Transcript slides - Smith Lab

Introduction to Immunology of the Eye
Nonself
Self
Other people’s cells
Material from other life forms
Worms
Dangerous Self and Commensal Nonself
NON-SELF
SELF
Dangerous
Not Dangerous
Two Theaters of the Immune System
Humoral: Target is Outside the Cell
A soluble protein or other molecule
that interacts with a cell free Nonself molecule or organism and
neutralizes or eliminated the bound
item.
Examples: Antibodies and
Compliment
Cell Mediated: Target is Inside
the Cell or the cell itself
Immune cell interacts with
other cells, and destroys or
alters the function of that cell
Examples: CD8 T cell, NK Cells,
Gamma Delta T cells
Innate Receptors
How the Immune System Recognizes
Nonself and Danger
•
•
•
•
•
Ancient: Found in worms and flies
Recognize molecular motifs common to microbes
Encoded in your Germline DNA
Expressed throughout the body
Toll-like Receptors (TLR) are the “Flagship” Innate immune
receptor
• TLRs are expressed on the corneal epithelium
Adaptive Receptors
– (Mucosal Immunology (2015) 8, 1000–1008)
TLR: Germline Receptors that recognize
Pathogen Associated Molecular Patterns
Adaptive Receptors
Innate Receptors
How the Immune System Recognize
Non-self and Danger
•
•
•
•
•
•
Ancient: Found in worms and flies
Recognize molecular motifs common to microbes
Encoded in your Germline DNA
Expressed throughout the body
Toll-like Receptors (TLR) are the “Flagship” Innate immune receptor
TLRs are expressed on the corneal epithelium
– (Mucosal Immunology (2015) 8, 1000–1008)
•
•
•
Found in vertebrates
Expressed only in specialized immune cells
Generated through recombination of the DNA in specialized immune cells
–
–
•
•
•
B cells
T cells
Receptor Specificity is Random
Molecular Targets/Ligands are referred to as “Antigens”
The B and T cell populations needs to be “educated” and activated in order to
respond to Antigen
Adaptive Immune Receptors:
RAG Generate Receptors
The Adaptive Immune Response:
Clonal Expansion and Selection is a microcosm of
natural selection
The Immune System
Adaptive
Innate
Humoral
Cell Mediated
Natural Killer Cells
Complement
The branches of the immune system
interact and overlap
Cell Mediated
Adaptive
Humoral
Innate
Immunology of the Eye
Innate Humoral
The Tear Film
• Tear film has antiseptic properties
• Tear Film Contains Amylases, Peroxidases, Lysozyme
• Defensins are small, 29-45aa, proteins
– α-defensins: released by immune effector cells
– β-defensins: expressed by epithelial cells
– θ-defensins: these exist, at least in non-human primates
• They have intrinsic anti-microbial properties
• Defensins in the tear film
– α-defensins: found in the tear film after insult: infection or
transplant
– β-defensins: constituently present in the eye and in
response to infection of the cornea
β-defensins
• hBD-1 constitutively expressed in the eye
• hBD-2 is upregulated in corneal epithelial cells
following exposure to PAMPs
• hBD-3 fits both parameters
• Mutations in Defensins have been shown to
increase replication of bacteria on corneal
surface
Ocul Surf. 2004 Oct; 2(4): 229–247.
Corneas of Beta-Defensin KO mice (Left) and WT (right)4.5 h after inoculation P. aeruginosa
strain PAO1-GFP (green).
Danielle K. Augustin et al. Infect. Immun. 2011;79:595-605
Immunology of the Eye
<12 hrs post-Insult
Innate Cell Mediated:
Attracting Immune Cells
• TLR ligation can activate
corneal resident immune
cells-dendritic cells
• TLR ligation of dendritic
cells and the corneal
epithelium induces
chemokine expression
• Chemokines are soluble
molecules that attract cells
• CXCL1 (KC): Attracts
Neutrophils
• CXCL2 (MIP-2): Attracts
Macrophages and
Neutrophils
Dendritic Cells
Surface Markers
CD11b
CD11c
MHC II
CD8alpha
Neutrophil Surface Marker:
CD11bHI
Gr-1HI
Ly6C lo ly6Ghi
Macrophage Surface Markers:
CD11b
F4/80
MHC II
ViralNeutrophils
Replicationand
in the
Macrophages
Cornea TwoCan
Days
Not
After
Directly Target
HSV-1
theInfection
Virally Infected Cell
Neutrophils and Macrophage
Migrate into the cornea within 24hrs
• Non-specific tissue
damage through the
release of reactive
oxygen species
• Phagocytosis of dead,
dying or stressed cells
• Release of more
Chemokines and
Cytokines
Innate Cell Mediated:
Attracting More Immune Cells
• TLR ligation of cells in the
corneal epithelium induces
chemokine expression
• Chemokines are soluble
molecules that attract cells
• CXCL1 (KC): Attracts
Neutrophils
• CXCL2 (MIP-2): Attracts
Macrophages and
Neutrophils
• CCL5: Attracts NK cells
• CXCL10: Attracts NK cells
NK Cells
• Express lytic
granules
• NK are always ready
to kill
• Decision to kill is
result of integration
of activation
Receptors and
Inhibitory
Receptors Signals
Surface Markers
CD56 (humans)
CD16 (humans)
