Transcript NK Cells

Cell Mediated Immunity
W. Robert Fleischmann, Ph.D.
Department of Urologic Surgery
University of Minnesota Medical School
[email protected]
(612) 626-5034
Objectives
• To understand the role of cell-mediated
immunity
• To understand the identity, mechanism of
activation, and method of killing by cytotoxic T
cells
• To understand the identity, mechanism of
activation, and method of killing by natural
killer (NK) cells
• To understand the method of killing by
antibody-mediated cellular cytotoxicity
(ADCC)
Arvak Petrosian, age 2 years, is brought from
Armenia to a private clinic in the U.S. that
specializes in diagnosis and treatment of
unusual diseases.
Arvak has a history of failure to gain weight.
He appears to be anorexic. In addition, he is
noted to have eczema.
He has recently been diagnosed with Type I
diabetes. According to his Armenian
physicians, he is now showing signs of
hypothyroidism.
What are your thoughts?
What tests would you wish to request?
A complete blood cell count shows
Low levels of neutrophils
Low levels of RBCs
Low levels of platelets
T and B cell mitogen stimulation is normal.
Antibody isotypes and sub-types are at normal levels.
What are your thoughts?
What tests would you wish to request?
Urinalysis shows the presence of protein in the urine.
Blood chemistry shows elevated levels of serum
creatinine and urea nitrogen.
Liver enzymes are significantly elevated.
What are your thoughts?
What tests would you wish to request?
Arvak is diagnosed as having
Eczema
Enteropathy (destruction of cells in the intestine)
Diabetes
Neutropenia
Anemia
Thrombocytopenia
Thyroiditis
Glomerulonephritis
Hepatitis
What are your thoughts?
Role of Cell-Mediated Immunity
• Humoral immunity mediated by antibody is responsible for
the protection of the extra-cellular environment from
pathogens and toxins.
• The principal roles of cell-mediated immunity are to
recognize and destroy pathogen-infected cells and cells
that have undergone genetic alterations (tumor cells).
• The importance of cell-mediated immunity can be seen in
patients who lack a thymus (DiGeorge Syndrome).
– Although antibodies are not made, these patients deal relatively
well with extracellular bacteria.
• DiGeorge patients do suffer repeated infections because no memory.
– Infections with viruses, intracellular bacteria, and fungi are difficult
to clear and become life-threatening in DiGeorge patients.
The Mediators of CellMediated Immunity
• Antigen-nonspecific effector cell
– Natural Killer cells
– Macrophages
– Neutrophils
– Eosinophils
• Antigen-specific effector cells
– CD4+ T cells (aka, helper T cells, Th cells)
– CD8+ T cells (aka, cytotoxic T cells, CTLs,
Tc cells)
Different Effector T Cells Can Be
Recognized by the Effector Molecules
They Produce
Cytotoxic T Cells
Cytotoxic T Cells
• Naïve CTL cells are called CTL precursors (CTL-P) to
indicate their immature state.
• A CTL-P cell matures only after being activated by
interaction with a Th1 cell.
• Maturation requires three sequential signals.
– Antigen-specific signal is transmitted by the TCR upon
recognition of peptide:Class I antigen presented by a
professional antigen presenting cell (licensed antigenpresenting cell) or by a tissue cell.
– Costimulatory signal is transmitted by CD28:B7 interaction
between the CTL-P cell and the licensed antigen-presenting
cell.
– A signal induced by IL-2 secreted by a Th1 cell, results in
proliferation and differentiation of the antigen-activated CTLP cell to a CTL.
•
Before an APC can
activate a CTL-P
cell by peptide:MHC
class I presentation,
it must be “licensed”
to do so.
Activation of
Effector CTLs from
Naïve CTL-Ps
– APC must present
peptide via MHC
Class II to a Th1
cell.
– Somehow, this
interaction
licenses the APC
to present peptide
via MHC Class I
antigen to the
CTL-P cell.
