Transcript Leukocyte/endothelial interactions are a major event in the

Leukocyte Endothelial Interactions
Judith Berliner, Professor
Departments of Biology and Medicine, UCLA
Leukocyte/endothelial
interactions are a major event in
the inflammatory process
Inflammation is a reaction to
injury or infection
Inflammation and repair
1. Tissue exposed to infection, toxin, trauma
2. Damaged cells rapidly produce endothelial
activators
3. In some cases, venules transiently increase
permeability in response to activators
4. Plasma proteins enter the tissue and react
with bacteria and toxins producing more
activators
Inflammation and repair
5. Activated endothelial cells upregulate
leukocyte adhesion molecules and
chemotactic factors
6. Leukocytes bind to and enter the vessel
wall
7. Acute: Neutrophils kill and macrophages
engulf bacteria and toxins. Resolution
8. Chronic: macrophages are unable to
remove source of injury. Granuloma
Inflammation and repair
9. Cytokines and growth factors produced by
injured cells stimulate replication of
nearby cells. Fibrosis
10. Angiogenesis occurs in response to
additional growth factors.
11. Tissue architecture is restored
Activators of the endothelium to
induce leukocyte EC interactions
1. Bacteria
2. Toxins
3. Autoantibodies
4. Oxidized Phospholipids
5. Advanced glycosylation end products
6. Cytokines: IL-1, TNF
LIPOPOLYSACCHARIDE
OM
peptidoglycan
IM
lipid A
INITIATION OF INFLAMMATION
• RECOGNITION OF MICROBES BASED
ON CONSERVED PATTERNS
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LIPOPOLYSACCHARIDE (Gm-)
LIPOPEPTIDES (BACTERIA)
PEPTIDOGLYCAN (MOST BACTERIA)
FLAGELLA (MANY BACTERIA)
UNMETHYLATED CpG IN DNA
DOUBLE-STRANDED RNA (VIRAL)
TOLL-LIKE RECEPTORS
single stranded
RNA
TLR8
Akira et al. Nature Rev Immun 2004
TLR SPECIFICITY -2004
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TLR1
– Triacyl lipopeptides (Bacteria, mycobacteria)
– Soluble factors (Neisseria meningitidis)
TLR2
– Lipoprotein/lipopeptides (Various pathogens)
– Peptidoglycan (Gram-positive bacteria)
– Lipoteichoic acid (Gram-positive bacteria)
– Lipoarabinomannan (Mycobacteria)
– Glycolipids (Treponema maltophilum)
– Porins (Neisseria)
– Zymosan (Fungi)
TLR3
– Double-stranded RNA (Viruses)
TLR4
– Lipopolysaccharide (Gram-negative bacteria)
– Taxol (Plants)
– Fusion protein (Respiratory syncytial virus)
– Envelope protein (Mouse mammary-tumor
virus)
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TLR5
– Flagellin (Bacteria)
TLR6
– Diacyl lipopeptides (Mycoplasma)
– Lipoteichoic acid (Gram-positive
bacteria)
– Zymosan (Fungi)
TLR7
– Imidazoquinoline (Synthetic)
– Single-stranded RNA (Viruses)
TLR8
– Imidazoquinoline (Synthetic)
– Single-stranded RNA (Viruses)
TLR9
– CpG-containing DNA (Bacteria and
viruses)
Regulation of Endothelial Cell gene
expression by TLR4 activation
1. P-Selectin and E-Selectin
2. MCP-1, IL-8, Platelet activating factor
3. ICAM-1, VCAM-1
4. Many cytokines
NFkB important in this activation
*5. Macrophages activation of TLR4 increases
TNF and IL-1 that activate endothelial cells.