DX5 (mice and humans)
NKp46 (mouse)
NK1.1 (C57BL/6 mice)
The Lytic Hit
Cornea 4 days After HSV-1 Infection
4000
3000
2000
1000
te
B
d
S
0
N
K
D
e
p
le
P
pfu/cornea
• Lytic Granule: Low pH to keep
enzymes inactive
• Release into a extracellular
space between the target cell
and NK cells
• Granzyme A: enter the
nuclease and activates DNA
repair machinery
• Granzyme B: Acts on the
mitochondria to induce the
release of cytochrome C and
induce Apoptosis
• Perforin: Pore forming protein
necessary for granzyme entry
into the cell
Immunology of the Eye
48 hrs post-Insult
Dendritic Cells Junction between
Innate and Adaptive Immune
Response
Dendritic Cells
Surface Markers
CD11b
Corneal Dendritic Cells
CD11c
-PROFESSIONAL ANTIGEN PRESENTING CELLS
MHC II
-Activated During Infection
CD8alpha
-Produce Chemokines
-Travel to the Draining lymph node
Dendritic Cells Junction between Innate
and Adaptive Immune Response
• Both Macrophage and
Dendritic Cells are
“Professional Antigen
Presenting Cells” APCs
• Both will take up
Antigen
• Both will present
antigen to T cells
• Dendritic cells will
leave the cornea
Professional APCs and T cells
• T cells can only interact with
Antigen when it is loaded into
a MHC molecule
• Two Types of T cells
– CD8+: Killer T cells
– CD4+: Helper T cells
• CD8 Cells Interact MHC Class 1
• CD4 Cells Interact MHC Class 2
• MHC 1 expressed by every cell
• MHC 2 expressed only by APCs
Adaptive Cell Mediate Immunity
• Dendritic Cells Express
Both MHC 1 and MHC 2
• Activated Dendritic cells
express Co-stimulatory
molecules
– B7 Family (CD86, CD80)
and TNFR Family(OX40L,
CD70)
• Naïve T cells that
encounter antigen
without co-stimulation
will not become effector
cells
Dendritic Cells Take Antigen Out of the
Cornea in Search of T cells (>48hrs)
• Depletion of Corneal
Dendritic Cells Prior to
HSV-1 infection is lethal
• Antigen Can Also Drain
From the Cornea
Independent of DCs
• Primed T cells Return to
the Cornea (7 days
post-insult)
Priming of B cells
• B cells are Present in the
Draining Lymph Node
• B Cells Bind Extra Cellular
Antigens
• Strong Ligation of the BCR
will lead to B-cell
proliferation
• Strong Ligation of the BCR
will lead to production of
IgM
B cell
Antibodies: Adaptive Humoral
Immunity
Antigen Binding
(Variable) Region
Constant Region
IgA is necessary for
adaptive humoral
immunity in the eye
Priming of B cells
B cell
• B cells are Present in the
Draining Lymph Node
• B Cells Bind Extra Cellular
Antigens
• Strong Ligation of the BCR
will lead to B-cell
proliferation
• Strong Ligation of the BCR
will lead to production of
IgM
• B cells require CD4 help for
antibody class switching
CD4 T cell are required for Antibody
class switching
• Typically, antigen alone
is not enough to prime
a B cell response
• CD40L expressed by
CD4 cells is necessary
for B cell class switching
and survival
• The cytokines produced
by the CD4 cells direct
antibody class switching
CD4 T cells are required for Antibody
Class Switching (7-14 days post-insult)
The eye maintains it own plasma cell
population (>14 days post-insult)
• IgA Plasma Cells
are resident in
Lamina
Propria/lacrimal
gland
• This allows
antibody transfer
to tear film and
ocular surface
• Deficiencies in IgA ,
class switching or
CD4 T cells leads to
chronic
conjunctivitis
• <12 hrs tears film destroys
bacteria
• >12 hrs PAMP driven
Inflammation leads to increase
chemokine production
• <24 hrs Neutrophils and
immature Macrophages enter the
cornea
• 24-48 hrs Macrophages and
Dendritic cells mature
• 48 hrs NK cells enter the cornea
• >48 hrs Dendritic cells travel to
the DLN
• 3-7 days Dendritic Cells Prime the
T cell responses
• 7 days T cells travel to cornea
• 7 days IgM production
• >7 days B cells class switching
• >14 days Plasma Cells become
resident in the lacrimal gland
• Infiltrate will shrink
• IgA will be present in tear film
Allergies
• There are 4 types of
allergic responses
• The common rapid
allergic response that
causes itchy eyes and
air way stress is Type 1
• Type 1 is mediated by
IgE responses
Type 1 Allergies are mediated by innate cells
armed with adaptive molecules
• Mast cells and Eosinophils
are granulocytes
• They express FC receptors
with a high affinity for IgE
• These innate cells
become armed with
adaptive receptors
• IgE ligation cause rapid
degranulation
• Common therapeutics in
eyed drops are mast cell
stabilizers (cromolyn)
The choice between IgE or IgG is
control CD4 T cell subtypes
ILC (innate Lymphoid Cells)
Regulate the Innate Response
• ILCs are present in tissue
• Respond to alarmins
– IL-25
– IL-33
• Based on the alarmin
ratios ILCs will
differentiate into ILC
subtypes
• IL-33 is constitutively
expressed in cornea
epithelium and most
mucosal tissue