University of Colorado Denver – School of Medicine
Disease and Defense Course IDTP5004A
Instructor: Francisco G. La Rosa, MD
February 9th, 2009
CHEMICAL MEDIATORS OF INFLAMMATION
Recommended Reading: Chapter 2 - "Acute and Chronic Inflammation", in Kumar, Robbins and
Cotran: Pathologic Basis of Disease, 7th ed. WB Saunders Co. This is also available free online at:
http://www.mdconsult.com/das/book/body/94286448-2/0/1249/0.html
Learning Objectives:
1.
List the major inflammatory effects of bacterial endotoxin.
2.
List the major plasma-derived inflammatory mediators from the following systems and
describe their inflammatory effects:
a.
b.
c.
d.
Clotting system
Fibrinolytic system
Kinin system
Complement system
3.
List the major lipid mediators and their role in inflammation.
4.
Describe the pathways (including the enzymes involved) for the production of the
inflammatory mediators in objectives #2 and #3.
5.
Discuss cytokines and their effects.
6.
Describe the sources and effects of vasoactive amines.
7.
Describe the sources, synthesis and effects of nitric oxide.
8.
Discuss the biochemical events involved in stimulus response coupling during neutrophil
activation.
9.
Describe the neutrophil-derived inflammatory mediators and their effects.
10.
How is the oxidative burst reaction of neutrophils important in inflammation?
11.
How is neutrophil degranulation important in inflammation?
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“Chemical Mediators of Inflammation”
Page 2
CHEMICAL MEDIATORS OF INFLAMMATION
“Chemical substances mediate all phases of acute inflammation” (Thomas Lewis)
1. Reactions are similar irrespective of tissue or type of injury
2. Reactions occur in absence of nervous connections
Mechanism of action:
1. Most mediators act through cell surface receptors. Exceptions are those that have a direct
enzymatic effect (lysosomal enzymes) or direct toxic effect (reactive oxygen species).
2. Stimulus response coupling leads to biological response.
3. Mediators can stimulate target cells to release secondary mediators.
4. Mediators are short lived. They are rapidly inactivated (helps prevent harmful effects).
Classification of chemical mediators by source:
1. Exogenous – bacterial products
a. Bacterial Endotoxin (lipopolysaccharide)
 Stimulates neutrophils and macrophages. Triggers production of cytokines (TNF, IL-1)
 Increases neutrophil adhesion to endothelium
 Activates Hageman factor and complement system
b. Bacterial Peptides (containing N-formyl methionine) – chemotactic factors
2. Endogenous – produced internally by host
a. Plasma derived. Proteins circulate in plasma as inactive precursors and undergo
proteolytic cleavage to become active.
b. Cell derived. Cell stimulation can result in secretion of preformed mediators and/or
synthesis and secretion of new mediators.
PLASMA-DERIVED MEDIATORS – Series of inactive proteins circulating in the plasma that are
converted to proteolytic enzymes that activate other proteins.
Plasma Proteases:
Interrelated systems that are triggered by activation of Hageman factor (Factor XII of the
coagulation cascade). Activated by endotoxin, activated platelets or contact of plasma with
damaged tissue (collagen, basement membrane).
a. Clotting System – results in production of thrombin, factor Xa and formation of
fibrinopeptides
1. thrombin cleaves fibrinogen to form fibrin and enhances leukocyte adhesion
2. fibrinopeptides are chemotactic and increase vascular permeability
3. factor Xa increases vascular permeability and leukocyte emigration
b. Fibrinolytic System – results in formation of the proteolytic enzyme, plasmin.
Plasmin cleaves fibrin to form fibrin degradation products (increase vascular
permeability) and complement fragments (anaphylatoxins).
c. Kinin System – results in bradykinin formation (nonapeptide)
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Bradykinin is a vasodilator, increases vascular permeability, bronchial smooth muscle
contraction, pain. It is short-lived (inactivated by kininases).
d. Complement System – series of plasma proteins (C1-C9) that play a role in
inflammation and immune defense against microorganisms.
Important inflammatory mediators in complement system:
C3a, C5a – Increase vascular permeability and cause vasodilation by binding to mast cells
and inducing histamine release (anaphylatoxins).
C5a – Chemotactic for neutrophils, monocytes, eosinophils, and basophils. Increases
adhesiveness of neutrophils to endothelium. Stimulates synthesis and secretion of arachidonic
acid metabolites.
