A. Characteristics
- Duration of inflammation minutes to a few days
- Exudation of fluid and plasma proteins
- Emigration of leukocytes, predominantly neutrophils
- Response to many types of injury, including trauma, heat, cold, radiation, chemicals
- Clinical signs are: heat (calor), swelling, redness (rubor), pain (dolor)
- Lymphadenopathy (swollen glands) in draining lymph node areas may occur
- Elevated levels of CRP and Interleukin 6 (IL6) associated with increased mortality risk [4]
B. Physiology
- Hemodynamic changes with vasodilation
- Increased vascular permeability, especially capillaries, arterioles
- Antibodies and/or immune complexes can play a major role in initiation of inflammation
- Exudation of leukocytes and production of antimicrobial agents
- Neutrophils are major effector cells [12]
- Production of highly reactive oxygen species (ROS) which have antimicrobial activity
- Includes superoxide (O2-), hydrogen peroxide (H2O2), other peroxides
- Toxic halogenated and nitrogenated compounds are produced
- Result is excellent antimicrobial activity, but with collateral tissue damage
- ROS also play a role in reperfusion injury
- IgE plays major role in many cases of hypersensitivity
C. Hemodynamic Changes in Inflammation
- Arteriolar microvascular dilatation
- Venular constriction
- Responses due to vasoactive compounds:
- Histamine - potent vasodilator
- Arachidonic Acid Metabolites: prostaglandins (PGs), thromboxanes, leukotrienes (LTs)
- Nitric Oxide - vasodilation
- Prostaglandins (PG) [9]
- Various classes: vasodilators and vasoconstrictors, pro- and anti-inflammatory
- Vascular dilatation primarily mediated by prostacyclin (PGI-2) and PGE2
- Vasoconstriction mediated by other prostaglandins
- Prostaglandin E2 (PGE2) is pro-inflammatory
- PGD2 and PGF2a are anti-inflammatory
- PGs are produced from arachidonic acid by several cyclooxygenases
- Cyclooxygenases also called PG synthetases
- At least two, and possibly three, PG synthetase isoenzymes exist
- Nitric Oxide (NO) [3]
- Potent relaxer of smooth muscle including blood vessel, esophageal, bronchial
- Production of NO in brain inhibits alpha adrenergic tone and sympathetic outflow
- Inhibits platelet aggregation
- Inhibits smooth muscle proliferation
- Inflammatory mediator (particularly in combination with reactive oxygen species, ROS)
- In this regard, plays a role in host defense against microbes
- Can react rapidly with free radicals, certain amino acids, transition metal ions
- NO can also react with with ROS such as H2O2, superoxide
- ROS reaction with NO can lead to formation of peroxynitrite (-NO2) which is toxic
- In vascular space, NO is sacenvged by oxyhemoglobin forming methemoglobin and nitrate
- Most absorbed NO is converted to nitrate and excreted in urine
- Leukotrienes (LT) [2]
[Figure] "Leukotriene Synthesis"
- LTs are primarily responsible for vasoconstriction and leukocyte exudation
- They comprise the "slow reacting substance of anaphylaxis" or SRSA
- SRSA is a combination of LT-C4, -D4 and -E4
- Thromboxanes (TBX) are potent platelet activators and vasoconstrictors
- Chemokines (see below)
- Released by endothelium and fibroblasts
- Potently leukocytes attractants
D. Vascular Permeability
- Small and medium vessels, especially capillaries
- Endothelial (pericyte) contraction allows "pore" formation in capillaries
- Mediated by
- Serotonin
- Histamine
- Bradykinin (BK)
- Potentiated by vasodilators such as PGs and nitric oxide (see above)
- Antibodies and/or immune complexes can also play a central role
- Role of Kinins
- Peptide mediators of acute and subacute inflammation
- Generated by action of kallikreins (tissue and serum forms) - "contact-system"
- Kallikreins act on high- and low-molecular weight kininogens
- BK and kallidin (lysyl-BK) are produced
- Cause vasodilation, vascular leak, pain and neurotransmitter release
- Leads to local edema, blood pressure reduction, and pain
- BK is degraded by angiotensin converting enzyme (ACE)
