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Title: P1253553542JxWqy


1
Ch.14 - Cell-Mediated Effector Response (C-MER)
2
Ch.14 - Cell-Mediated Effector Response (C-MER)
  • The principle role of the cell-mediated immunity
    is to detect and eliminate cells that harbor
    intracellular pathogensas well as tumor cells
    that have undergone genetic modification so that
    they present antigen not typical of normal cells
    (p.319)

3
Cell Organization of C-MER
  • Cells with direct cytotoxic activity
  • Antigen specific
  • CD8 Cytotoxic T cells (TC cells or CTLs)
  • Nonspecific
  • Natural Killer Cells (NK cells)
  • Macrophages
  • Cells that mediate the delayed-type
    hypersensitivity reactions (DTHR)
  • CD4 TH cells
  • TH1 cells
  • TH2 cells

CD8 Cytotoxic T cells (TC cells or CTLs)
TH1 cells TH2 cells
4
Effector T Cells
  • Composed of three kinds of cells
  • CD8 TC cells
  • TH1 TH2 cells
  • Characterized by
  • Less Stringent activation requirements
    (Table14-1)
  • CD28-B7 interaction NOT necessary for activation
  • Increased expression of cell-adhesion molecules
  • 2-4x increased expression of CD2 integrin LF1
  • Production of effector molecules (Table14-2)
  • Membrane-bound
  • Soluble

CD8 TC cells TH1 TH2 cells
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6
Figure 10-15
7
Effector T Cells
  • Composed of three kinds of cells
  • CD8 TC cells
  • TH1 TH2 cells
  • Characterized by
  • Less Stringent activation requirements
    (Table14-1)
  • CD28-B7 interaction NOT necessary for activation
  • Increased expression of cell-adhesion molecules
  • 2-4x increased expression of CD2 integrin LF1
  • Production of effector molecules (Table14-2)
  • Membrane-bound
  • Soluble

CD8 TC cells TH1 TH2 cells
8
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9
Functions of T-cell Effector Molecules
  • Mediate target-cell destruction by TC cells
  • Fas ligand (membrane-bound)
  • perforins (soluble)
  • granzimes (soluble)
  • Promote macrophage activity
  • TNF-b (soluble membrane-bound)
  • INF-g (soluble)
  • GM-CSF (soluble)
  • Play role in B-cell activation by TH2 cells
  • CD40 (membrane bound)
  • IL-4, IL-5, IL-6 (soluble)

TC cell
TH1
TH2
10
Focus Cytotoxic T cells
  • Generated by Immune activation of TC cell
    precursors
  • Have lytic capabilities
  • Play critical role in recognition of altered self
    cells
  • MHC I restricted (generally)
  • All nucleated cells in body express MHC I
    molecules

11
Generation of Effector Cytotoxic T cells
  • Requires three specific signals
  • Signals for Activation
  • Primary antigen specific TCR(CD8)-Ag-MHC I
    interaction
  • Co-stimulatory CD28-B7 interaction
  • (may not be necessary for Memory TC cell
    precursors )
  • Signals for Proliferation Differentiation
  • Signal from IL-2 interaction with high-affinity
    IL-2 Receptor
  • Memory TC cell precursors may produce enough
    IL-2 to self-proliferation
  • Generally TC cell precursors (CTL-Ps) need IL-2
    produced from TH1 cells for proliferation

12
Figure 14-1
13
Figure 14-2
14
Figure 14-3
  • Attach
  • Four MHC I molecules bound to a specific peptide.
  • Biotin
  • Fluorescently labeled Streptavidin
  • MHC Tetramers bind to specific TC cell with
    affinity for specific Ag-MHC I chosen.
  • TC cell can be counted and separated by flow
    cytometry
  • sensitivity to cell populations as
    low as 0.1

15
Ag-MHC I complex
16
Ag-MHC I complex
17
Figure 14-4
  • Study demonstrated that acute infections result
    in varying percent distributions of Ag-MHC
    specific Cytotoxic T cells
  • (in respect to total population of TC cells in
    respective tissues)
  • Ranging from 0.6 in peripheral lymph nodes to
    41 in kidneys.

