Innate Immunity: Physical Barriers & Microbiota

• Chapter 2 focuses on innate immunity and the body's immediate response to infection

Physical Barriers & Microbiota

• Physical barriers colonized by commensal microorganisms protect against infection by pathogens
• Mammalian babies have no commensal microorganisms before birth
• After birth, commensals from environment (family members, pets) begin to populate the skin and mucosal surfaces
• The microbiota is an integral part of a healthy human body and influences the development of the immune system
• Surface epithelia provide mechanical, chemical, and microbiological barriers to infection
• Skin: Epithelial cells joined by tight junctions, fatty acids, antimicrobial peptides, and normal microbiota
• Gut: Epithelial cells joined by tight junctions, longitudinal flow of air or fluid, low pH, antimicrobial enzymes, antimicrobial peptides, and normal microbiota
• Lungs: Epithelial cells joined by tight junctions, movement of mucus by cilia, pulmonary surfactant, and antimicrobial peptides
• Eyes/nose/oral cavity: Epithelial cells joined by tight junctions, tears, nasal cilia, antimicrobial enzymes in tears and saliva, and antimicrobial peptides

Pathogens

• Intracellular and extracellular pathogens require different types of immune responses
• Extracellular pathogens live and replicate in the spaces between human cells and are accessible to soluble, secreted immune system molecules
• Intracellular pathogens live and replicate inside human cells
• Host cells must be killed in order to expose intracellular pathogens to the soluble immune system molecules
• Bacteria, viruses, fungi, and parasites can be present in the extracellular spaces of infected tissue
• Viruses and some bacteria need to enter cells to replicate

Complement system

• The complement system is composed of plasma proteins that mark pathogens for destruction
• Soluble proteins are made by the liver and circulate in the blood, lymph, and extracellular fluid
• Many complement proteins are proteases that circulate as inactive zymogens until activated upon infection
• Complement proteins activate each other in a cascade via proteolytic cleavage
• Activation results in covalent attachment of C3b to a pathogen's surface
• C3 cleavage produces C3b and C3a fragments
• C3b is chemically reactive and covalently attaches to the pathogen's surface, marking it as dangerous
• C3a recruits phagocytic cells to the site of infection

Complement Activation Pathways

• Alternative: the first pathway to be activated and a part of innate immunity initiated by direct interaction with a pathogen
• Lectin: part of innate immunity that is initiated by mannose-binding lectin in plasma
• Classical: Initiated in the innate response by the binding of C-reactive protein to pathogen surfaces, and in the adaptive response by the binding of antibodies to pathogen surfaces
• All pathways result in C3 cleavage to C3a and C3b and marking the pathogen for destruction, recruitment of immune cells, and/or perforation of the pathogen cell membrane

Alternative Pathway

• Complement activation proceeds by the alternative pathway at the start of the infection
• Bacterial surfaces induce changes in the local physiochemical environment
• Changes trigger the hydrolysis of serum C3, giving C3(H2O), also called iC3
• iC3 binds to inactive complement factor B
• Factor D cleaves the bound factor B, giving iC3Bb, a soluble C3 convertase, which cleaves C3 into C3a and C3b
• C3b fragments covalently attach to the pathogen surface
• The alternative C3 convertase, C3bBb, works at the surface of the pathogen
• The alternative C3 convertase C3bBb works at the surface of the pathogen
• It functions similarly to the soluble iC3Bb, but since it is pathogen-bound, it stays nearby
• C3bBb molecules cleave more C3 and fix more C3b at the pathogen surface
• This process is a positive feedback loop that leads to rapid coating with C3b

Regulatory Proteins

• Formation and stability of C3bBb on the cell surface is determined by complement control proteins
• Factor P: extends the lifetime of C3 convertase on the microbial surface
• Factor I: assisted by factor H, cleaves and inactivates C3bBb
• DAF and MCP: disrupt C3bBb on the human cell surface

Phagocytosis by Macrophages

• Macrophages are mature forms of circulating monocytes that have left the blood to take up residence in the tissues
• They're prevalent in connective tissues and GI and respiratory tracts
• Macrophages participate in both innate and adaptive immunity
• Complement receptors on phagocytes trigger the uptake and breakdown of C3b-coated pathogens
• CR1 on the surface of macrophages recognizes C3b-coated pathogens, and generates intracellular signals that enhance phagocytosis and the fusion of the phagosome with lysosomes