- BK may be protective against development of left ventricular hypertrophy
- Kinins also enhance cytokine, eicosanoid and nitric oxide (NO) release
- Kinin type 1 receptors (B1) are inducible by inflammatory mediators such as IL1ß
- Kinin type 2 recpetors (B2) are constitutively expressed
E. Leukocyte Exudation [10]
- Leukocytes are part of the innate immune system with rapid responses to invaders [11]
- The innate immune system is designed to react to a few, highly conserved, antigens
- These antigens include lipopolysaccharide (LPS), teichoic acids, bacterial DNA, manins, and glucans
- Various specific receptors have evolved to recognize these antigenic structures
- Neutrophils and macrophages express many of these receptors
- Activation of these cell types through these receptors leads to expression of costimulatory surface molecules, cytokine, and chemokine production
- Some of the receptors are secreted and can stimulate the complement (C') system
- Innate Immune System [11,16]
- Mannan-binding lectin - initiates C' cascade
- Macrophage mannose receptor
- CD14 - one of the receptors for LPS, found on macrophages and B cells
- Toll-like receptors (TLR) - various family members; TLR-4 involved in LPS binding
- Activation of innate immunity leads to recruitment of adaptive immunity
- Formation of Ab-Ag complexes is main way of activating classical C' pathway
- Ag coupled to C' protein C3dg reduces threshold for B cell activation up to 10,000X
- Leukocytes traverse vessel walls between endothelial cells in response to stimuli
- Stimuli include leukotrienes (LTB4) and complement components (C5a > C3a >> C4a)
- These may affect expression of cell adhesion molecules (CAMs)
- Binding of CAMs to receptors is usually required for migration of effector cells
- Inhibition of complement component formation can substantially block inflammation
- Pexelizumab, a monoclonal antibody which binds C5 complement, showed no benefit in percutaneous coronary interventions [26]
- Orderly Recruitment of White Blood Cells
- Neutrophils (PMNs) enter first, then monocytes, then lymphocytes
- Probable role of Cell Adhesion Molecules including LFA1, MAC1, VCAM-1, others
- May also related to chemotactic factor production at specific site
- These factors include potent chemotactic cytokines called chemokines
- Leukocyte Migration [19]
- Leukocytes migrate out of blood into tissue in particular fashions
- Initially, adhesion of cells to blood vessel wall (endothelium) is required
- Adhesion cascade includes: tethering, rolling, activation, firm adhesion, transmigration
- Tethering requires selectins; these are proteins which bind to sugars (special lectins)
- L-Selectin (CD62L) by leukocytes, P-Selectin by platelets, E-Selectin by endothelium
- Rolling uses Integrins: Mac1 binds ICAM-1 (CD43), LFA-1 binds ICAM-1, and -2, and VLA4 binds to VCAM-1
- Macrophages use Mac1, lymphocytes use LFA-1 and VLA4, eosinophils use VLA4
- Transmigration across endothelium uses CD31 and other molecules
- Chemokines
- Two main types, alpha (CXC) and beta (CC) families
- Families based on cysteine (C) residues
- alpha chemokines (including IL-8) act mainly on neutrophils
- ß-chemokines on other cells
- Roles of PMNs in Inflammation
- Phagocytosis of bacteria, opsonized organisms
- Includes killing of organisms with free radicals (superoxide) and halides (see below)
- Tissue damage: oxidants, free radicals, proteases
- Induction of distant effects: PMNs and monocytes make highly inflammatory IL1
F. Vasoactive Inflammatory Mediators
- Histamine
- Preformed vasoactive (arteriolar dilation) amines, mainly from mast cells
- Release stimulated by IgE-R crosslinking and/or C3a/C5a binding to cells (see below)
- Serotonin
- Causes vasoconstriction is most vessels
- Increases platelet aggregation (synergizes with collagen)
- Direct constrictor effects on vascular smooth muscle
- Preformed vasoactive (arteriolar dilation) amines, mainly from mast cells
