(2nd semester B.Pharm Pathophysiology) inflammation notes

1. Inflammation and Healing
DEFINITION
Inflammation is defined as the local response of living mammalian tissues to injury from any agent. It
is a body defense reaction in order to eliminate or limit the spread of injurious agent, followed by
removal of the necrosed cells and tissues.
CAUSES
The injurious agents causing inflammation may be as under:
1. Infective agents like bacteria, viruses and their toxins, fungi, parasites.
2. Immunological agents like cell-mediated and antigen antibody reactions.
3. Physical agents like heat, cold, radiation, mechanical trauma.
4. Chemical agents like organic and inorganic poisons.
5. Inert materials such as foreign bodies.
 Inflammation is distinct from infection—inflammation is a protective response by the body
to variety of etiologic agents (infectious or non-infectious), while infection is invasion into
the body by harmful microbes and their resultant ill-effects by toxins.
 Inflammation involves 2 basic processes with some overlapping, viz. early inflammatory
response and later followed by healing.
 Though both these processes generally have protective role against injurious agents,
inflammation and healing may cause considerable harm to the body as well e.g. anaphylaxis
to bites by insects or reptiles, drugs, toxins, atherosclerosis, chronic rheumatoid arthritis,
fibrous bands and adhesions in intestinal obstruction.
SIGNS OF INFLAMMATION
i) Rubor (redness)
ii) Tumor (swelling)
iii) Calor (heat)
iv) Dolor (pain).
v) Functio laesa (loss of function)

2. TYPES OF INFLAMMATION
Depending upon the defense capacity of the host and duration of response, inflammation can be
classified as acute and chronic.
A) Acute inflammation
 It is of short duration (lasting less than 2 weeks) and represents the early body reaction,
resolves quickly and is usually followed by healing.
 The main features of acute inflammation are:
o accumulation of fluid and plasma at the affected site
o intravascular activation of platelets
o polymorphonuclear neutrophils as inflammatory cells.
B) Chronic inflammation
 It is of longer duration and occurs after delay, either after the causative agent of acute
inflammation persists for a long time, or the stimulus is such that it induces chronic
inflammation from the beginning.
 A variant, chronic active inflammation is the type of chronic inflammation in which
during the course of disease there are acute exacerbations of activity.
 The characteristic feature of chronic inflammation is presence of chronic inflammatory
cells such as lymphocytes, plasma cells and macrophages, granulation tissue formation,
and in specific situations as granulomatous inflammation.
C) Subacute inflammation is used for the state of inflammation between acute and chronic.
ACUTE INFLAMMATION

3. it can be divided into following two events:
I. Vascular events
II. Cellular events
I. VASCULAR EVENTS
 Alteration in the microvasculature (arterioles, capillaries and venules) is the earliest
response to tissue injury.
 These alterations include: haemodynamic changes and changes in vascular permeability.
Haemodynamic Changes
1. Irrespective of the type of cell injury, immediate vascular response is of transient vasoconstriction
of arterioles. With mild form of injury, the blood flow may be re-established in 3-5 seconds while
with more severe injury the vasoconstriction may last for about 5 minutes.
2. Next follows persistent progressive vasodilatation which involves mainly the arterioles. This
change is obvious within half an hour of injury. Vasodilatation results in increased blood volume in
microvascular bed of the area, which is responsible for redness and warmth at the site of acute
inflammation.
3. Progressive vasodilatation, in turn, may elevate the local hydrostatic pressure resulting in
transudation of fluid into the extracellular space. This is responsible for swelling at the local site of
acute inflammation.
4. Slowing or stasis of microcirculation follows which causes increased concentration of red cells,
and thus, raised blood viscosity.
5. Stasis or slowing is followed by leucocytic margination or peripheral orientation of leucocytes
(mainly neutrophils) along the vascular endothelium. The leucocytes stick to the vascular
endothelium briefly, and then move and migrate through the gaps between the endothelial cells into
the extravascular space. This process is known as emigration
Altered Vascular Permeability