Activators of the endothelium to
induce leukocyte EC interactions
1. Bacteria
2. Toxins – Hydrogen Peroxide
3. Autoantibodies – Rheumatoid Arthritis
4. Oxidized Phospholipids - Atherosclerosis
5. Advanced glycosylation end products – Diabetes
6. Cytokines – TNF, IL-1
Activators of the endothelium to
induce leukocyte EC interactions
1. Bacteria
2. Toxins – Hydrogen Peroxide,Silica
3. Autoantibodies – FC receptor
4. Oxidized Phospholipids – CD 36
5. Advanced glycosylation end products – RAGE
6. Cytokines – TNF, IL1 - TNF and IL-1 receptors
Rolling Molecules
Selectins and their ligands
Integrins and their ligands
Endothelial molecule
Leukocyte Receptor
P-Selectin
Sialyl-Lewis X
PSGL-1
E-Selectin
Sialyl-Lewis X
GlyCam-1
L-Selectin
*VCAM-1
a5b1
a4b7
SELECTINS
• SELECTINS--LEUKOCYTE AND ENDOTHELIAL
PROTEINS THAT MEDIATE CELL ADHESION TO
CARBOHYDRATES
– P-SELECTIN, ON ENDOTHELIA AND PLATELETS, UPON
ACTIVATION RAPIDLY TRANSFERRED TO CELL
MEMBRANE
– L-SELECTIN, ON LEUKOCYTES, EXPRESSED
CONSTITUTIVELY, BECOME MORE ADHESIVE IN
STIMULATED LEUKOCYTES BUT SHED WITHIN MINUTES
OF ACTIVATION
– E-SELECTIN, ON ENDOTHELIA, INDUCED WITHIN
HOURS BY INFLAMMATORY MEDIATORS
Regulation of Chemokines Levels
and Activity
Attachment to GAGs- localizes activity
Cleavage by metalloproteinases
Formation of homo and heterodimers
m-RNA Degradation
Protein Degradation
Effects of chemotactic factors on
leukocyte activation
1. Chemotactic factors bind to GPCR
2. This leads to integrin activation
3. Integrin activation causes arrest followed
by spreading and migration
Structure and Function
of Integrins
Integrins are heterodimers (formed
by a and bchains) receptors
located on the surface of cells. The
b subunit contains the RGDbinding site. Calcium is required
for integrin binding activities. The
cytoplasmic end is associated with
cytoskeletal proteins.
FAK
Leukocyte transmigration
1. Leukocyte migrates to EC junction
2. Leukocyte activates transient endothelial
retraction
3. Leukocyte extends pseudopod into
junctions
4. Leukocyte binds to homotypic molecules
on EC and moves across monolayer
5. Leukocyte produces proteases and migrates
across basement membrane in response to
chemotactic factors
Activation of the Leukocyte for
killing and phagocytosis
PHAGOCYTOSIS AND
KILLING
PHAGOCYTIC KILLING:
-BY GRANULE CONTENTS
-BY REACTIVE OXYGEN PRODUCTS
EXTRACELLULAR
KILLING
NEUTROPHILS: ARSENAL
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REACTIVE OXYGEN PRODUCTS
NUTRIENT-BINDING PROTEINS
PROTEASES
LYSOZYME
MICROBICIDAL PEPTIDES
–DEFENSINS
–CATHELICIDINS
Genetic Disease
Defect
Leukocyte adhesion deficiency 1
b-chain of CD11/CD18
Leukocyte adhesion deficiency 2
Fucosyl transferase required for
sialylated oligosaccharide synthesis
Chronic granulomatous disease
X-linked
Autosomal recessive
Decreased oxidative burst
NADPH oxidase (membrane)
NADPH oxidase (cytoplasm)
Myeloperoxidase deficiency
Absent MPO-H2O2 system
Chediak-Higashi syndrome
Protein involved in organelle
membrane docking and fusion
Diseases Associated with
Inflammation and Angiogenesis
Cancer
Rheumatoid Arthritis
Atherosclerosis
Diabetic Retinopathy
Sources of Inflammatory
Molecules in Tumors
Extrinsic Pathway – Inflammation due to
infection of cell injury
Intrinsic Pathway – Oncogene Activation
Both lead to increased macrophages in tumors
Macrophage Phenotypes and
Cancer
M1
M2
Markers:
IL12, TNF, IL6,ROS
IL-4, IL-10, TGFB
Functions:
Attract lymphocytes
Decrease lymphocyte entry
Activate lymphocytes
Decrease in lymphocyte
activation
Kill tumor cells
Increase in angiogenesis
Increase tumor cell growth