C3b – Opsonin, binds to microbial surface and promotes phagocytosis.
CELL-DERIVED MEDIATORS
1.
Lipid Mediators – arachidonic acid metabolites (prostaglandins, thromboxane,
leukotrienes, lipoxins), platelet-activating factor.
a. Produced by stimulated neutrophils, macrophages, platelets, mast cells, endothelial cells
b. Autacoids – are biological factors that are primarily characterized by the effect they have
upon smooth muscle. With respect to vascular smooth muscle, there are both
vasoconstrictor and vasodilator autacoids. They act locally, rapidly destroyed.
Arachidonic Acid Metabolites (eicosanoids) – Arachidonic acid is a 20 carbon fatty
acid with 4 double bonds. Oxygenated metabolites of arachidonic acid are found in
inflammatory lesions and participate in all phases of inflammation. They also
regulate normal physiological processes such as hemostasis. Their production is
inhibited by important anti-inflammatory drugs (NSAIDs, steroids).
Important Mediators – they act through G-protein coupled receptors.
TXA2 (platelets) – vasoconstrictor, platelet aggregation
PGI2 (endothelial cells) – vasodilator, inhibits platelet aggregation
PGD2 (mast cells), PGE2/PGF2 (widely distributed) – vasodilators, potentiate edema
LTC4, D4, E4 – vasoconstrictors, bronchoconstrictors, increase vascular permeability
LTB4 – chemotactic
Lipoxins – vasodilators, chemotactic, increase monocyte adhesion, decrease neutrophil
chemotaxis and adhesion, negative regulators of leukotriene action.
i.
ii. Platelet-activating Factor (PAF) – phospholipid mediator (acetyl glycerol ether
phosphocholine). Acts through G-protein coupled receptor, inactivated by
acetylhydrolase.
Sources – neutrophils, monocytes, basophils, endothelium, platelets, others.
Actions – platelet stimulation, vasoconstriction, bronchoconstriction, vasodilation,
increased vascular permeability, leukocyte activation (adhesion, chemotaxis,
degranulation, oxidative burst).
Production – derived from membrane phospholipids by action of phospholipase A2.
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2.
“Chemical Mediators of Inflammation”
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Cytokines – Regulatory polypeptides produced by many cell types (lymphocytes,
macrophages) and include IL-1 (interleukin 1), IL-2, TNF (tumor necrosis factor), interferongamma (IFN), chemokines (IL-8).
1. Autocrine effects – act on same cell that produces them.
2. Paracrine effects – act on cells in immediate vicinity. Increase adhesiveness of
endothelium.
3. Endocrine effects – act systemically. Induce acute phase reactions.
Types of cytokines based on action or target cell:
a. Cytokines that regulate lymphocyte function – IL-2 stimulates lymphocyte growth and
transforming growth factor  (TGF) inhibits lymphocyte growth.
b. Cytokines involved in innate immunity (primary response to injurious stimuli) – tumor
necrosis factor (TNF) and IL-1.
c. Cytokines that activate inflammatory cells (macrophages) – interferon  (IFN-) and IL12.
d. Chemokines – chemotactic for leukocytes. They bind to G-protein coupled receptors.
CXCR4 and CCR5 are co-receptors for HIV entry into lymphocytes.
CXC chemokines have one amino acid separating two cysteines. IL-8 is an activator
and chemoattractant for neutrophils that is produced by macrophages, endothelium and
fibroblasts exposed to IL-1 and TNF.
CC chemokines have adjacent cysteine residues (monocyte chemoattractant protein
(MCP-1) and macrophage inhibitory protein (MIP-1).
3.
Vasoactive Amines – Histamine, serotonin
a. Stored preformed in granules of mast cells, basophils and platelets.
b. Mast cells (connective tissue) and basophils (circulating) are induced to degranulate by
Ag/Ab (IgE) complexes binding to Fc receptors, physical injury (trauma, heat), C3a and
C5a complement fragments (anaphylatoxins), leukocyte-derived proteins,
neuropeptides, cytokines (IL-1, IL-8).
c. Induce arteriolar dilation and the immediate phase of increased vascular permeability.
Binds to receptors on endothelial cells, inducing contraction and opening of junctions.
d. Rapidly inactivated by histaminase.
e. Serotonin is found in platelet granules and is released when platelets aggregate
(collagen, thrombin, PAF, Ag/Ab complexes, ADP, TXA2).
