Inflammation and Repair

INTRODUCTION
WAR IS THE METAPHOR FOR INFLAMMATION. Both are necessary evils. Both are
more-or-less stereotyped responses to outside threats. There are specialized troops (white
cells), including suicide-commandos (neutrophils), long-term siege armies (granulomas),
and many others. There are supply routes (vessels), communications and intelligence
(mediators), and a huge array of lethal weapons (inflammatory enzymes). In war as in
inflammation, there will be damage to both the enemy and to friendly forces, and there
will very likely be severe damage to the battlefield itself. Despite idealistic rhetoric about
"the laws of war", when the fighting starts, there is really only one law for the soldiers:
"Kill your enemy." Like it or not, if you want peace, you must be prepared to fight under
certain conditions. Like it or not, if you want to be healthy, your body must be able to
mount an inflammatory response. Force will always rule our world. Our best hope is that
this will be the force of good laws. And the best for which we can hope from the
inflammatory response is that, for most of our lives, it will do us rather good than harm.
Probably your own death will be caused by your last inflammatory response.

Definition: "Big Robbins" has defined INFLAMMATION as "the reaction of

vascularized living tissue to local injury". Inflammation is the name given to the more-or-
less stereotyped ways our tissues respond to noxious stimuli, with blood vessels and
white blood cells as its twin centerpieces and a host of proteins as actors. Inflammation
"destroys, dilutes, or walls off the injurious agent" and sets in motion the limited powers
of the body to heal itself. Inflammation and repair can and do themselves damage the
body.

Causes of inflammation:
Physical agents like Cold, Heat, UV rays
Chemical agents like poisons, Toxins of Bacteria
Infective agents like Bacteria, Virus, and Fungus
Immunological agents like Antigen Antibody reactions, Cell mediated.
Signs of inflammation – Celsius described 4 signs of inflammation
Rubor
Tumor
Dolor
Calor
Fifth sign – functiolaesa was later added by virchow

Types of Inflammation:
Depending upon the rapidity and duration of response
Acute inflammation – It is almost completely stereotyped -- over minutes to a few
days, blood vessels widen and longer duration ("late-phase inflammation) is more
variable, with variable participation by lymphocytes, plasma cells, macrophages, and
healing cells (fibroblasts and angioblasts).disgorge protein, and neutrophils leave the
bloodstream and rampage through surrounding tissues.

Chronic Inflammation - ("late-phase inflammation") is a response to prolonged

problems, orchestrated by T-helper lymphocytes. It may feature recruitment and
activation of T- and B-lymphocytes, macrophages, eosinophils, and/or fibroblasts.
Again, the process is complex. You will recognize lymphocytes in tissue section by
their small, "blue button" nuclei.

Depending on the morphologic types

 Adhesive Inflammation – in which amount of fibrin is sufficient to result in
slight or moderate degree of adherence
 Catarrhal Inflammation – hyperemia of mucosal vessels, enlargement of
secretory epithelial cells which secrete a layer of viscous, mucinous material on
surface.
 Degenerative Inflammation – degenerative changes in cytoplasm and nucleus
which undergo necrosis.
 Interstitial Inflammation – inflammatory reaction in the supportive fibrous
connective tissue stroma of organ.
 Proliferative Inflammation – increase in the number of cells especially
reticuloendothelial macrophages.
 Pseudomembranous Inflammation – involves mucous and serous membranes
characterized by tenacious membrane like covering made up of fibrin that is
firmly adherent to underlying acutely inflamed tissue.
  Purulent Inflammation – accumulation of PMNs in great number and their
enzymes causes liquefaction.

Terms for abnormal accumulations of fluid

A Transudate is protein-poor salt water squeezed through blood vessels by
hydrostatic pressure, i.e., it has specific gravity of extracellular fluid, 1.010 or
thereabouts.
An exudate is an abnormal, protein-rich fluid that has leaked out of inflamed vessels.

A body fluid (either an exudate or an area of liquefaction necrosis ) containing

neutrophilic leukocytes and necrotic debris is PUS. The preferred adjective to describe
things with lots of pus is Purulent. To produce pus is to Suppurate. Pus that literally
fills an important body cavity is called an Empyema. (This is most common in the
pleural cavities.) If you've got a lot of pus, you need it drained by a surgeon. Pus requires
no description, but it is worth mentioning that pus is not always the same color or
thickness.