•
It is not known
whether the Th1 cell
and the CTL-P cell
are bound to the
APC at the same
time.
Note: Naïve CTL-P = naïve CTL precursor
Importance of Licensing of APC
• Licensing of APC requires interaction of the
APC with the Th1 cell.
• Thus, presentation of antigen to the Th1 cell via
MHC class II and to the CTL-P cell via MHC
class I acts a control to prevent recognition of
self antigen (antigen must be a phagocytosed
antigen to be expressed on Class II).
– This is a function best carried out by a dendritic cell
acting as an APC.
– Infection of a dendritic cell by a virus assists in the
presentation of antigen associated with MHC class I.
Activation of Memory CTLs
• Th1 cell not required to provide IL-2.
• Autocrine IL-2 is sufficient.
Naïve CTL-P Versus Mature CTL
• Naïve CTL-P
– Does not express IL-2 or IL-2R and only begins to express them
after it begins to be activated.
– Expresses the CD45RA isoform of CD45.
– Expresses a low level of cell adhesion molecules CD2 and LFA-1.
– Does not exhibit cytotoxic activity.
• Mature CTL
– Expresses high affinity IL-2 receptor and requires high levels of IL2 to proliferate.
– Synthesizes low levels of IL-2.
– Expresses the CD45RO isoform of CD45.
– Expresses a high level of cell adhesion molecules CD2 and LFA-1
– Exhibits cytotoxicity.
• Memory CTL cells may not require Th1 help.
– Requires only low levels of IL-2 (produced by activated CTL-Ps)
for memory CTLs to become mature effector CTLs.
Overview of CTL Killing
•
Primary events
– Conjugate formation
– Reorientation of cytoplasmic
granules in CTL
– Granule exocytosis
– Dissociation
– Target cell death by apoptosis
Pictures of
CTL Killing
Note that the T cell approaches
the target cell, forms a close
binding to the target cell, and
reorients its granules (small
arrow) toward the target cell.
CTL Killing: Binding of Target Cell
• The TCR-CD3 membrane complex on the CTL
recognizes peptide bound to MHC class I on a
target cell.
• LFA-1 on CTL binds to ICAMs on target cell.
• Antigen activation converts LFA-1 from a lowaffinity state to a high-affinity state, resulting in
the formation of the conjugate.
• After 5-10 minutes, the LFA-1 returns to a lowaffinity state, resulting in dissociation of the
CTL from the target cell.
Bound CTLs Kill Targets by
Two* Mechanisms
• Perforin/Granzyme are secreted from granules
– Perforin molecules form pore
– Granzyme molecules activate apoptosis by
cleavage of caspases
• FasLigand protein on cell membrane surface
– Membrane-bound FasL binds to Fas on the
membrane of the target cell to initiate cell killing
– Activates apoptosis by cleavage of caspases
* CTLs can also kill by TNF production and
secretion
CTL Killing: Degranulation Events
• The CTLs (but not naïve CTL-Ps) contain intracellular granules.
– Contain monomers of perforin
• Perforin monomers polymerize to form pore in the target cell
• Perforin has some homology with C9 and forms pores similar to complement pores
– Contain serine proteases called granzymes or fragmentins
• Granzymes taken up by receptor mediated endocytosis (granzyme binds to
mannose 6-phosphate receptor) or through perforin pore (perforin-assisted
pathway)
• Granzymes induce apoptosis
Apoptosis Induction
by FasL and
Granzyme
• FasL on the CTL binds to Fas
(TNF family receptor) on the
target cell, activating its death
domains to bind to FADD (FASassociated Death Domain) that,
in turn, activates caspase-8
• Granzymes initiate apoptosis by
activating caspase-8
Natural Killer Cells
Natural Killer Cells (NK Cells)
• NK cells make up 5-10% of the population of
circulating lymphocytes.
• NK cells play major roles in killing virusinfected cells, intracellular pathogen-infected
cells, and tumor cells.