- Release stimulated by IgE-R crosslinking and/or C3a/C5a binding to cells
- Bradykinin
- Opposes effects of angiotensin II by inducing arteriolar vasodilatation and increasing vascular permeability
- Kininogen converted to bradykinin (9-mer) through clotting cascade
- Destroyed by ACE (Angiotensin converting enzyme; also called kininase)
- Other effects include slowing of heart rate and smooth muscle contraction
- Prostaglandins (PG)
- Cyclooxygenase pathway (arachidonic acid), inhibited by NSAIDS, glucocorticoids
- Prostacyclin: PG-I2 causes arteriolar dilatation along with PGE2
- Thromboxanes: extremely potent, mediate vascular constrict and platelet aggregation
- Leukotrienes (LT)
[Figure] "Leukotriene Synthesis"
- Lipoxygenase pathway of arachidonate metabolism through 5-lipoxygenase (5-LO)
- Synthesis blocked by glucocorticoids and 5-LO inhibitors
- LT receptor blockers are also available
- LTs include slow reacting substance of anaphylaxis (SRSA): LTE4
- LTE4 causes vasoconstriction, bronchoconstriction, increased vascular permeability
- Acetylated glycerol ether phosphocholine (AGEPC)
- Originally called platelet activating factor (PAF)
- PAF antagonists are being studied in sepsis and other inflammatory diseases
G. Complement (C') [5]
[Figure] "Complement Cascade"
- The C' system consists of classical and alternative pathways
- The classical pathway depends on antibodies (Abs) for activation
- Abs may exist bound in solid phase to targets, or in circulating immune complexes (IC)
- The classical pathway may be activated by either, provided the correct Fc is present
- The alternative pathway can be activated directly by foreign cell membranes
- Activation of the C' system by either pathway leads to C3a and C5a formation
- C3a and C5a are potent activators of mast cells
- C5a stimulates neutrophils as well
- Inhibitors of C' Activation [22]
- C1 Inhibitor - covalently binds C1r and C1s and blocks further activity
- C4bp - accelerates decay of classical pathway C3 convertase (C4b2a)
- Factor H (HF1) - accelerates decay of alternative pathway C3 convertase (C3bBb)
- Factor I - proteolytically cleaves and inactivates C4b and C3b (cofactors needed)
- Carboxypeptidase N - removes terminal arginine residues from C3a, C5a (inactivates)
- Vitronectin (S Protein) - binds C5b-7 complex and prevents membrane insertion
- SP-40 - modulates membrane attack complex formation
- C' Receptor 1 (CR1, CD35) - dissociation of all C3 convertases, cofactor
- Membrane Cofactor Protein - cofactor for Factor I mediated C3b and C4b cleavage
- Decay Accelerating Factor (DAF) - accelerates decay of all C3 convertases
- CD59 - inhibits lysis of bystander cells (C7 and C8 interactions)
- Homologous Restriction Factor (HRF) - inhibits bystander lysis, C8 and C9 interactions
- Disease and C' Inhibitors
- Absence of C1 inhibitor leads to hereditary angioedema
- Mutations in Factor H associated with familial hemolytic uremic syndrome (HUS) [22]
- Mutations of membrane cofactor protein also associated with familial HUS [22]
- Mutations in DAF or CD59 associated with Paroxysmal Nocturnal Hemoglobinuria (PNH)
- Eculizumab is a recombinant antibody against complement protein C5
- Blocks terminal complement activation by inhibiting C5 cleavage to C5a and C5b
- Reduces intravascular hemolysis, hemoglobinuria, need for transfusion in PNH [24]
- Pexelizumab, another Ab against C5, of no benefit in PCI (see above) [26]
- Blockade of C' activation with soluble CR1 (sCD35) reduces tissue damage
H. Phagocytes and Microbial Oxidants [12,14]
- Neutrophils and other phagocytes produce a variety of toxic oxidants
- These molecules are called reactive oxygen species (ROS)
- ROS include superoxide (O2-), hydrogen peroxide (H2O2), free radicals (OH·)
- These ROS then form halide or nitrate derivatives
- Hypochlorite (OCl-) and peroxynitrite (ONOO-) are highly toxic to microorganisms
- However, these compounds also harm normal tissue, causing "collateral damage"
- ROS are generated by four major enzymes in phagocytes