Pus always has a yellow-green tinge because of myeloperoxidase.

Classic yellow pus (as in a staphylococcal boil) also includes some lipid from necrotic
tissue, hence the stronger yellow color.

Without necrosis (as in a streptococcal phlegmon), pus is more yellow-gray.

Pseudomonas bacteria make a pigment that imparts a blue-green fluorescent sheen to pus.

Clostridia produce extensive hydrolysis of tissue, and the discharge from clostridia
wound contains free lipid and other small molecules that impart a "dirty dish-water" look.

ACUTE INFLAMMATION

It is a stereotyped response to recent or ongoing injury. Although the process is complex,

the principal features are dilatation and leaking of vessels, and involvement of circulating
neutrophils.
You can recognize neutrophils in tissue sections by their segmented nuclei. Pus is
neutrophils plus liquefaction necrosis . Usually, the neutrophils themselves caused most
of the necrosis.

The "triple response of Lewis" to a superficial scratch (after the momentary

vasoconstriction): (1) a red scratch mark; (2) then a red flare around the scratch mark; (3)
then a red swollen area ("wheal") around the flare. (Try it!) Dr. Lewis found that he could
eliminate the flare, but not the others, by cutting the autonomic nerve supply (i.e.,
preventing the "axon reflex").

Sequence of Events in acute inflammation

Injury

Transient and inconstant vasoconstriction of arterioles (3-5 sec) may last for min
( neurogenic and adrenergic in nature)

Vasodiatation (first in arterioles and then capillaries – hallmark of early haemodynamic

changes in acute inflammation)

Increased Permeability of Vasculature

Escape of Protein Rich Fluid into Perivascular Tissue

Increased Concentration of RBC in Small Blood Vessels Leading To Increased Viscosity

Stasis of Blood Flows

Leukocyute Margination and Migration from the Vessel to the Site of Injury
(Duration in mild injury – 15-30 min or even greater
In severe injury – few minutes)

The various mechanisms proposed to explain the increased permeability of endothelium

of blood vessels are:
1) Endothelial contraction: immediate transient response
 It occurs in venules leaving arterioles and capillaries unaffected
 Most common mechanism of leakage and is elicited by histamine, bradykinin,
leukotriene and sub P.
 Leads to the formation of widened intercellular junctions
 Reversible and short lived( 15-30 min)

2) Endothelial retraction:
 Cytoskeleton of endothelial cells undergoes structural organization in such a way
that endothelial cells retract from one another.
 Reversible and can be induced in vitro by cytokine mediators (IL-1, TNF, INF),
hypoxia.
 Onset is delayed ( 4-6 yrs) but prolonged > 24 hours

3) Increased transocytosis across the endothelial cytoplasm:

 Occurs across channels consisting of clusters of interconnected, uncoated vesicles
and vacuoles called vesiculovacuolar organelle.
 Induced by histamine and most of the chemical mediators.

4) Direct endothelial injury: (immediate sustained response)

Results in endothelial cell necrosis and detachment. Leakage starts immediately after the
injury and is sustained at higher level for several hours.

5) Leukocyte mediated endothelial cell injury:

Occurs due to release of toxic oxygen species and proteolytic enzymes from activated
leukocytes.

6) Leakage from new blood vessels: New vessels which from remain leaky until
endiothelial cells differentiate and form intercellular junctions.

