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