Phagocytosis: Phagocytes (eating cells) are body cells specialized for capture, ingestion and destruction of antigens (as bacteria and fungi), debris, and produce inflammatory molecules which regulate other components of the immune system. They express a wide range of surface receptors that allow them to identify microorganisms. Also, phagocytosis can be enhanced by antibodies, complement and acute phase proteins (all called opsonins and act as a bridge between the antigens and phagocytic cells). They are of TWO types:

1.Polymorphnuclear leucocytes (microphages); mainly neutrophils. They are short-lived cells with a half life of 6 hours. 2. Mononuclear cells (macrophages); Monocytes after 7-10 hours in the blood they migrate to tissues. In C.T. (histiocytes), or fixed in RES (e.g., liver, spleen, L.N., B.M… etc.). The process of phagocytosis include the following stages:

Mononuclear Phagocytes (RES) Granulocytes Polymorphonuclear leukocytes (PMN, neutrophils) Eosinophils Basophils (blood) Mast Cells (tissues) Mononuclear Phagocytes (RES) Monocytes (blood) Macrophages (tissue) fight pathogens inflammation allergic and hypersensitivity reactions fight pathogens inflammation cytotoxicity immune regulation

Neutrophils (PMN) Present in blood (60-70% of WBC) Not normally present in tissues Short lifespan - 12 hours Functions: First cell at the site of infection/injury Ingest and kill microbes after bactericidal mechanisms activated (binding to pathogen)

Mononuclear Phagocytes Blood - monocytes (1-6% WBC) Tissues - macrophages mature form of monocytes found in tissues (ex., gastrointestinal tract, lung, liver, brain, skin, spleen); reticuloendothelial system (RES) Functions: Phagocytize and kill after bactericidal mechanisms activated Produce cytokines/chemokines (initiate inflammation) Antigen presentation (activate adaptive immunity) Tumor surveillance and cytotoxicity

Reticuloendothelial System Consists of fixed and wandering macrophages located throughout the body (tissues, sinusoids, lymph system) Major locations: Liver and Lung Functions: antibacterial resistance, tumor resistance, defense against shock, antigen processing, lipid metabolism, protein turnover, iron metabolism

A. Chemotaxis,Migration, & Attachment: If epith. lining is breached by microbes, resident phagocytes are attracted (influx) to the site by chemotactic substances as bacterial endotoxins (LPS), serun C5a, IL-8, and leukotriene B4.

The macrophages produce cytokines as IL-1 & TNF The macrophages produce cytokines as IL-1 & TNF.These activate endothelial cells of nearby venules to produce adhesion molecules (selectin, integrins, & ICAM) and chemkines (e.g.,IL-8) which mediate MIGRATION of leucocytes & monocytes from the blood to tissues (diapedesis). The phagocytes have receptors on their surface through which they ATTACH non-specifically to m.o. Examples: Receptors for bacterial endotoxines, for mannose residues on glycoprotein of many bacteria, unmethylated bacterial DNA & double stranded RNA of many viruses. The attachment and ingestion is enhanced if microbes are coated with IgG or C3b (opsonization).

Pattern Recognition Receptors (PRR) Three broad classes based on expression profile, localization, function PRR that signal an infection: 1. Toll Receptor Family (Toll-like Receptor “TLR “ 1-11) Expressed externally or internally Binding activates “pro-inflammatory” signaling pathways 2. Phagocytic (endocytic) PRR Expressed on the surface of phagocytic cells Mediate uptake of microbe into phagocytes 3. Secreted PRR Secreted by MP, epithelial cells, hepatocytes Activate complement, opsonins, function as accessory proteins for Pathogen-Associated Molecular Pattern (PAMP) recognition on target cells such as microbes. These signaling pathways are evolutionarily conserved.

Toll-Receptor Family:

How do Macrophages Identify Microbes? Pattern Recognition Receptors (PPR ) Recognize pathogen associated molecular patterns (PAMP); conserved molecular patterns on microbes Identify a class of microbes; ex., LPS, LTA, peptidoglycan, lipoarabinomannan, dsRNA, mannose, b-glycans PAMP are often essential for microbe survival Action Time Immediate activation of effectors Delays need for adaptive immunity LPS – from Gram negative LTA – lipotechoic acid – from Gram positive Peptidogkycans Lipoarabinomannan - from microbacteria) dsRNA -produced by most viruses Mannans – from fungi B-glucans – from fungi

Leukocyte Adhesion selectins

B. Ingestion: The phogocytes engulf the antiges (e.g. organism) by extending pseudopods around them. Then the organism is included into vacuole called PHAGOSOME. Lysosomal granules fuse with the phagosome forming PHOGOLYSOSOME. Then is followed by DIGESTION of organism.