Cellular Changes

Leukcytes Extravasation and Phagocytosis

During inflammation reaction, leukocytes are delivered to the site of injury that ingest
offending agent.
Events that may occur during the journey from intravascular to extravascular space are:
1) In the lumen: Margination, rolling
Margination and rolling – in normally flowing blood, red and white cells are
confined to the axial central column, leaving relatively cell free layer of plasma in
contact with the venules. In stagnated blood flow, RBC,s tend to stick together and
forms clumps or rouleax, a process known as sludging. These clumps becomes larger
than WBC,s and occupy most rapidly moving central axis of slowly moving stream
displacing WBC,s to the periphery. Them WBC,s come closer to the vessel wall, this
is known as pavementing.
2) Adhesion and Transmigration across the endothelium (diapedesis)
It occurs by binding molecules on the leukocytes and endothelial surfaces. These
belong to 4 groups
 Selectins : these include three
E – selectins : confined to endothelium
P – selectins : endothelium and platelets
L – selectins : most leukocyte type
 Immunoglobulins : include 2 endothelial adhesion molecules
ICAM – 1 (intercellular adhesion molecule)
VCAM – 1(vascular cell adhesion molecule)
 Integrins
 Mucin like glycoprotein
Type of emigrating leukocyte varies i.e. in acute conditions mainly neutrophils
during 6-24 hours and then monocytes during 24-48 hours appear.

Notable exceptions to the "neutrophils first, monocytes later" rule:

(1) In viral and rickettsial infections, the lymphocytes are the principal cell;
(2) In classic allergy and in some parasitic infections, eosinophils dominate from start to
finish;
(3) In typhoid fever, the predominant cells are always the macrophages;
(4) In most forms of acute dermatitis, lymphocytes are most abundant;
(5) In clostridial gas gangrene, don't expect to see any white cells;
(6) In many kinds of bacterial infections (including chlamydial ones), there are few or
no other cells besides the neutrophils. In pseudomonas infection neutrophils appear
over 2-4 days.
Applied
Leukocyte adhesion molecules go by names such as LFA-1 and MO-1. These are
members of a homologous set.,
Their deficiency states are known. Obviously these patients mount a good increase in
neutrophils in response to infection, but the neutrophils don't enter the tissues very
well.

* Leukocyte adhesion deficiencies.... LAD I: Lack a CD11b/CD18 integrin; LAD II:

lack the selectin-binding Lewis X glycoprotein.
Endothelial adhesion molecules include ELAM-1 (for neutrophils) and ICAM-1 (for
neutrophils, lymphs, and monos). Various mediator proteins increase the numbers of
some or all of these.
* Alcoholism, diabetes, and glucocorticoid therapy all reduce the numbers of
adhesion molecules. You'll be impressed with the difficulty that such people have
with fighting off bacteria.
Pain itself (i.e., stimulation of pain fibers) causes the local neutrophils to lose their
ability to express L-selectin adhesion molecules. This seems to be one mechanism of
feedback that limits the acute inflammatory response

3) Migration in the interstitial tissue towards the chemotactic stimulus –

chemotaxis
It is the process of migration of leukocytes towards the site of injury. All the
granulocytes, monocytes and to lesser extent lymphocytes responsible for it.
Various agents responsible are:
For neutrophils : compliment c3a and c5a and leukotriene b4
For monocytes : c567, c3a, c5a and bacterial products, activated clotting factors
fibrinolytic , kinins and lymphokines
For eosinophils: ECF- A, products of parasites and products of complement.

Mechanism of chemotaxis
Chemotactic agent – on specific receptor on leukocyte surface membrane
 Activation of phospholipase