18
Cytotoxic T cells Granule Mediated Homicide
  • 1) Conjugate formation
  • 2) Membrane attack
  • 3) TC cell dissociation
  • 4) Target cell obliteration (destruction)

19
Figure 14-6
20
Figure 14-5 SEM of Cytotoxic T cell-target cell
conjugate
21
Cytotoxic T cells Granule Mediated Homicide
  • 1) Conjugate formation
  • Recognition TCR-Ag-MHC I interaction
  • Embrace LFA-1 receptor (TC cell) binds to ICAMs
    on target cell membrane
  • LFA-1 is induced to a high-avidity state
    mediated by antigen
  • The LFA-1 high-avidity state regulates TC cell
    adhesion only to appropriate target cells
    displaying specific antigen.
  • Note The LFA-1 high-avidity state is only
    active for 5-10min and its deactivation may
    facilitate TC cell dissociation.
  • 2) Membrane attack
  • 3) TC cell dissociation
  • 4) Target cell obliteration (destruction)

22
Figure 14-7 Antigen activated CTL activation
23
Cytotoxic T cells Granule Mediated Homicide
  • 1) Conjugate formation
  • 2) Membrane attack
  • Cytoplasmic rearrangement
  • Brings Golgi and storage granules into closer
    proximity to target cell
  • Granule Secretion (exocytosis)
  • Perforin - 65kDa monomer
  • Undergoes conformational change upon contact with
    target cell membrane which exposes amphipathic
    domain, enabling insertion into membrane.
  • Once in membrane perforins polymerize and create
    5-20nm pores (w/Ca2).
  • Granzyme
  • 3) TC cell dissociation
  • 4) Target cell obliteration (destruction)

24
Figure 14-9a Perforin pore formation in target
cell membrane
25
Figure 14-9b EM of perforin pores in target cell
membrane
26
Cytotoxic T cells Granule Mediated Homicide
  • 1) Conjugate formation
  • 2) Membrane attack
  • Cytoplasmic rearrangement
  • Granule Secretion (exocytosis)
  • Perforin - 65kDa monomer
  • Granzyme
  • Binds to mannose 6-phosphate receptor and
    internalized into target cell. The Perforin pores
    allow the Granzyme to exit internalized vesicles.
  • Once inside cytoplasm of target cell, initiates
    reaction cascade culminating in activation of
    endonucleases which in turn digest DNA into
    oligomers of 200bp (typical of apoptosis).
  • 3) TC cell dissociation
  • 4) Target cell obliteration (destruction)

27
Cytotoxic T cells Fas Ligand Mediated Homicide
  • Fas
  • Transmembrane protein
  • Member of the TNF-receptor family
  • Can deliver death signal when crosslinked with
    its natural ligand
  • Natural ligand is a TNF called Fas ligand (FasL)
  • 2) FasL
  • Found on the membrane of TC cells
  • Interaction with Fas protein triggers target cell
    apoptosis
  • 3) Fas-FasL interaction
  • elucidated by experiments with perforin KO and
    1pr strain mice
  • Homozygous 1pr strain mice express little to no
    Fas
  • Fig 14-10 Fig 14-10 summary

28
Figure 14-10 Fas-FasL interaction experiments
with KO mice
29
Figure 14-10 Fas-FasL interaction experiments
with KO mice
30
Figure 14-10 Summary
  • Results
  • only two apoptotic signaling mechanism
  • granule mediate and Fas mediated homicide

31
Cell Death by Apoptosis
  • Caspase
  • Family of cysteine proteases which cleave after
    Asp residue
  • Normally present in cell as inactive proenzymes -
    procaspases
  • gt12 caspases with different specificity have been
    identified
  • Cleavage of procaspase produces an active
    initiator caspase, which in turn cleaves other
    caspases.
  • Both Granule and Fas mediated apoptotic signaling
    induces the caspase cascade (Fig14-11) by
    activating Procaspase-8.
  • Results in systematic disassembly of the cell

