2 5 Innate immunity.pdf

MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi Innate Immunity Learning Objectives 1. Compare and contrast innate and adaptive immunity. 2. Discuss the natural barriers to infection. 3. List the cellular and non-cellular components of innate immunity and outline their contribution to host defen...

MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi Innate Immunity Learning Objectives 1. Compare and contrast innate and adaptive immunity. 2. Discuss the natural barriers to infection. 3. List the cellular and non-cellular components of innate immunity and outline their contribution to host defense. 4. Discuss the principles involved in pathogen recognition by innate cells, e.g., phagocytes. List the types of receptors found on innate cells and explain how each functions. 5. Outline the process of neutrophil recruitment and activation. 6. List the major participants in the NF- B signal transduction pathway and explain how the process works. 7. Describe the process of intracellular killing by phagocytes, including the roles of specific enzymes and their products. 8. Explain how the innate system affects the adaptive system. 9. Discuss tests to demonstrate the efficacy (or detect defects) of innate immunity and give specific examples of tests and their application. References Basic Immunology: Functions and Disorders of the Immune System. 5th Ed. 2016. Abbas, AK, Lichtman, AH, & Pillai, S. Elsevier. St. Louis, MO. Chapter 2, pp 27-53 How the Immune System Works, 6th edition, 2019. Lauren Sompeyrac. Wiley-Blackwell, Lecture 2 Immunology at a Glance. 10th Ed., 2013. Playfair, JHL, & Chain, BM. Wiley-Blackwell. Hoboken, NJ. 1 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi Innate Immunity The innate immune defenses constitute an old and well-established system that form early barriers to infectious disease agents and play a critical role in preventing infection. The microorganisms that are encountered daily in the life of a normal healthy individual only occasionally cause clinical disease. Most are detected and destroyed within minutes or hours by defense mechanisms that do not require a prolonged period of induction because they do not rely on the clonal expansion of antigen-specific lymphocytes. Indeed, the initial response to a perceived threat (the fight/flight response) is essentially nonimmunologic, and it may help explain how stress influences the immune response to infection. While the individual may consciously respond to a physical threat, infections occur silently, and the immune response proceeds autonomously. The innate immune system is a series of non-antigen-specific mechanisms that are present at a constant level or slightly increase on challenge but are not selected or amplified. There are two broad categories of innate mechanisms: cell-based systems (external body surface barriers and internal phagocytic cells) and a circulating protein system (e.g., complement, acute phase proteins, defensins). Instead of antigen specific receptors, the phagocytes have invariant receptors that recognize bacterial components. While distinguishable from the adaptive system, the innate system has important roles in the activation of B and T lymphocytes and in the effector functions of antibodies. The innate system of host defense against infection is made up of several distinct components. The first mechanism is barrier functions of the body’s epithelia that prevent the establishment of infections. Cells and various molecules that destroy pathogens that have gained access to the body constitute the second mechanism. The third mechanism comprises induced responses by elements that recognize and react against pathogens non-specifically. The innate immune system can be viewed as one that forms the first line of short-term defense that acts mainly to frustrate the establishment of an infectious focus. It acts locally at the site of an infection and involves phagocytic cells (macrophages, neutrophils) as well as a specialized subset of lymphocytes that kill infected cells nonspecifically (NK cells). It also activates the adaptive immune system, which provides longer-term defense against pathogens. 2 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi Innate immune responses are mediated by natural, constitutive, inborn defense mechanisms that act nonspecifically to limit and eliminate pathogens. The major areas at which natural defense factors are found are The skin The respiratory tract The digestive tract, and The genitourinary tract 3 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi The Components of Innate Immunity The intact skin constitutes a formidable barrier against infections. The outer most layer of the skin is composed of dead cells, the corneocytes, but the deeper layers contain living keratinocytes that may secrete such cytokines as interleukin-8 (IL-8) and tumor necrosis factor (TNF- ). IL-8 is a potent chemoattractant for neutrophils. The actions of TNF- are numerous and affect a broad range of cells. Resting macrophages can be induced by TNFto synthesize IL-1 (interleukin 1) and PG-E2 (prostaglandin E2) both of which are proinflammatory substances. The skin also contains Langerhans cells and dendritic cells, which present processed antigen to T cells of the SALT (skin associated lymphoid tissue) and the local lymph node. The tight junctions between cells at all locations provide a barrier to penetration by pathogens. The upper respiratory tract contains mucus secreting goblet cells and ciliated epithelium, making up the mucociliary escalator that traps particles and sweeps them upwards for 4 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi expulsion by sneezing or coughing. The lower tract is protected by surfactants (collectins) secreted by Type II pneumocytes. Surfactants perform the dual role of lubricating the alveoli and preventing their collapse during expiration