4. Several mechanisms may contribute to increased vascular permeability in acute inflammatory
reactions.
 Endothelial cell contraction leading to intercellular gaps in postcapillary venules
o It is the most common cause of increased vascular permeability.
o It is a reversible process elicited by histamine, bradykinin, leukotrienes, and many
other chemical mediators.
o It is usually short-lived (15-30 minutes), and is called the immediate transient
response.
o A slower and more prolonged retraction of endothelial cells, resulting from changes
in the cytoskeleton, may be induced by cytokines
 Endothelial injury results in vascular leakage by causing endothelial cell necrosis and
detachment (immediate sustained response).
o Direct injury to endothelial cells is usually seen after severe injuries (e.g., burns and
some infections).
o In most cases leakage begins immediately after the injury and persists for several
hours (or days) until the damaged vessels are thrombosed or repaired. Venules,
capillaries, and arterioles can all be affected, depending on the site of the injury.
o Examples include mild to moderate thermal injury, certain bacterial toxins, and x- or
ultraviolet irradiation (i.e., the sunburn that appears the evening after a day in the
sun).

5.  Leukocyte-mediated endothelial injury may occur as a consequence of leukocyte
accumulation along the vessel wall. Aactivated leukocytes release many toxic mediators that
may cause endothelial injury or detachment.
 Increased transcytosis of proteins via an intracellular vesicular pathway augments venular
permeability, especially after exposure to certain mediators such as vascular endothelial
growth factor (VEGF). Transcytosis occurs via channels formed by fusion of intracellular
vesicles.
 Leakage from new blood vessels
o Tissue repair involves new blood vessel formation (angiogenesis). These vessel
sprouts remain leaky until proliferating endothelial cells mature sufficiently to form
intercellular junctions.
o New endothelial cells also have increased expression of receptors for vasoactive
mediators, and some of the factors that induce angiogenesis (e.g., VEGF) directly
induce increased vascular permeability via transcytosis.
II. CELLULAR EVENTS (Leukocyte Recruitment and Activation)
The cellular phase of inflammation consists of 2 processes:
1. exudation of leucocytes
2. phagocytosis.
1. Exudation of Leucocytes
The changes in blood flow and vascular permeability are quickly followed by an influx of leukocyte
into the tissue
The journey of leukocyte from the vessel lumen to the tissue is a multistep process that is mediated
and controlled by adhesion molecules and cytokines
The process can be divided into
i) In the lumen:
Margination, rolling and adhesion
ii) Migration across the endothelium and vessel wall
iii) Migration in the tissue towards chemotactic stimulus
i. Leukocyte adhesion to endothelium

6.  Vascular endothelium in its normal, inactivated state does not bind circulating cells
or impede their passage
 In normally flowing blood in venules, RBC are confined to a central axial colum,
displacing the leukocytes towards the wall of vessel
 Because of blood flow slows early in inflammation (stasis), haemodynamic condition
change, and more white blood cells assume a peripheral position along the
endothelial surface. This process of leukocyte redistribution is called Margination
 The peripherally marginated and pavemented neutrophils slowly roll over the
endothelial cell lining (Rolling phase)
 This is followed by transient bond between leukocytes and endothelial cells
becoming firmer (Adhesion)
 The cell adhesion molecules like selectins, integrins and immunoglobulin gene
superfamily adhesion molecules are responsible for rolling and adhesion phase
ii. Leukocyte migration through endothelium
 Migration of leukocytes through the endothelium is called transmigration or diapedesis
 It occurs mainly in post capillary venules
 The expression of cell adhesion molecule is enhanced by cytokines, thus ensuring the
leukocytes are recruited to the tissue
 Adhesion molecules present in the inter cellular junction between endothelial cells are
involved in the migration of leukocytes
 After traversing the endothelium, leukocytes pierse the basement membrane, by secreting
collaginases, and enter the extravascular tissue ( emigration)
 The cell then migrate towards the chemotactic gradient created by cytokines and
accumulate in the extravascular site
iii. Chemotaxis of leukocytes
 After exiting the circulation, leukocytes will move towards the site of injury by a process
called chemotaxis
 It is defined as the locomotion along a chemical gradient
 Both exogenous and endogenous substance can act as chemoatractants
 Exogenous agents: bacterial products including peptides that posses an N terminal amino
acid
 Endogenous agents: cytokines, Arachidonic acid metabolites
 Chemotactic agents bind to specific g-protein coupled receptors on the surface of leukocytes

