, without prior exposure (sensitization). NK cells lyse target cells by releasing granules of the enzymes perforin, which creates pores in cell membranes, and granzyme, which enters the perforated cell and induces programmed cell death (apoptosis). Activated NK cells are also important secretors of IFN-γ, and are thus important activators of macrophages in the vicinity, increasing their phagocytic and respiratory burst capabilities.

Adaptive Immunity

Adaptive immunity is stimulated by infection, and by signaling from the innate immune system. With subsequent re-exposure to the infectious organism, the magnitude, specificity, and speed of the response increases, hence the term adaptive immunity. Adaptive immunity is the domain of the T and B lymphocytes, whereby humoral (antibody) responses or cellular responses are generated against specific molecules termed antigens (Figure 3).

Eicosanoids

Eicosanoids are a group of lipid messengers synthesized from the 20-carbon polyunsaturated fatty acids (PUFA) dihomo-γ-linolenic acid (DGLA; 20:3n-6), arachidonic acid (ARA; 20:4n-6) and eicosapentaenoic acid (EPA; 20:5n-3). Eicosanoids include prostaglandins (PGs), thromboxanes (TXs), leukotrienes (LTs), lipoxins, hydroperoxy-eicosatetraenoic acids (HPETE) and hydroxyeicosatetraenoic acids (HETE).

The fatty acid precursor for eicosanoid synthesis is released from cell membrane phospholipids, usually by the action of phospholipase A2, which is activated in response to a noxious cellular stimulus (Figure 6). Generally, the membranes of cells in cats on most commercial diets contain 5 to 10 times more ARA than EPA; thus ARA is usually the principal precursor for eicosanoid synthesis, giving rise to the 2-series PGs and TXs, and the 4-series LTs (Plantinga et al, 2005). However, the exact proportion of other 20 carbon PUFA in cell membranes is determined by the relative proportions of them, and their shorter 18 carbon precursors in the diet of the animal.

Figure 6. The production of eicosanoids from fatty acid precursors released from cell membrane phospholipids by the action of phospholipase A2.

PGE2 has a number of pro-inflammatory effects including inducing fever, increasing vascular permeability and vasodilation and enhancing pain and edema caused by other agents such as histamine (Harris et al, 2002). PGE2 suppresses lymphocyte proliferation and natural killer cell activity and inhibits production of tumor necrosis factor (TNF)-α, interleukin (IL)-1, IL-6, IL-2 and interferon (IFN)-γ. In these respects then, PGE2 is also immunosuppressive and anti-inflammatory. PGE2 does not affect the production of the Th2-type cytokines IL-4 and IL 10, but promotes immunoglobulin E (IgE) production by B lymphocytes. Therefore PGE2 supports a Th2-biased adaptive response, and inhibits Th1 responses.

LTB4 increases vascular permeability, enhances local blood flow, is a potent chemotactic agent for leukocytes, induces release of lysosomal enzymes, enhances the respiratory burst, inhibits lymphocyte proliferation and promotes natural killer cell activity. LTB4 enhances production of TNFα, IL-1 and IL-6 by monocytes and macrophages, and enhances Th1 cytokine production.

To add to the complexity, PGE2 inhibits 5-lipoxogenase, thereby interfering with production of LTB4, and ARA also gives rise to anti-inflammatory lipoxins. Thus, eicosanoids are pro- as well as anti-inflammatory, and together they regulate inflammation. The overall effect will depend upon the timing of production of the different eicosanoids, the sensitivity of target cells, and the concentrations of the different eicosanoids produced.