and facilitating phagocytosis by binding to pathogens. The main defense factors in the gastrointestinal tract are stomach acidity and the intestinal flora. The gut, especially the large intestine, contains the highest concentration of resident bacteria in the body and these perform a number of important functions including the production of bacteriocins, vitamin synthesis, and participation in sterol metabolism. An intact epithelium and the presence of antimicrobial peptide secreting Paneth cells keep the intestinal organisms in check. The genitourinary tract is protected by the flushing action of urine, as well as by low pH of urine and vaginal secretions and the presence of bactericidal enzymes such as lysozyme. The other principal cells of innate immunity are phagocytes (macrophages, neutrophils), dendritic cells, epithelial cells, and natural killer (NK) cells. Phagocytes are widely distributed throughout the body, and they acquire special designations depending on their location. They are functionally similar however and the basic mechanisms of recognition, activation, and effector function are the same. Dendritic cells are antigen-presenting cells and are derived from bone marrow precursors. They may be classified as myeloid dendritic cells (mDC) derived from monocytes, plasmacytoid dendritic cells (pDC) derived from lymphoid cells, and follicular dendritic cells (FDC) derived from mesenchymal stem cells. Epithelial cells provide a barrier to infection and also modulate the recruitment and activation of phagocytes. Natural killer (NK) cells are large granular lymphocytes. They are sometimes called “null” cells because they do not express characteristic B or T cell markers. Natural killer cells target virusinfected or abnormal cells and destroy such cells in the absence of a specific trigger. About 5% of peripheral blood lymphocytes are NK cells. NK cells express receptors for interleukin-2 (IL- 2R) and can proliferate in response to T cell mitogens and IL-2. NK cells use killing mechanisms induced via receptors such as Fc RIII (CD16) and KAR. Other substances that are necessary for effective functioning of innate immunity include complement components, acute phase proteins such as C-reactive protein and mannan binding lectin, kinins and various enzymes (phospholipase, ribonuclease, amidase, etc.). Acute inflammation is the result of innate immune response to infection or injury. Changes occur in the vasculature, which permit phagocytic cells, notably neutrophils, to upregulate the expression of integrins that facilitate their emigration from the vascular to the extravascular compartment where they may ingest and destroy microorganisms. The cardinal signs of inflammation are redness (rubor), swelling (tumor), heat (calor), and pain (dolor). The pus that forms at the site of a ‘pyogenic’ infection is composed of dead neutrophils and bacteria. Cells that are unable to express integrins due, for example, to a mutation in CD18, fail to migrate and cannot affect an inflammatory response. 5 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi Recognition/Cognitive Function The recognition or cognitive function of cells of the innate system occurs by a mechanism that is clearly distinctive. Innate immune recognition is mediated by germ-line encoded receptors. Thus, the specificity of each receptor is genetically predetermined, and it is limited in number by the size of the organism’s genome. Unlike the adaptive immune response, in which, as we shall see, there is the capacity for gene recombination and somatic hypermutation events with the generation of billions of receptors, the innate immune system is programmed to respond rapidly to just a few thousand highly conserved structures present on most pathogens. These receptors are known as Pattern Recognition Receptors (PRRs). There are three types of PRRs: Type 1 PRR: Secreted molecules that circulate in blood and lymph Type 2 PRR: Cell-surface receptors that bind pathogens for engulfment Type 3 PRR: Cell-surface receptors that bind pathogens and initiate signals for the release of cytokines The expression of pattern recognition receptors is not clonal. All such receptors displayed by cells of a given type have identical specificities. This explains why all phagocytes in any one individual are reactive to the same pathogens. In addition, because the receptors are pre-existent, the response by phagocytes is immediate, an important feature of an innate immune response. Type 1 Secreted PRRs include the mannan-binding lectin, a surfactant protein-like collectin, that binds microbial carbohydrates to initiate the lectin pathway of complement activation. Mannan binding protein is synthesized in the liver and is secreted into the serum in an acute phase response. C-reactive protein is also a secreted PRR. 6 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi Type 2 Phagocytosis PRRs, e.g., scavenger receptors, mediate the uptake and delivery of pathogens into lysosomes where they are destroyed by phagocytosis. Pathogen derived proteins are also processed and are presented in association with MHC molecules. Phagocytosis PRRs appear to possess a high affinity for carbohydrates with large numbers of mannose residues. Type 3 Signaling PRRs are found on the cell membrane as well as within the cell. Cell membrane signaling PRRs are called Toll-like receptors (TLRs) – because of their homology to the toll family of receptors found in drosophila. Some TLRs are also found in the endosome. TLRs are found on and in macrophages, dendritic cells and epithelial cells. The binding of TLRs to pathogens results in the activation of signal transduction pathways that induce the expression of a variety of cytokine genes. Cytoplasmic signaling receptors are called NLRs or NOD-like receptors (nucleotide oligomerization domain-like receptors). 