Hydrolysis of P1P2, IP3 and DAG

Release of Ca initially from intracellular stroma then from extracellular Ca

Increased activity of contractile element responsible for cell movement

Phagocytosis : 2 main cells
a) PMNs : In acute cases referred to as microphages
b) Circulating monocytes and fixed tissue mononuclear phagocytes refeffred to as
macrophages.
1. Recognition and attachment - by opsonins – Fc fragment of IgG
C3b
PP – collectins
2. The particle will then be engulfed and a lysosome membrane fused with the
phagosome membrane, causing digestion within the phagolysosome. (If only C3b
is present in the opsonin, additional molecules will be required to trigger
engulfment.) Some of the lysosomal enzymes will leak out of the neutrophil and
into the intercellular fluid.
3. Killing and degradation: a) killing of phagocytized bacteria is mediated through
the H2O2-myeloperoxidase-halide system
b) O2 dependent H2O2 production.
c) O2 independent mechanism by production of
bactericidal permeability increasing protein (BPI), lysozyme, lactoferrin and defensin.
Neutrophils product, including Lysosomal enzymes, H2O2, free radicals, and
Arachidonic acid metabolites are released during the process by "regurgitation during
feeding", "frustrated phagocytosis" (i.e., the neutrophil tries to eat something too big,
such as a huge immune complex or a splinter; it can't engulf it so it drools), and
"cytotoxic release". The latter is a euphemism for stuff leaking out of dead cells.
Applied
Neutrophil defects worth learning now:
1. Insufficient circulating neutrophils ("neutropenia"; "agranulocytosis"), as in radiation
injury, cancer therapy, drug sensitivity
2. Neutrophil adherence molecule defects, due to heredity, glucocorticoid administration,
diabetes, or ethanol in the bloodstream.
3. Failure of neutrophils to move properly (notably in diabetes) or to respond to
chemotactic stimuli
4. Failure of neutrophils to phagocytize (diabetics, people with complement or
immunoglobulin deficiencies)
5. Defective microbial killing. This may be due to
 Insufficient production of H2O2 ("Chronic granulomatous disease)"; affected
children must wall off catalase-producing microbes such as staphylococci with
monocytes instead. * Genetics and clinical picture J. Inf. Dis. 188: 1593, 2003.
 myeloperoxidase deficiency (less serious),
 * In HIV infection, the neutrophil defect is minor but probably real (Clin. Exp.
Immuno. 121: 311, 2000).
6. Mixed defects. Remember that diabetes and glucocorticoids interfere with most white
blood cell functions.

In CHEDIAK-HIGASHI SYNDROME (a problem with membrane synthesis), there are

too few neutrophils, they do not respond properly to chemotactic stimuli and their
(abnormally large) lysosomes fail to fuse with phagosomes.

Principal Mediators of Inflammation

Mediator: any substance or anatomical structure that transmits information

between 2 reagents, cells or organs.
1) Mediators originate either from plasma or from cells
2) They act by binding to specific receptors on target cells
3) A chemical mediator can stimulate the release of mediators by target cells
themselves
4) They may act on a single or few types of cells or they may have widespread
targets
5) They have very short life after their release
6) Most of them have the potential to cause harmful effects.

Vasoactive Amines
Histamine:
1) Richest source is mast cells and connective tissue adjacent to blood
vessels;also found in basophills and platelets.
2) Important in early inflammatory response
3) Released after degranulationin response to variety of stimuli like physical
injury, complement system.
4) Cause arteriolar dilatation and increased permeability of venules.

Serotonin: ( 5 hydroxytryptamine)

1) Stored in platelets and enterochromaffin cells

2) released from platelets when they aggregate in response to tissue injury
3) Causes arteriolar dilatation and sensitize afferent nerve endings leading to
pain.
4) Less potent than histamine.

Kinins:
Bradykinin – causes slow contraction of the smooth muscles.
 Derived from specific plasma proteins called kininogens by the action of specific
proteases called kallikreins
 Increases vascular permeability, dilates the blood vessels and cause algesia

Clotting System:

1) Thrombin: acts on soluble fibrinogen to produce an insoluble fibrin. Thrombin also

causes increased leukocyte adhesion and fibroblast proliferation.

2) Factor Xa: causes increased vascular permeability and leukocyte exudation.

Complement system

Consists of 20 complement proteins

Classic pathway alternate pathway

 (Ag – Ab reaction) (Non immunogenic – bacteria, toxins
IgA)
C1 C3

Activated C1 activated C3

C1a4b2a C3 C3bBb
(C3 convertase) C3a (C3 convertase)
C3b
C1a2a3b C5 C3bBb3b (C5 convertase)
C5a
C5b
C6, C7, C8, C9

C5 – 9
(Membrane attack complex)

Interrelationship b/w four plasma mediator systems triggered by activation of factor

X11

Factor X11 (high mol. Wt kininogen)

(Hagmen factor or surface active agent)

Kinin system clotting system

X11a
Prekallikerin kallikerin X1 X1a
HMWK bradykinin prothrombin thrombin
Fibrinolytic system
Plasminogen plasmin fibrinogen fibrin

Complement system fibrinopeptidase

C3 C3a fibrin split products

C3a and C5a – increase vascular permeability and cause vasodilation.