7.  This leads to increased cytosolic calcium, which triggers the assembly of cytoskeletal
contractile elements necessary for movement. Leukocytes move by extending pseudopods
that anchor to the ECM and then pull the cell in the direction of the extension.
 The direction of such movement is specified by a higher density of receptor-chemotactic
ligand interactions at the leading edge of the cell.
 Leukocyte Activation
 Once leukocytes have been recruited to the site of infection or tissue necrosis, they must be
activated to perform their functions.
 Stimuli for activation include microbes, products of necrotic cells, and several mediators that
are described later.
 Leukocytes express on their surface different kinds of receptors that sense the presence of
microbes. These include Toll-like receptors, which recognize endotoxin (LPS) and many other
bacterial and viral products
 Engagement of these receptors by microbial products or by various mediators of
inflammation induces a number of responses in leukocytes that are part of their normal
defensive functions and are grouped under the generic term leukocyte activation
2. Phagocytosis.
 Recognition of microbe or dead cells induces several responses in leukocytes
 The functional responses that are most important for destruction of microbes and other
offenders are phagocytosis and intracellular killing
 Phagocytosis is defined as the process of engulfment of solid particulate by the cells (cell
eating)
 Types of phagocytic cells
o Polymorphonuclear neutrophils (PMNs) which appear early in acute inflammatory
response, sometimes called as microphages
o Circulating monocytes and fixed tissue mononuclear phagocytes, commonly called
as macrophages.
 Phagocytosis of the microbe by polymorphs and macrophages involves the following 3 steps
o 1. Recognition and attachment
o 2. Engulfment
o 3. Killing and degradation
1. Recognition and attachment
o Phagocytosis is initiated by the expression of cell surface receptors on macrophages
which recognise microorganisms: mannose receptor and scavenger receptor.

8. o Then the microorganisms are coated with specific proteins, opsonins, from the
serum and the process is called opsonisation (meaning preparing for eating).
o Opsonins establish a bond between bacteria and the cell membrane of phagocytic
cell. The main opsonins present in the serum are as under:
IgG opsonin
C3b opsonin
Lectins
2. Engulfment
o The opsonised particle or microbe bound to the surface of phagocyte is ready to be
engul fed.
o This is accomplished by formation of cytoplasmic pseudopods around the particle due to
activation of actin filaments beneath cell wall, enveloping it in a phagocytic
vacuole(phagosomes)
3. Killing and degradation
o The phagosomes then fuses with lysosome granules
o This stage resulting in discharge of the granules contents into the phagolysosome
o Resulting in the killing and degradation of microbes
In general, following mechanisms are involved in disposal of microorganisms:
A. Intracellular mechanisms:
i) Oxidative bactericidal mechanism by oxygen free radicals
a) MPO-dependent

9. b) MPO-independent
ii) Oxidative bactericidal mechanism by lysosomal granules
iii) Non-oxidative bactericidal mechanism
B. Extracellular mechanisms
A. INTRACELLULAR MECHANISMS
 Oxidative bactericidal mechanism by oxygen free radicals An important mechanism of
microbicidal killing is by oxidative damage by the production of reactive oxygen metabolites
(O’2, H2O2, OH’, HOCl, HOI, HOBr).
 A phase of increased oxygen consumption (‘respi ratory burst’) by activated phagocytic
leucocytes requires the essential presence of NADPH oxidase

 a) MPO-dependent killing. In this mechanism, the enzyme MPO acts on H2O2 in the
presence of halides (chloride, iodide or bromide) to form hypohalous acid (HOCl, HOI, HOBr).
It is more potent antibacterial system
in polymorphs than H2O2
alone:

 b) MPO-independent killing. Mature macrophages lack the enzyme MPO and they carry out
bactericidal activity by producing OH– ions and superoxide singlet oxygen (O’) from H2O2 in
the presence of O’2 (Haber-Weiss reaction) or in the presence of Fe++ (Fenton reaction):