- NADPH Oxidase
- Superoxide Dismutase (SOD)
- Nitric Oxide Synthetase (NOS)
- Myeloperoxidase
- Xanthine oxidase may play a major role in liver and intestine (but not heart)
- Many other phagocyte oxidants are generated by noneyzmatic reactsion involving these ROS
- NADPH Oxidase
- Membrane-bound enzyme that catalyzes production of superoxide (O2-)
- 2 O2 + NADPH --> 2 O2- + NADPH+ + H+
- NADPH oxidase is dormant in resting phagocytes
- NADPH oxidase is activated by a variety of inflammatory and bacterial stimuli
- O2- is delivered to the external environment and into phagocytic vesicles
- Chronic granulomatous disease (CGD) is due to defects in NADPH oxidase
- Nitric oxide inhibits the function of NADPH oxidase [13]
- Superoxide Dismutase (SOD)
- Two forms of SOD: coper-zinc (cytoplasmic, SOD1) and manganese (mitochondrial)
- Superoxide O2- is highly reactive and toxic
- SOD catalyzes superoxide conversion to H2O2: 2 O2- + 2H+ --> O2 + H2O2
- H2O2 is less toxic than than O2- but still highly reactive
- H2O2 is normally detoxified by glutathione peroxidase (GPO) in higher organisms
- Catalase can also detoxify H202, forming H20 and O2
- Glutathione Peroxidase (GPO)
- Catalyzes reduction of H2O2 to H20 and oxidation of glutationine
- Also prevents oxidation of lipids to maintain biological membranes
- Converts oxidized lipids (which are atherogenic) back to non-oxidized lipids
- Strong inverse relationship between a patient's red blood cell GPO levels and risk of subsequent cardiovascular events [23]
- Myeloperoxidase (MPO)
- Heme enzyme which catalyzes oxidation of Cl-, Br-, I- (or SCN-) by hydrogen peroxide:
- Cl- + H2O2 --> OCl- + H2O
- OCl- is antimicrobial (but redundant systems are present)
- MPO deficiency is common (~1/1000 persons), usually with mild or no symptoms
- MPO is an abundant leukocyte enzyme elevated in fissured atherosclerotic plaques [21]
- Plasma levels evaluated as predictor of any cardiovascular event in patients with angina
- MPO levels at presentation also predicted risk of major cardiac events at 1 and 6 months
- Risk ratios in the 2-4X range, even after correction for baseline troponin levels [21]
- Major Nonenzymatic Reactions and ROS Products
- Hydroxyl Radical (OH·)
- Oxygenated Halogens (OCl-, others)
- Singlet Oxygen (1O2)
- Reactive Nitrogen Species
- Hydroxyl Radical (OH·)
- Generated by "Fenton Reaction": H2O2 + Fe2+ (Cu+) --> OH· + OH- + Fe3+ (Cu2+)
- OH· can initiate a variety of free radical reactions involving biomolecules
- OH· is therefore very destructive to biological systems
- Fe3+ is reduced back to Fe2+ by ascorbic acid (vitamin C) and other antioxidants
- Oxygenated Halogens
- Initially, OCl- / HOCl are formed by MPO (major oxygenated halogen in body)
- HOCl (hypochlorous acid) reacts with many amines to form chloramines
- Lipophilic chloramines are highly toxic: chloramine (NH2Cl) and putrescine (H2N-C4H8NHCl)
- Taurine chloramine (SO3=CH2-CH2-NHCl) is water soluble and nontoxic
- HOCl can also react with amino acids to form chloramines and aldehydes
- Aldehydes are highly reactive and toxic (used as fixatives in histology)
- Oxygenated halogens are highly toxic to microbes, may be most important in neutrophils
- Singlet Oxygen (1O2)
- Oxygen normally has two unpaired electrons (diradical)
- Singlet oxygen is much more reactive, in which these two electrons are paired
- Singlet oxygen is produced in excess in patients with certain types of porphyria
- Singlet oxygen is also produced by neutrophils: H2O2 + OCl- --> 1O2 + H2O + Cl-
- Likely that singlet oxygen is responsible for some antimicrobial effects and tissue damage
- Nitric Oxide Synthetase (NOS) [13]
- At least three forms of the enzyme exist: 2 constitutive forms and one inducible
- Constitutive NOS in brain (neuronal NOS or nNOS) and endothelial NOS (eNOS)
- Inducible NOS is associated primarily with inflammation (iNOS)
- Inducible form produced by phagocytes and other cells when stimulated