C5a – lipoxygenase pathway activated chemotactic agent for neutrophils, monocytes and
eosinophils.
C3b – acts as osonins and helps in phagocytosis.

Arachidonic Acid Metabolites:

Arachidonic acid is 20 carbon polysaturated fatty acid derived from either dietary source
or conversion from linoleic acid( essential fatty acid). Arachidonic acid metabolites are
also referred to as eicosanoids – synthesized by 2 major enzyme classes
Cyclooxygenase and lipoxygenase.

Cell membrane phospholipids

Phospholipase (steroids inhibit this)

Arachidonic acid chemotactic lipids

Lipoxygenase cycloxygenase (cox)

SHPETE (hydroperoxy eico satetraenoic acid) prostaglandin endoperoxideG2

Leukotriene A4 (LTA4) prostaglandin H2 (PGH2)

Vasoconstriction LTC4 prostacyclin thromboxaneA2
PGI2 TXA2
Bronchospasm LTD4 - inhibit platelet aggregation vasoconstriction
- vasodilation platelet aggregation
vascular permeability LTE4 - potentiates oedema

PGE PGD2 PGF2 HHT

Vasodilation and oedema chemotaxis

Generation of AA metabolites and their role in inflammation

2 isoforms of COX enzymes are there – COX 1 AND COX 2
COX 1 COX 2
- found in most normal cells and tissues - induced in inflammation by cytokines
- and mediators
- Involved in PGs synthesis in gastric mucosa - involved in PGs synthesis in the
platelets and kidney inflammation
- inhibited by non selective NSAIDS - inhibited by selective COX 2
inhibitors
Celecoxib,nimesulide, valdecoxib

Actions Metabolites
Vasconstriction : thromboxane, A2, Leucotrienes C4,D4,E4
Vasodilatation :PGI 2, PGE1, PGE2, PGD2
Increased Vascular Permeabilty : Leucotrienes C4,D4,E4
Chemotaxis, Leukocyte Adhesion : Leukotrienes B4, HETE, Lipoxins

Cytokines:
Proteins produced by many cell types that modulate function of other cell types. These
are generated by mononuclear phagocytes referred to as monokines and those by
lymphocytes are referred to as lymphokines. For eg cytokines responsible for
inflammation are IL- 1, TNF or cachetin.
Actions:
1) Acute phase reaction: Fever, increased sleep and acute phase proteins, decreased
appetite, neutrophilia
 2) Endothelial Effects: increased leukocyte adherence, PGI synthesis, increased
procoagulant activity, decreased anticoagulant activity
3) Fibroblastic effects: increased proliferation, increased collagen and PGE
synthesis, increased collagenase and protease
4) Leukocyte Effects: increased cytokine secretion.

Chemokines:
These are cytokines that share ability to stimulate leukocyte movement. These are Super
family of small proteins that act primarily as activators and chemoattractants for specific
types of leukocytes.

Platelet activating factors

These are released from IgE and they sensitize basophils, mast cells and leukocytes
which causes
Increased vascular permeability
Vasodilatation
Bronchoconstiction
Adhesion of leukocyte endothelium
Chemotaxis

Lysosomal component
Neutrophils and monocytes have these granules.
Neutrophilic granules – release lysozyme, alkaline phosphatase, lactoferrin, collagenase,
myeloperoxidase.
Monocyte granules – acid protease, collagenase, elastase, plasminogen activator.

Nitric Oxide:
 Soluable gas produced by endothelial cells, macrophages and specific neurons in
brain
 It is a potent vasodilator, reduces platelet aggregation and is also involved in the
pathogenesis of shock.
 Inhibit mast cell induced inflammation.

Oxygen Derived Free Radicals:

 Released extra cellularly from activated leukocytes. Major types are superoxide ion,
hydrogen peroxide and hydroxyl radical
Various processes in which they are implicated are:
1 Endothelial cell damage
2 Inactivation of antiproteases
3 Injury to cell types ( RBCs, parenchyma cells, tumor cells)
Body has antioxidant protective mechanism in serum, tissue fluids and target cells.
These include;
Serum protein - ceruloplasmin and tranferrin
Enzyme – peroxide dimutase, glutathione peroxidase and catalase which detoxifies
H2O2.