10.  ii) Oxidative bactericidal mechanism by lysosomal granules
o The preformed granule-stored products of neutrophils and macrophages are
discharged or secreted into the phagosome and the extracellular environment.
o Substances liberated by degranulation of macrophages and neutrophils are MPO,
protease, trypsinase, phospholipase, and alkaline phosphatase.
o Progressive degranulation of neutrophils and macrophages along with oxygen free
radicals degrades proteins i.e. induces proteolysis.
iv) Non-oxidative bactericidal mechanism
o Some agents released from the granules of phagocytic cells do not require oxygen
for bactericidal activity.
o These include the following: a) Granules. Some of liberated lysosomal granules do
not kill by oxidative damage but cause lysis of microbe within phagosome.
o These are lysosomal hydrolases, permeability increasing factors, cationic proteins
(defensins), lipases, proteases, DNAases. b) Nitric oxide.
o Nitric oxide is a reactive free radicals similar to oxygen free radicals which is formed
by nitric oxide synthase.
o It is produced by endothelial cells as well as by activated macrophages. Nitric oxide
is another potent mechanism of microbial killing.
B. EXTRACELLULAR MECHANISMS
i) Granules : Degranulation of macrophages and neutrophils exert its effects of proteolysis outside
the cells as well.
ii) Immune mechanisms : Immune-mediated lysis of microbes takes place outside the cells by
mechanisms of cytolysis, antibodymediated lysis and by cell-mediated cytotoxicity.
MEDIATORS OF INFLAMMATION
These are a large and increasing number of endogenous chemical substances which mediate
the process of acute inflammation. Mediators of inflammation have some common properties as
under:
 These mediators are released either from the cells or are derived from plasma proteins ”
 All mediators are released in response to certain stimuli. These stimuli may be a variety of
injurious agents, dead and damaged tissues, or even one mediator stimulating release of
another (secondary mediators)

11.  Mediators act on different targets. They may have similar action on different target cells or
differ in their action on different target cells.
 Ranges of actions of different mediators are: increased vascular permeability, vasodilatation,
chemotaxis, fever, pain and tissue damage.
 Mediators have short lifespan after their release. After release, they are rapidly removed
from the body by various mechanisms
I. CELL-DERIVED MEDIATORS
1. VASOACTIVE AMINES
o Two important pharmacologically active amines that have role in the early inflammatory
response are histamine, 5-hydroxytryptamine (5-HT) or serotonin and neuropeptides
o Histamine
 It is stored in the granules of mast cells, basophils and platelets.
 It is released from these cells by various agents as under:
 Stimuli or substances inducing acute inflammation e.g. heat, cold,
irradiation, trauma, irritant chemicals, immunologic reactions etc.
 Immune reactions
 Anaphylatoxin
 The main actions of histamine: vasodilatation, increased vascular (venular)
permeability, itching and pain.
 Stimulation of mast cells and basophils also releases products of arachidonic acid
metabolism
o ii) 5-Hydroxytryptamine (5-HT or serotonin)
 It is present in tissues like chromaffin cells of GIT, spleen, ner vous tissue, mast cells
and platelets.

12.  The actions of 5-HT are similar to histamine but it is a less potent mediator of
increased vascular permeability and vasodilatation than histamine.
o Neuropeptides
 Another class of vasoactive amines is tachykinin neuropeptides such as substance P,
neurokinin A, vasoactive intestinal polypeptide (VIP) and somatostatin.
 These small peptides are produced in the central and peripheral nervous systems.
 The major proinflammatory actions of these neuropeptides are as follows: a)
Increased vascular permeability. b) Transmission of pain stimuli. c) Mast cell
degranulation.
2. ARACHIDONIC ACID METABOLITES (EICO SANOIDS)
o These are the most potent mediators of inflammation, much more than oxygen free
radicals.
o Arachidonic acid is a constituent of the phospholipid cell membrane
o Arachidonic acid is released from the cell membrane by phospholipases.
o It is then activated to form arachidonic acid metabolites or eicosanoids by one of the
following 2 pathways: via cyclo-oxygenase pathway or via lipooxygenase pathway:
Metabolites via cyclo-oxygenase pathway:
a) Prostaglandins (PGD2, PGE2 and PGF2-a).