- All NOS catalyze production of nitric oxide (NO) from arginine
- Arginine + O2 + NADPH --> NO + citrulline + NADP+
- NOS is highly complex, containing FAD, FMN, heme, and biopterin
- Major role of NO in inflammation is conversion to peroxynitrite and other toxic metabolites
- Note that NO is a weak free radical
- Inhibition of iNOS can worsen tissue damage in some inflammatory settings [13]
- Therefore, the physiological role of iNOS in inflammation is complex
- Reactive Nitrogen Species
- Formed by combination of NO with superoxide (O2-) or other ROS
- A variety of species have been discovered including peroxynitrite (ONOO-)
- Peroxynitrite can combine with carbon dioxide, HOCl or other molecules
- Nitration of tyrosines in proteins leads to damage and dysregulation
- Free-Radical Scavenger NXY-059 [25]
- Reduces size of infarct in various animal stroke models and is neuroprotective
- Administration within 6 hours in acute stroke reduced disability at 90 days
- No effect on NIH Stroke Scale or Barthel index
I. Fever [4,20]
- Mediated by exogenous and endogenous pyrogens
- Central Nervous System (CNS) and Thermoregulation
- Temperature regulation is primarily at the CNS level
- Hypothalamus and limbic system outputs are key
- Preoptic region, including medial and lateral aspects of preoptic area, is critical
- The anterior hypothalamus and septum also play major roles
- Outputs from these areas travel down through reticular formation and spinal cord
- Thermoregulatory centers integrate inputs from skin thermosensors and core body areas
- A physiological thermal set point exists in the preoptic area
- This set point is developmentally determined and varies amongst individuals
- Normal set point is ~37°C (98.6°F)
- Major Exogenous Pyrogens
- Gram Negative Lipopolysaccharide (Endotoxin)
- Gram Positive Bacterial Molecules - lipoteichoic acids
- Viruses
- Fungi
- Protozoa
- Bacterial superantigens - toxic shock and other toxins
- Major Pyrogenic Cytokines (Endogenous Pyrogens)
- Interleukin 1 (IL1)
- Interleukin 6 (IL6)
- Tumor Necrosis Factor alpha (TNFa)
- Interferon Gamma (IFNg)
- Prostaglandin E2 is likely the common pathway at hypothalamic level
- IL6 is the major inducer of acute phase reactants
- TNFa and IL1ß levels predict relapse in Crohn's (inflammatory bowel) Disease [6]
- Fever Suppression [20]
- Acetaminophen, aspirin, non-steroidal antiinflammatory drugs (NSAIDs) are mainstay
- Primarily work by inhibition of prostaglanding E2 synthesis (mainly through COX2)
- Aspirin and NSAIDs have peripheral antiinflammatory activities also
- Acetaminophen only has central actions
J. Acute Phase Reactants (APR) [4,7]
- Cytokines stimulate fever as well as regulate the synthesis of a variety of proteins
- These proteins are called "acute phase" reactants or proteins (APR)
- There are both positively and negatively regulated ACR
- The erythrocyte sedimentation rate (ESR) is an indirect measure of APR
- IL6 is the major stimulator of APR
- IL1ß also plays a role
- CRP (C-reactive protein) is the most common clinical marker for inflammation
- IL6 directly stimulates CRP production, mainly from hepatocytes
- Elevated CRP is strongly associated with atherosclerosis [17] and type 2 diabetes [18]
- Elevated CRP and IL6 are associated with increased mortality in elderly [4]
- Chronic expressin of APR likely contributes to cachexia [15]
- The following APR levels increase with inflammation:
- Fibrinogen
- Plasminogen
- Tissue plasminogen activator (TPA)
- Plasminogen activator inhibitor 1 (PAI-1)
- Urokinase
- Protein S
- Vitronectin
- C-Reactive Protein (CRP)
- Serum Amyloid A
- Haptoglobin
- alpha1-Acid Glycoprotein
- Fibronectin
- Ceruloplasmin
- Haptoglobin (moreso than hemopexin)
- Ferritin
- Complement Proteins C3, C4 and C9
- Complement C1 Esterase Inhibitor
- Complement C4b Binding Protein
- Complement Factor B
- Mannose binding protein (lectin)
- alpha1-antichymotrpsin
- Pancreatic secretory trypsin inhibitor
- alpha1-Protease Inhibitor