Outcomes of Acute Inflammation

1) Complete Resolution: It means complete return to normal tissue architecture and

function following acute inflammation. It occurs when injury is limited or short lived or
where there has been little tissue destruction and damaged parenchymal cells can
regenerate.
2) Abscess Formation: an abscess can be defined as a circumscribed collection of pus
appearing in acute or chronic localized area.
3) Healing by connective tissue replacement: It occurs when there is substantial tissue
destruction, abundant fibrin exudation or inflammation injury to the tissues that don’t
regenerate
4) Progression to chronic inflammation: It occurs when acute inflammation can’t be
resolved either due to interference in normal healing or persistence of injurious agent

Systemic Effects of Inflammation

1. Bacterial Infection of blood:

It can manifest in the following ways:
Bactermania: It is defined by the presence of bacteria that do not multiply
significantly
Septicemia: defined as the condition where highly pathogenic bacteria are multiplying
rapidly in blood.
 Pyaemia: formation and dissemination of small septic thrombi in blood that cause
their effect at the site where they are
THE SYSTEMIC INFLAMMATORY RESPONSE SYNDROME ("total-body
inflammation") represents toxicity from excessive production of the cytokines and/or
other white-cell products.
Venous return to the heart (i.e., venous responsivity) is compromised; perhaps
myocardial function is depressed, etc., etc., etc., etc., etc.
When it is caused by bacterial infection of the bloodstream, it's called SEPSIS.
* Deltibant ("Bradycor"), an anti-bradykinin antagonist, was once promoted as
improving survival in sepsis (JAMA 482: 277, 1997 -- didn't come into use) and for
post-traumatic neuroprotection (J. Neurotrauma 16: 431, 1999 -- again, doesn't seem
to have come into use.)
2. Fever: due to bactermia and endogenous pyrogens which is basic protein of low
molecular wt and act on thermoregulatory centre of hypothalamus
3. Leukocytosis: commonly accompany the acute inflammatory reaction and is in
the range of 15,000 – 20,000/microlitre. When 40000 – 1lakh – leukaemic reaction.
4. Malaise
5. Metabolic derangements
6. Shock and Hypotension
This is due to direct action of pyrogens/ PGs

Ant hypothalamus

Post hypothalamus

Vasomotor centre

Sympathetic nerve stimuli

Vasoconstriction of skin vessels and decreased heat dissipation

Fever
 CHRONIC INFLAMMATION

Definition - it is the type of inflammation which persists for a long duration usually for
weeks to months. "). It is a response to prolonged problems, orchestrated by T-helper
lymphocytes. It may feature recruitment and activation of T- and B-lymphocytes,
macrophages, eosinophils, and/or fibroblasts. Again, the process is complex. You will
recognize lymphocytes in tissue section by their small, "blue button" nuclei.

Etiology - persistent infection like TB, prolonged exposure to toxic substance and
autoimmunity.
H/F – 1. infiltration with mononuclear cells like macrophages, lymphocytes and plasma
cells.
2. Tissue injury
3. Attempt of healing by CT replacement of the damaged tissue, proliferation of the blood
vessels and fibrous tissue.
Other cells involved are –
Lymphocytes
Mast cells
Neutrophils

Granulomatous inflammation

GRANULOMAS are seen in certain chronic inflammation situations. They are

clusters of macrophages that have stuck tightly together, typically to wall
something off. Such macrophages are called epithelioid cells. You will recognize
granulomas in tissue sections by their characteristic appearance, or the presence of
giant cells.

 Distinct pattern, predominant cell is - activated macrophages which are modified

epitheloid cells.
 Surrounded by leukocytes – lymphocytes and plasma cells
 Epitheloid cells fuse to form giant cells in periphery and sometimes in the centre.
Peripheral arrangement occurs as two types:
Langhan’s type in – TB and sarcoidosis
Foreign body type – TB and infectious granulomaThe classic granulomatous diseases
include tuberculosis , tuberculoid leprosy , foreign body reactions (*  including the
reactions to everything from sutures to schistosome eggs ), the deep fungal infections,
berylliosis, and the mysterious disease "sarcoidosis".