• NK cells produce a number of important
cytokines, including IFN-.
– IFN- can tilt the immune response toward Th1
cells by inhibiting Th2 and by inducing IL-12
production by macrophages and dendritic cells.
– IFN- can activate macrophages and NK cells.
• NK cell activity is stimulated by IFN-, IFN-,
IFN-, TNF-, IL-12 and IL-15.
NK Cells Are an Early
Defense Against Viruses
NK Cells versus CTLs
• NK cells express CD16, FcRIII.
• NK cells do not need to be educated by the thymus.
• NK cells do not undergo rearrangement of receptor
genes and, thus, do not express T cell receptors or
CD3.
• NK cell killing is not MHC restricted (identical killing
levels are seen for allogeneic and syngenic tumor
cells).
• NK cells do not show immunologic memory that can
be primed by re-exposure to antigen.
• NK cell killing is similar to CTL killing
– FasL expressed on surface and can kill cells expressing Fas
– Perforin and granzyme released from granules
– TNF expressed on surface and secreted
NK Cell Receptors
• NK cells have both activation and inhibition
receptors.
• NK cell receptors fall into two categories but
members of each group can be activation or
inhibition receptors.
– Lectin-like receptors
• Bind proteins rather than polysaccharides
• Can be activation or inhibition receptors
– Immunoglobulin-like receptors (aka, KIR = killer
cell immunoglobulin-like receptors)
• Bind to most class I MHC molecules
• Can be activation or inhibition receptors, but most are
inhibition receptors
Opposing Signals Model of
NK Cell Killing
• If an antigen recognition
signal is given to the NK
cell and the level of MHC
class I is high, no killing
occurs.
• If an antigen recognition
signal is given to the NK
cell and the level of MHC
class I is low, killing
occurs.
NK Cells and CTL Have
Complementary Activities
NK Cells
• Kill cells that mask
the presence of
foreign antigen on
MHC class I
CTLs
• Kill cells that
express foreign
antigen on MHC
class I
NKT Cells
NKT Cells
• NK cells express lectin-like receptors and KIR.
• CTL express T cell receptor and CD3.
• NKT cells express T cell receptor but differs from CTL.
– Maturation is in the thymus.
– The T cell receptor is invariant (specific gene segments encode
TCR and TCR chains).
– The T cell receptor does not recognize MHC-bound peptides but
does recognize a glycolipid presented by CD1d.
– NKT cells do not exhibit memory.
– NKT cells express cell surface markers characteristic of NK cells.
– NKT cells may play a role in killing bacteria and tumor cells but
precise role is uncertain.
• Bacteria killing via expression of glycosyl ceramides (glycolipid)?
• Tumor cell killing via expression of glycolipids that are specific to
tumor cells?
Antibody-Dependent
Cell-Mediated Cytotoxicity
(ADCC)
ADCC Killing
• Effector cells associated with
ADCC
–
–
–
–
–
NK cells
Macrophages
Monocytes
Neutrophils
Eosinophils
• Effector cells bind antigen via
antibody bound to Fc receptor
• Killing is mediated by
– Cytolytic enzyme release by
macrophages,neutrophils,and
eosinophils
– TNF release by NK cells,
monocytes, and macrophages
– Perforin release by NK cells and
eosinophils
– Granzyme release by NK cells
Arvak Petrosian
Because of the number of organ
systems under autoimmune attack, Arvak
is diagnosed with a rare condition called
IPEX syndrome (immune dysregulation,
polyendocrinophathy, enteropathy, Xlinked syndrome).
This syndrome is caused by mutations
in the FOXP3 gene.
Arvak Petrosian
FOXP3 gene function is required for the
development of Treg cells.
In the absence of appropriate Treg cell
function, CD8+ cytotoxic T cells are active
and destroy normal tissue cells that they
recognize. Thus, peripheral tolerance is
lost, resulting in autoimmunity.