32
Figure 14-11 Caspase Cascade
33
Cell Organization of C-MER
  • Cells with direct cytotoxic activity
  • Antigen specific
  • CD8 Cytotoxic T cells (TC cells or CTLs)
  • Nonspecific
  • Natural Killer Cells (NK cells)
  • Macrophages
  • Cells that mediate the delayed-type
    hypersensitivity reactions (DTHR)
  • CD4 TH cells
  • TH1 cells
  • TH2 cells

CD8 Cytotoxic T cells (TC cells or CTLs)
Natural Killer Cells (NK cells)
TH1 cells TH2 cells
34
Fig. 2-1 Hematopoiesis
35
Natural Killer (NK) Cells
  • Discovered when negative control (unimmunized)
    mice showed significant lysis of tumor cells.
  • Compose 5-10 of recirculating lymphocyte
    population
  • Involved in immune defense against virus and
    tumors
  • Play important role in immune regulation
  • Influence both adaptive and innate immunity via
    cytokine production/excretion
  • INFg
  • Affects phagocytic and microbial activities of
    macrophages
  • Influences TH1 cells vs TH2 cells commitment of
    development
  • First line of defense in viral infections
  • Number of NK cells peaks 3 days after infection
  • Rapid rise in NK population closely follows rise
    in levels of cytokines (Fig14-12)

36
Fig. 14-12 Time course of viral infection
37
Comparison between NK and T Cells
38
Comparison of NK and T-cell Assassination
Mechanism
  • Similar to processes employed by CTLs
  • Express FasL on membrane surface
  • Contain Granules of perforin and granzimes
  • Target cell degradation occurs via perforins and
    granzymes
  • Different from CTLs cytotoxicity
  • NK always cytotoxic, do not need to be activated
    to produce granules
  • Do not express Ag specific T-cell receptors or
    CD3
  • Recognition of target cells is NOT MHC restricted
  • NK immune response generates no immunological
    memory
  • No greater immune response upon secondary
    infection

39
Natural Killer (NK) Cells
  • Express inhibition and activation receptors on
    cells surface
  • Many inhibition and many activation receptors
    create an opposing-signal model.
  • The balance between the opposing signals is
    believed to enable NK to differential between
    healthy and infected cells (Fig 14-14)
  • Additional NK activator signals can be delivered
    by soluble factors
  • TNF-a, IL-12, and IL-15
  • NK cells may target cells that produce aberrant
    MHC expression
  • Many virus-infected and tumor cells have reduced
    MHC expression

40
NK Cells Inhibitor-Receptor Superfamily
  • C-type-lectin-inhibitory receptor (CLIR)
  • In humans CD94/NKG2 - disulfide bonded
    heterodimer of two glycoproteins
  • Recognizes HLA-E on potential target cells
  • HLA-E serves as indicator of overall level of MHC
    I biosynthesis
  • Thus CD94/NKG2 are not specific for specific HLA
    allele
  • Killer-cell-inhibitory receptors (KIR)
  • A group of Ig-superfamily-inhibitory receptors
    (ISIR)
  • more than 50 family members have been found
  • Specific for one or more of polymorphic HLA
    products
  • Inhibitory receptors have veto power over
    activation receptors
  • Thus, cells expressing normal levels of MHC I
    receptors tend to escape all forms of NK
    assassination.
  • Thus cells that lack normal MHC I expression
    lack of normal self expression DIE!!!

41
Fig. 14-14 Opposing-signals model of cytotoxic
activity
42
Fig. 14-15 Time course of viral infection
43
Fig. 14-16 One-way mixed lymphocyte reaction
(MLR)
44
Fig. 14-16 In vitro cell-mediated lympholysis
(CML) assay
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