C. Intracellular killing or Digestion: Anti-microbial and cytotoxic substances are produced that destroy phagocytosed m.o. by TWO mechanisms: 1. Oxygen-dependent (Respiratory Burst): This killing system is mediated by NADPH oxidase enzymes complex, which converts oxygen into reactive oxygen species such as hydrogen peroxide and superoxide that are lethal to micoorganisms. When combined with myeloperoxidase, hypochlorous ions (HOCl -, analogous to bleach) are highly effective oxidants and anti-microbial agents.

2. Oxygen-independent: Killing in this system is the result of lysozomal contents which include lysozyme, lactoferrin, a group of cationic proteins (defensins), and hydrolytic and proteolytic enzymes. Some bacteria resist destruction for long periods, others multiply within the phagocytes (M. tuberculosis, M. leprae). Wandering phagocytes transport these m.o. to new places (spread infection).

Functions of macrophages: Initiation and amplification of the inflammatory response. Killing of microorganisms. Resolution and repair of inflammation. Link between innate and adaptive immune system.

Cont…/Second line of innate immunity… 3. Natural Killer (NK) cells: They comprise 10-15% of the peripheral lymphocytes. They are CD3 (-), CD11b (+), CD16 (+), and CD 56 (+). They have non-specific CYTOTOXIC activity against tumour cells, graft cells, & virus infected cells. Their function is NOT restricted by MHC. Their activity is increased by INF, IL-2 & 12. They have surface receptor to IgG - Fc (CD16).

4. Inflammatory barriers: Inflammatory response has 3 events: 1. Vasodilatation of nearby capillaries leading to redness of tissues, increase tissue temperature, increase capillary permeability and influx of fluid and cells (exudate) causing oedema. 2. Influx of phagocytes which engulf m.o. & release lytic enzymes that can result in tissue damage. 3. Chemical mediators are released from damaged tissues, m.o., leucocytes & plasma enzyme systems ( histamine,kinin,fibrine,cytokines &APP).

Functions of Phagocytes: Inflammatory Responses Injury or Hallmarks of an Inflammatory Response Redness Heat Swelling Pain

Constitutional Factors affecting Innate Immunity: 1. Species : Some organisms are pathogenic to certain species, e.g., M.leprae affects man, but not monkey. 2. Race : Example; Negroes & red Indians are more susceptible than whites to M. tuberculosis. 3. Individuals: GENETIC influences susceptibility (e.g., G6PD deficiency resistant to malaria). Extremes of AGE ; due to immaturity or aging of immune system in children & old respectively.

4. Nutritional status: Under nutrition increases susceptibility to disease. 5. Hormones: Example: corticosteroid & other immunosuppressive drugs, diabetics (lack insulin) or pregnancy increase susceptibility to infections. Cytokines of innate Immunity: From activated macrophages: IL-1, IL-2,TNF which increase extravasation of neutrophils, induce coagulation & increase vascular permeability. Also IL-12 which activate NK cells. INF-gamma (from NK) activate macrophages. INF-alpha (from viral infected cell) inhibits V.

4. Nutritional status: Under nutrition increases susceptibility to disease. 5. Hormones: Example: corticosteroid & other immunosuppressive drugs, diabetics (lack insulin) or pregnancy increase susceptibility to infections. Cytokines of innate Immunity: From activated macrophages: IL-1, IL-2,TNF which increase extravasation of neutrophils, induce coagulation & increase vascular permeability. Also IL-12 which activate NK cells. INF-gamma (from NK) activate macrophages. INF-alpha (from viral infected cell) inhibits V.

Summary: Aims of the immune system: I. Protection: by specific &non-specific immunity 2.Hypersensitivity (I,II,III& IV). 3.Tolerance and autoimmunity. 4. Immunodeficiency diseases. Introduction to immunological words as antigens, antibodies, complement ,T cells, B-cells, HLA (MHC) system, CD markers, Monoclonal antibodies & cytokines. Branches of recent and current IMMUNOLOGY. Innate immunity. Details.