Terminal Complement Proteins

• Terminal complement proteins lyse pathogens by forming membrane pores
• C5, C6, C7, C8, and C9 form the membrane-attack complex
• Activation of C5 yields the soluble C5b fragment, initiating membrane-attack complex assembly
• C6 binds to and stabilizes C5b and forms a binding site for C7
• C7 binds to C5b6 and exposes a hydrophobic region for attachment to the cell membrane
• C8 binds to C5b67 and exposes a hydrophobic region that inserts into the cell membrane
• C9 polymerizes on the C5b678 complex, forming a membrane-spanning channel that disrupts cell integrity, resulting in cell death
• Binding of C3b to the alternative C3 convertase produces C3b2Bb, the alternative C5 convertase
• The C3b component of C3b2Bb cleaves C5 into C5a and C5b
• C5b initiates the formation of a membrane-attack complex (MAC), which can make holes in the membranes of bacterial and eukaryotic cells

CD59

• The human cell surface protein CD59 binds to the C5b678 complex
• CD59 prevents the recruitment of C9 and its polymerization in the membrane to form a pore, preventing the assembly of the MAC on the cell surface
• Homologous Restriction Factor (HRF) works the same way

Inflammation

• C3a and C5a fragments induce anaphylactic shock
• C3a and C5a are referred to as anaphylatoxins
• The receptors are located on phagocytes, endothelial cells, and mast cells
• Roles in inflammation include increased blood flow and vascular permeability, and act as chemoattractants (C5a)

Inflammation & The Spread of Infection

• Anaphylatoxins act on blood vessels to increase vascular permeability and promote a greater innate immune response
• Increased permeability results in increased fluid leakage from blood vessels, plus extravasation of complement and other plasma proteins
• Migration of monocytes and neutrophils from blood into tissue is increased
• Microbicidal activity of macrophages and neutrophils is also increased
• Coagulation, Kinin and Protease Inhibitor systems limit the spread of infection

Plasma Protein Classes

• Several classes of plasma protein limit the spread of infection
• Coagulation System: Plasma enzymes form blood clots, immobilize pathogens, and prevent them from entering the blood and lymph
• Kinin System: Plasma proteins cause vasodilation and increase the supply of innate immunity components to the infected site
• Protease Inhibitors: Inhibit pathogenic cell-surface or secreted proteases; examples include a2-Macroglobulins

a2-Macroglobulin

• a2-Macroglobulin inhibits potentially damaging proteases and contains a highly reactive thioester bond that inhibits microbial proteases
• It traps the microbial protease with a 'bait' region, and binds the protease covalently through activation of the thioester group
• It enshrouds the protease so that it cannot access other protein substrates, even though the protease is still catalytically active

Antimicrobial Peptides

• Antimicrobial peptides kill pathogens by perturbing their membranes
• Human B1-defensin is a short alpha-helix segment resting against three strands of antiparallel beta-sheet
• Defensin is an amphipathic peptide with separate regions having charged or hydrophobic residues
• Defensin interacts with the charged surface of the cell membrane, and inserts into the lipid bilayer which forms pores that leads to a loss of membrane integrity

Defensins & The Immune System

• Alpha-defensins HD5 and HD6 (cryptdins) are only made by Paneth cells located in the crypts of the small intestine
• Paneth cells also secrete other antimicrobial factors, including lysozyme and phospholipase A2
• Though considered epithelial, rather than hematopoietic in origin, Paneth cells classify as cells of the immune system

Pentraxins

• Pentraxins are plasma proteins of innate immunity that bind microorganisms and target them to phagocytes, their role is similar to antibodies
• 2 types of Pentraxins: Short Pentraxin and Long Pentraxin
• Short Pentraxin: Serum amyloid P component, which is produced by liver hepatocytes
• Long Pentraxin: PTX3, produced by monocytes, macrophages, dendritic cells, endothelial cells, epithelial cells
• Serum C-reactive protein (CRP) is a principal member of the pentraxin family

Chapter 2 explores innate immunity, focusing on the body's immediate response to infection. It highlights physical barriers and the role of microbiota in protecting against pathogens. Surface epithelia provide mechanical, chemical, and microbiological barriers to infection in the skin, gut and lungs.