4.
Neuropeptides – Small proteins (substance P) secreted by nerve fibers prominent in the
lung and GI tract, that transmit pain signals, regulate vessel tone, and modulate vascular
permeability.
5.
Nitric Oxide (NO) (free radical gas) – ubiquitously produced with diverse local effects.
a. It has a short half-life (sec), it acts locally and the amounts produced are controlled by
the rate of synthesis from L-arginine by the enzyme nitric oxide synthase (NOS).
b. There are three types of NOS isoforms that have different tissue distribution and
regulation.
i. Type I nNOS is constitutively expressed in neuronal tissue and is regulated by Ca2+.
In CNS, NO regulates neurotransmitter release and blood flow.
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ii. Type II iNOS is an inducible enzyme present in many cell types (endothelium,
smooth muscle cells, macrophages). It is calcium independent. It is upregulated by
cytokines (IL-1, TNF, IFN) and endotoxin. NO produced by macrophages is
cytotoxic for microbes and tumor cells.
iii. Type III eNOS is constitutively expressed by endothelial cells. It is regulated by Ca2+
and activated in endothelial cells by bradykinin, thrombin and shear stress. NO
produced by endothelial cells induces smooth muscle relaxation (vasodilation) by
increasing cGMP levels.
Roles of NO in inflammation
i. Relaxation of vascular smooth muscle
ii. Antagonism of platelet activation (adhesion, aggregation, degranulation)
iii. Reduction of leukocyte recruitment at inflammatory sites
iv. Action as a microbicidal agent in macrophages
6.
Neutrophil activation and mediators produced
a. Receptor-Ligand Binding – Neutrophils have receptors for many mediators (C5a, C3b,
LTB4, N-formylated peptides, endotoxin, immunoglobulin, platelet-activating factor,
cytokines (TNF, IL-1).
b. Stimulus-response Coupling – Intracellular signals trigger biological responses.
Phospholipase C is activated leading to PIP2 breakdown to form DAG and IP3.
c. Cell Activation
i. IP3 induces release of CA2+ from intracellular stores which is important for assembly
of cytoskeletal elements necessary for movement, activation of phospholipase A2
leading to production of arachidonic acid metabolites, and increased adhesiveness.
ii. DAG activates protein kinase C which is necessary for activation of the oxidative
burst and secretion/degranulation of lysosomal enzymes.
d. Oxidative Burst (increased oxygen consumption and glucose utilization).
i. Occurs in neutrophils (and monocytes/macrophages) activated by chemotactic
factors, immune complexes, and during phagocytosis.
ii. NADPH oxidase pathway is activated leading to production of oxygen-derived free
radicals. Superoxide anion is converted to H2O2, OH, and toxic NO derivatives.
iii. At low levels they can increase expression of chemokines, cytokines and adhesion
proteins, but at high levels they cause tissue injury (endothelial damage leading to
increased vascular permeability).
iv. toxic oxygen metabolites are rapidly converted to nontoxic metabolites by catalase,
superoxide dismutase, glutathione.
e. Degranulation – Preformed mediators are secreted from neutrophil (and monocyte)
granules.
Neutral proteases (collagenase, elastase, cathepsin) – degrade extracellular matrix components
resulting in tissue destruction. Can also cleave C3 and C5 to generate anaphylatoxins. Neutral
proteases are inactivated by anti-proteases such as 1-macroglobulin and 1-antitrypsin.
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Disclaimers:
1. The primary goal of this chapter is to study the learning objectives outlined at the beginning of
this handout. The material to study is provided in the lectures, the handouts and the
recommended textbooks. All these sources provide the content over which you will be tested.
The lectures are intended to provide broad information of the material found in the textbooks and
handouts, and to give the students the opportunity to ask questions on subjects not clear in the
texts. The handouts do not seek to follow up the sequence of the lectures, and most importantly,
they are not a surrogate of the books.
2. The text presented in this handout has been edited by Dr. La Rosa from material found in your
books, from published articles and other educational works. This handout is solely for
educational purpose and not intended for commercial or pecuniary benefit (see USA Copyright
Law, Section 110, “Limitations on exclusive rights: Exemption of certain performances and
displays”). Reproduction and use of this handout can be done only for educational use.
[Download] the USA Copyright Law version, October 2007.