- alpha2-Macroglobulin
- Phospholipase A2
- Pancreatic secretory trypsin inhibitor aa. Inter-alpha protease inhibitor
- The following ARP levels decrease with inflammation:
- Albumin
- Transthyretin (Prealbumin)
- Transferrin
- alpha2-HS Glycoprotein
- alpha-fetoprotein (AFP)
- Thyroxine binding globulin
- IGF-1
- Clotting Factor XII
- Role of APR [7]
- Replaces normal set of homeostatic proteins
- Resets equilibrium / homeostasis set point
- Likely net effect to improve defensive and adaptive capabilities
- Patterns of cytokine production and APR different with different stimuli
- Elevated IL-6 and C-reactive protein levels are associated with mortality in elderly [8]
K. Summary of Acute Inflammatory Response
- Vascular Leakage
- Histamine
- Kinins and substance P (pain)
- Prostaglandins and Leukotrienes
- Serotonin
- Neutrophil Migration
- Activation by C5a >> C3a
- IL-8 and other chemokines
- Eosinophilia may also occur
- Fever Induction
- Exogenous pyrogens stimulate PGs and various cytokines (endogenous pyrogens)
- Necrotic tissue stimulates endogenous pyrogens
- Endogenous pyrogens stimulate CNS preoptic center neurons
- CNS outputs stimulate heat production by visceral organs in the body
- Result is usually a fever
- Increased shivering or very high levels of pyrogens may lead to hypothermia
- Antibodies (if preformed or made locally) can potentiate these responses
- Acute Tissue damage due to effector cell lysosomal / oxidative products
- Induction of Acute Phase Reactants (ARP) as described above
Histopathologic Patterns in Inflammation
A. Serous- Mild injury, with epithelial destruction
- Examples: water blister, burns
- Primarily fluid accumulation (transudate)
B. Fibrinous
- Exudation of fluid rich in plasma proteins, containing fibrinogen
- Fibrinogen converted to fibrin by thrombin
- Cells are not involved in this process (humoral process)
- Fibrin forms and serves as scaffolding for growth of connective tissue
- Example: pericarditis
C. Pseudomembrane Formation
- Formation of new pseudomembrane on top of original one
- Similar to fibrous: covering of mucosal surface with fibrin, inflammatory cells, necrosis
- Often, only the very top of the original membrane is hurt, but function is compromised
- Examples
- Pseudomembranous colitis (C. difficele)
- Diphtheria
D. Suppurative (Purulent)
- Fluid is rich in neutrophils (dead) and target organism
- Frequent with gram+ infections, especially Staphylococcus
- Local collection of pus within tissue, organ etc, result is an abscess
- Abscess formation often by gram negative organisms: E. coli, Klebsiella, Bacteroides ssp
- Congenital Immunodeficiency Diseases
- Associated with defects in anti-microbial activity
- Chediak-Higashi Syndrome
- Chronic Granulomatous Disease
- Glucose 6-P Dehydrogenase (G6PD) Deficiency: Impaired H2O2 Production
- Chediak-Higashi Syndrome
- Large azurophilic granules not degranulated
- Poor neutrophil locomation
- Autosomal recessive gene
- Chronic Granulomatous Disease (CGD) [12]
- Defect in antimicrobial oxidants due to one of four distinct mutations
- Most common (65% of CGD) is defect in 65K protein on chromosome Xp21
- This is a mutation in one of the two subunits of cytochrome B (gp91 phox)
- The other 35% of CGD are missing either the other subunit of cytochrome B or one of the two regulatory subunits; all of these forms are autosomal
- Other Diseases with High Risk Suppurative Infections
- Hypogammaglobulinemia
- C3 and other complement deficiencies
- Sickle Cell Disease
- Sickle Cell Anemia
- Patients "auto-splenectomize" due to infarction of spleen
- Especially susceptible to encapsulated organisms
- Increased risk of Salmonella infections
E. Ulcer
- Shedding of inflamed tissue on surface of an organ
- Crater which remains is referred to as an ulcer
- Strict definition requires denuding of epithelia, causing an ulcer at the particular point
Resources
Absolute Neutrophil Count
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