13.  PGD2 and PGE2 act on blood vessels and cause increased venular permeability,
vasodilatation and bronchodilatation and inhibit inflammatory cell function. PGF2-a
induces vasodilatation and bronchoconstriction.
b) Thromboxane A2 (TXA2). Platelets contain the enzyme thromboxane synthetase and
hence the metabolite, thromboxane A2, formed is active in platelet aggregation, besides
its role as a vasoconstrictor and broncho-constrictor.
c) Prostacyclin (PGI2). PGI2 induces vasodilatation, broncho dilatation and inhibits
platelet aggregation.
d) Resolvins are another derivative of COX pathway which act by inhibiting production of
pro-inflammatory cytokines.
Metabolites via lipo-oxygenase pathway:
a) 5-HETE (hydroxy compound), an intermediate product, is a potent chemotactic
agent for neutrophils
b) b) Leukotrienes (LT) are so named as they were first isolated from leucocytes. Firstly,
unstable leukotriene A4 (LTA4) is formed which is acted upon by enzymes to form
LTB4 (chemotactic for phagocytic cells and stimulates phagocytic cell adherence)
while LTC4, LTD4 and LTE4 have common actions by causing smooth muscle
contraction and thereby induce vasoconstriction, bronchoconstriction and increased
vascular permeability
3. LYSOSOMAL COMPONENTS
 The inflammatory cells— neutrophils and monocytes, contain lysosomal granules
which on release elaborate a variety of mediators of inflammation. These are as
under:
o i) Granules of neutrophils Neutrophils have 3 types of granules: primary or
azurophil, secondary or speci fic, and tertiary.
 a) Primary or azurophil granules are large azurophil granules which
contain functionally active enzymes. These are myeloperoxidase,
acid hydrolases, acid phosphatase, lysozyme, etc
 b) Secondary or specific granules contain alkaline phosphatase,
lactoferrin, gelatinase, collagenase, lysozyme, vitamin-B12 binding
proteins, plasminogen activator.

14.  c) Tertiary granules or C particles contain gelatinase and acid
hydrolases.
o ii) Granules of monocytes and tissue macrophages These cells on
degranulation also release mediators of inflammation like acid
proteases, collagenase, elastase and plasminogen activator.
4. PLATELET ACTIVATING FACTOR (PAF)
o It is released from IgE-sensitised basophils or mast cells, other leucocytes,
endothelium and platelets.
o Apart from its action on platelet aggregation and release reaction, the actions of
PAF as mediator of inflammation are:
i. increased vascular permeability;
ii. vasodilatation in low concentration and vaso cons triction otherwise;
iii. bronchoconstriction;
iv. adhesion of leucocytes to endothelium;
v. chemotaxis.
5. CYTOKINES
i) Cytokines are polypeptide substances produced by activated lymphocytes (lymphokines)
and activated monocytes (monokines).

15. 6. FREE RADICALS: OXYGEN METABOLITES AND NITRIC OXIDE
o i) Oxygen-derived metabolites are released from activated neutrophils and macrophages
and include superoxide oxygen (O’2), H2O2, OH’ and toxic NO products.
These oxygen-derived free radicals have the following actions in inflammation:
o a) Endothelial cell damage and thereby increased vascular permeability.
o b) Activation of protease and inactivation of anti pro tease causing tissue matrix
damage.
o c) Damage to other cells.
The actions of free radicals are counteracted by antioxidants present in tissues and serum which play
a protective role
o ii) Nitric oxide (NO) was originally described as vascular relaxation factor produced by
endothelial cells. Now it is known that NO is formed by activated macrophages during the
oxidation of arginine by the action of enzyme, NO synthase. NO plays the following roles in
mediating inflammation: a) Vasodilatation b) Anti-platelet activating agent c) Possibly
microbicidal action.
II. PLASMA PROTEIN-DERIVED MEDIATORS (PLASMA PROTEASES)

16. o These include various products derived from activa tion and interaction of 4 interlinked
systems: kinin, clotting, fibrinolytic and complement.
o Each of these systems has its inhibitors and accelerators in plasma with negative and
positive feedback mechanisms respectively.
o 1. THE KININ SYSTEM
o This system on activation by factor Xlla generates bradykinin, so named because of the slow
contraction of smooth muscle induced by it.
o First, kallikrein is formed from plasma prekallikrein by the action of prekallikrein activator
which is a fragment of factor Xlla. Kallikrein then acts on high molecular weight kininogen to
form bradykinin
o Bradykinin acts in the early stage of inflammation and its effects include: i) smooth muscle
contraction; ii) vasodilatation; iii) increased vascular permeability; and iv) pain.
o
o 2. THE CLOTTING SYSTEM
o Factor Xlla initiates the cascade of the clotting system resulting in formation of fibrinogen
which is acted upon by thrombin to form fibrin and fibrinopeptides
o The actions of fibrinopeptides in inflammation are: i) increased vascular permeability; ii)
chemotaxis for leucocyte; and iii) anticoagulant activity.
o 3. THE FIBRINOLYTIC SYSTEM
o This system is activated by plasminogen activator, the sources of which include kallikrein of
the kinin system, endothelial cells and leucocytes.
o Plasminogen activator acts on plasmino gen present as component of plasma proteins to
form plasmin. Further breakdown of fibrin by plasmin forms fibrino peptides or fibrin split
products