The activation of Rac and Rho helps to increase the microbicidal capacity of macrophages and neutrophils that have ingested pathogens. Upon phagocytosing microbes, macrophages and neutrophils produce a variety of toxic products that help to kill the engulfed microorganism (Fig. 3.4). The most important of these are the antimicrobial peptides described in Section 2-4, reactive nitrogen species such as nitric oxide (NO), and ROS, such as the super-oxide anion (O2–) and hydrogen peroxide (H2O2). Nitric oxide is produced by a high-output form of nitric oxide synthase, inducible NOS2 (iNOS2), whose expression is induced by a variety of stimuli, including fMLF.
Activation of the fMLF and C5a receptors is directly involved in generating ROS. Superoxide is generated by a multicomponent, membrane-associated NADPH oxidase, also called phagocyte oxidase. In unstimulated phagocytes, this enzyme is inactive because it is not fully assembled. One set of subunits, the cytochrome b558 complex (composed of p22 and gp91), is localized in the plasma membranes of resting macrophages and neutrophils, and it appears in lysosomes after the maturation of phagolysosomes. The other components, p40, p47, and p67, are in the cytosol. Activation of phagocytes induces the cytosolic subunits to join with the membrane-associated cytochrome b558 to form a complete, functional NADPH oxidase in the phagolysosome membrane (Fig. 3.5). The fMLF and C5a receptors participate in the process by activating Rac, which functions to promote the movement of the cytosolic components to the membrane to assemble the active NADPH oxidase.
Pattern recognition by cells of the innate immune system.
˜-Defensins (HNP1–4), °-defensin Cathelicidin, macrophage
HBD4, cathelicidin, azurocidin, Antimicrobial peptides bacterial permeability inducing protein (BPI), lactoferricin
Acidifcation Toxic nitrogen oxides Toxic oxygen-derived products pH°3.5–4.0, bacteriostatic or bactericidal Nitric oxide NO Superoxide O2 – , hydrogen peroxide H2O2, singlet oxygen 1O2 •, hydroxyl radical •OH, hypohalite OCl–
Lysozyme: digests cell walls of some Gram-positive bacteria Acid hydrolases (e.g., elastase and other proteases): break down ingested microbes Lactoferrin (sequesters Fe2+), vitamin B12-binding protein
The NADPH oxidase reaction results in a transient increase in oxygen consumption by the cell, which is known as the respiratory burst. It generates superoxide anion within the lumen of the phagolysosome, and this is converted by the enzyme superoxide dismutase (SOD) into H2O2. Further chemical and enzymatic reactions produce a range of toxic ROS from H2O2, including the hydroxyl radical (•OH), hypochlorite (OCl–), and hypobromite (OBr–). In this way, the direct recognition of bacterially derived polypeptides or previous pathogen recognition by the complement system activates a potent killing mechanism within macrophages and neutrophils that have ingested microbes via their phagocytic receptors. However, phagocyte activation can also cause extensive tissue damage because hydrolytic enzymes, membrane-disrupting peptides, and reactive oxygen species can be released into the extracellular environment and are toxic to host cells.