7 MDBS 1102 Innate Immunity 8 Dr. Raymond F Adebiyi MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi TLRs function as PRRs that bind to microbial products, leading to the activation of a transcription factor NF- κB (nuclear factor) signaling pathway. The initiation of this pathway turns on genes for such cytokines as TNF-α and IL-1. Mammals have many TLRs (TLR-2, TLR-3, TLR-4, TLR-5, TLR-9), and it appears that each recognizes a different microbial product (PAMP). The recognition of microbial lipopolysaccharide by TLR4 is mediated by co-factors CD14 and MD2. The PRRs described above recognize molecular structures that are unique to pathogens, especially, prokaryotes. These structures (1) are not shared with the host (2) are commonly found on many pathogens, and (3) are relatively invariant. 9 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi These microbial molecular structures are called the Pathogen Associated Molecular Patterns (PAMP). PRRs, on host cells, recognize PAMPs on pathogens. As mentioned above, some PRRs appear to recognize different PAMPs. For example: TLR-2 binds to peptidoglycan of Gram-positive bacteria, like staphylococci and streptococci TLR-3 binds to dsRNA TLR-4 binds to LPS (endotoxin) of gram-negative bacteria TLR-5 binds to flagellin of motile bacteria. Activation and Effector Functions Chemotaxis: Macrophages at the site of an infection secrete the cytokines IL-8, TNF- , IL- 1 and PGE2. These cytokines are responsible for the recruitment of neutrophils. In addition, there is an enhanced expression of P selectin and integrins (ICAM-1) by endothelial cells of local capillaries and CD18 by neutrophils. The result is that neutrophils become marginated and adherent to capillary endothelium. The cells undergo diapedesis, moving from the vasculature into the extravascular space, and then undergo directional 10 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi chemotaxis to the site of infection. In the absence of CD18, neutrophils are unable to leave the blood vessels and this may result in a form of leucocyte adhesion deficiency (LAD). Phagocytosis: An important function of the phagocytes of the innate immune system is to engulf and destroy microbes. Resting phagocytes, with the exception of alveolar macrophages, derive their energy from anaerobic glycolysis. The intracellular accumulation of lactic acid is mildly bactericidal. The phagocytic granules also contain many enzymes, such as lysozyme, cationic proteins, phagocytin, transferrin, and lactoferrin. These materials may mediate a relatively modest method of oxygen-independent killing of microorganisms. Following activation, phagocytes begin to consume oxygen (the respiratory burst) and the metabolic pathway shifts to the greater energy efficient hexose monophosphate shunt. Glucose-6-phosphate dehydrogenase is required for this shift to occur. Associated with the HMP shunt is the enzyme nicotinamide-adenine phosphate (NADPH) oxidase. The activity of this enzyme results in the conversion of oxygen to the super oxide radical and the formation of hydrogen peroxide and singlet oxygen. Nitric oxide (NO) is also produced following the induction of NO synthetase. Hydrogen peroxide, oxygen radicals and nitric oxide are potently microbicidal and are responsible for the effective killing of microbes by phagocytes. In the presence of myeloperoxidase and halide ions, hydrogen peroxide is converted to hypohalide ions, and the reaction proceeds spontaneously to produce water, halide ion, and singlet oxygen. Singlet oxygen is unstable and a return to the ground state emits light energy. This chemoluminescence is observed during phagocytosis. 11 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi The intracellular killing capacity of phagocytes may also be demonstrated by the reduction of nitroblue tetrazolium (NBT). A low NBT score indicates an inability to kill pathogens while a high NBT score indicates adequate intracellular killing capacity. This killing capacity is dependent on the activity of NADPH oxidase. Killing by NK Cells NK cells kill infected and abnormal host cells via two main mechanisms. 1) They possess killer activating receptors (KAR) that bind to PAMPs and initiate the release of perforin and granzyme. This is essentially a purely innate killing function. This action may however be abrogated by the killer inhibition receptors (KIR) that bind MHC-1 on normal host cells. KIR ensures that normal host cells are protected from NK action. 2) NK cells also possess Fc RIII (CD16) with specificity for Fc of IgG bound to microorganisms. Engagement of CD16 results in the expression of FasL and binding to Fas leads to caspase activation. This is an adaptive killing mechanism known as antibody mediated cellular cytotoxicity, ADCC. It should be noted that CD16 is also present on other cells such as neutrophils and macrophages which kill using reactive oxygen species (described above). 12 MDBS 1102 Innate Immunity Dr. Raymond F Adebiyi The phagocyte and the NK cell are obviously major components of innate immunity, but many accessory molecules are required for the proper functioning of the immune system. These include the complement proteins and their derivatives, cytokines (IL-1, IL-2, IL-6, IL- 8, IL-12, TNF- , PG-E2, etc.), acute phase proteins such as interferon (IFN- , IFN- γ, IFN- ), collectins, C-reactive protein as well as many enzymes. The innate system starts off the immunological reaction to pathogens and then activates the adaptive immune system. The adaptive immune system responds to a pathogen only after the pathogen has been recognized by the innate system. Cells of adaptive immunity, e.g., T cells, recognize peptides bound to MHC molecules. However, such recognition is not sufficient for T cell activation. A co-stimulatory signal is required, and the signal is controlled by the innate immune system. Recognition of a peptide in the absence of costimulatory signals results in apoptosis of T cells. 13