Chapter 3

Induced Innate Response

Recognition

• immune receptors distinguish features of microbial structure
• microbes look different
• innate cells have receptors that detect differences
• Pattern Recognition Receptors (PRRs)
• Pathogen Associated Molecular Patterns (PAMPs)
• most microbial ligands are carbs and lipids
• lectin: receptors that recognize carbs
• distinguishing self from non-self
• Natural killer cells recognize stress signals on virus infected cells

Elimination (phagocytosis)

• macrophages
• Toll-like receptors (PRR)
    * trigger signaling instead of phagocytosis
    * 10 of them
    * form dimers
      * some homo, some hetero
    * recognize all 4 types of pathogen
• TLR 4
    * LPS (endotoxin) binds to TLR 4, signal transduction, NFkB activated, expression of inflammatory cytokines
• defects have dire consequences
    * NEMO deficiency
    * NFkB activation impaired
    * development issues
• Cytokines
    * nomenclature
      * originally called monokines and lymphokines
    * proteins secreted by cells of innate and adaptive immune that mediate many functions of those cells
    * interleukins (IL_)
    * chemokines
    * specific names
      * interferons (viral response)
      * tumor necrosic factor (TNF)
• General Characteristics
    * proteins (polypeptides)
      * produced in response to microbes/other antigens
      * induce/regulate immune response
    * production is brief due to tight regulation
      * synthesis initiated by
      * mRNAs unstable
      * regulated post transcriptionally
    * fast
    * pleiotropism
      * 1 cytokine → multiple effects
    * redundancy
      * 2 cytokines → 1 job
    * synergy and antagonism
      * 2 cytokines → amplified response
      * 2 cytokines → opposite jobs
    * act locally or systemically
    * function by binding to specific membrane receptors on target cells
      * high affinity → very potent
      * receptor expression regulated → contributes to specificity
    * change gene expression in target cells
    * cellular response tightly regulated
      * feedback inhibition
      * inhibition of signaling
• What do activated macrophages do?
    * PRR + PAMP = NFkB activation an cytokine expression
    * internalized LPS triggers inflammasome that releases IL-1 beta

Cytokines

• NFkB turns of expression of genes to produce cytokines
• IL-1 beta
    * produced by macrophages in response to gram-negative bacteria
    * different pathway
    * make inflammasome
    * acts synergenically with TNF- alpha
• IL-6
    * increase rate of matabolism in fat and muscle cells
    * generates heat
    * slows down rate of replication for bacteria
    * increases rate of immune cell function
• CXCL8
    * recruits neutrophils from blood to site of infection
• CCL2
    * recruites monocytes from blood to tissue
    * monocytes become macrophages or dendritic cells
• IL-12
    * recruits and activates natural killer cells to secrete cytokines to help macrophages
• TNF- alpha
    * produced as a result of TLR stimulation
    * strong TNF response
      * local → good
      * systemic → causes septic shock symptoms
        * fluid from blood leaks into all tissue, edema, bp drops
        * decreased blood volume leads to vesicles collapsing on themselves → death

Phagocytes

• macrophages
    * from monocytes (CCL2)
    * long lived
    * reside in tissue
    * work from beginning
    * raise alarm
    * repair tissue after
• neutrophils
    * short lived
    * circulated in blood
    * receive “call” from macrophages
    * only function is to phagocytose
    * most abundant white blood cell
      * around 50 billion
      * can double or triple with infection
      * most respond to signal and migrate into tissue
• 2 types of macrophages
    * M1 macrophages
      * inflammatory
    * M2 macrophages
      * anti-inflammatory
      * gut and respiratory
• Neutrophils
    * selectin expressed on vascular cells, velcro interaction with s-Lex

Extravasation

• movement of cells from blood into tissue
• Rolling
    * velcro between selection and s-Lex
    * slows neutrophil down and rolls along
    * white blood cells have time to listen for the call before flying by
• Tight Binding
    * stronger adhesion molecules keep neutrophil in place
    * CXCL8 binds to neutrophil
    * TNFalpha leads to vasodilation, loosen tight junctions
    * TNFalpha tuns of expression of vascular adhesion molecule ICAMP-1
    * neutrophil is grabbed by ICAM-1 binding to LFA-1
    * CXCL8 binds to receptor on neutrophil, change in conformation leads to stronger bond between ICAM-1 and LFA-1
• Diapedesis
    * neutrophils squeezing through vasc. endo cells to get out of blood stream
    * neutrophils become pancake
• Migration
    * neutrophils follow CXCL8 conc. gradient to find SOI

Phagocytose

• neutrophils covered in receptors to detect pathogens (PRRs)
• PRR binds to PAMP and engulfs it into endosome called phagosome
• membrane remodels around it
• bacteria in phagosome
• granules: vesicles full of bad stuff to kill bacteria
• granules fuse with phagosome
• Specific granules have NADPH oxidase
    * causes respiratory burst
    * critical to start killing bacteria
    * raises pH inside of phagosome
• lysosome fuses to make phagolysosome
• phagolysosome lowers pH back down, degraded
• neutrophil can do about 6-8 bacteria before running out of granules
    * neutrophil dies
• macrophages eat dead neutrophils
• dead neutrophils create neutrophil extracellular trap
    * mostly chromatin
    * traps bacteria
• NET causes some collateral tissue degradation
• some bacteria secrete potent toxins making NET lead to a lot of inflammation and tissue degredation

Acute Phase Response

• part of inflammatory
• IL-6 binds to liver cells to turn on acute phase response
• makes C-reactive protein and mannose-binding lectin (and others)
• C-reactive protein
    * binds to bacteria
    * targets phosphocholine
    * acts as opsonin
• Mannose-binding lectin
    * binds to mannose on bacteria
    * opsonizes
• Opsonization and Activation of complement
• Lectin Pathway
    * triggered by mannose binding lectin
    * mannose binding lectin bound to lectin
    * C4 binds to mbl, cleaved into C4a and C4b with C4b left on the membrane
    * C2 binds and is cleaved with C2a staying behind and making convertase (C4bC2a)
    * C3 convertases
      * alternative: C3bBb
      * classical: C4bC2a
• Classical pathway
    * triggered by c-reactive protein
    * binds to pathogen surface
    * C1 binds to protein
    * C1 looks like mannose binding lectin
• Acute phase proteins (CRP and MBL) activate complement through respective pathways
• IL-6 binds to liver cell, turns on CRP and MB, opsonization and activate complement

Interferon Response

• all cells can induce interferon response
    * any cell with nucleus can get infected, so any can respond
• internal PRRs recognize viral nucleic acid
• activates transcription factors (NFkB) secrete inflammatory cytokines
• IRF3 activates interferon response
• produces type 1 interferon
    * interferon is a type of cytokine
    * inhibits viral replication
      * oligoadenylate synthetase
      * PKR
    * recruit and activate natural killer cells
    * warn neighbors
• IFN-alpha and IFN-beta are type 1 interferons
• viral response is systematic
• plasmacytoid dendritic cells (pDCs) make lots of type 1 interferons in blood
    * secrete 1000 fold more interferon than any other cells
    * “professional” interferon producing cells with weird lineage and morphology
    * catch wind of interferon signal and multiply it
• Natural Killer Cells
    * activated by type 1 interferons
    * change from naive NKs to effector NKs and proliferate
    * look for MIC ligand
      * activating ligand of NK
      * flag for infected cells
    * healthy cells have an inhibitory ligand
    * releases cytotoxins, inducing apoptosis
• inhibit viral response, warn neighbors, recruit and activate NK