* "Big Robbins" lists syphilis (the granulomas, if any, are small and loose) and
silicosis (the granulomas, if any, are very fibrous).
.
Wound Healing and Repair

Wound: it is defined as anatomic or functional interruption in the continuity of the tissue

that is accompanied by cellular damage and death.
Healing may be defined as the body response to injury in an attempt to restore normal
structure and function
It involves 2 distinct processes:
Regeneration: Replacement of injured cells by cells of same type sometimes leaving no
trace of previous injury.
Replacement/Repair: By connective tissue that leaves a permanent scar.

Both types of repair involves the formation of

1) Parenchyma or cellular structure of tissue
2) Stroma or connective tissue milieu in which cells reside

Normal Cell Cycle

It is divided into 4 phases:
M: Phase of mitosis
G: cell enters this phase immediately after mitosis
S: replication of DNA occurs in this phase
G2: between synthesis and mitosis phase

Repair:
Two processes involved in repair:
 Granulation tissue formation
 Contraction of wounds
Granulation tissue formation:
Slightly granular and pink appearance of tissue.
Each granule corresponds to proliferation of new blood vessels which are slightly lifted
on the surface by thin covering of fibroblast and collagen.
3 stages involved in the process:

Phase of inflammation: exudation of plasma, neutrophils and some monocytes with in

24 hours.
Phase of clearance: combination of proteolytic enzymes liberated from neutrophils ,
autolytic enzymes from dead tissue cells and phagocytic activity of macrophagesclear off
necrotic tissue, debris and RBC’s.
Phase of ingrowth of granulation tissue:
Angiogenesis or neovascularisation – by proliferation of endothelial cells from the
margins of severed blood vessels. New blood vessels are fragile giving oedematous
appearance to granulation tissue.
Formation of connective tissue - this occurs from fibrocytes which appear on sixth day.
As maturation proceeds, no of fibroblasts increase and that of blood vessel decrease
resulting in formation of inactive looking scar called as cicatrisation.

Contraction of the wound:

Wound starts contracting after two to three days and process completes by 14 th day.
During this wound is reduced by approx. 80% of original size. Contracted wound results
in rapid healing since less surface area of injured tissue has to be replaced.

Depending upon the relationship of the cell cycle and proliferative potential cells of
the body are classified into 3 main groups

Labile cells
 Continually dividing cells that follow the cell cycle from one mitosis to other and
proliferate throughout the life.
 Surface epithelia, cells of the bone marrow, haemopoitic tissue, epithelium GIT,
urinary tract, mucosa of oral cavity and lining mucosa of excretory ducts of the
glands of body contain labile cells.
Stable cells
 Normally show low level of replication but can undergo rapid cell division in
response to stimuli
 They are considered in G0 phase and can be stimulated to G1 readily.
 Parenchyma of the liver, kidneys , pancreas, mesenchymal cells like fibroblasts
and smooth muscle cells and vascular endothelial cells can be classified as stable
cells.

Permanent cells
Non dividing cells that have left the cell cycle and cant undergo cell mitosis irrespective
of any stimulus. e.g. neurons, cardiac muscle, cells of nervous system

Extracellular matrix
Secreted locally and assembles into network in the spaces surrounding the cells.
It functions like
 Retain water to provide turgor to the tissue
 Retains mineral to provide rigidity to the skeletal tissues
 Provides reservoir for growth factors controlling cell proliferation
 Provides substratum for cells to adher, migrate and proliferate

Components
Fibrous structural proteins – collagen and elastin
Adhesive glycoproteins – fibronectin and laminin
A gel of proteoglycans and hyaluronin

Wound Healing

It is a complex but orderly phenomenon involving number of processes like

 Inflammatory process following an initial injury
 Regeneration of the parenchymal cells
 Migration and proliferation of both parenchymal and connective tissue cells
 Synhesis of extracellular matrix proteins
 Remodeling f connective tissue and parenchymal components
  Collagenization and increase in wound strength

Factors affecting wound healing

Local Factors:
Location of wound: type of tissue in which injury has occurred. More the amount of
labile or stable cells in the tissue, more adequate is healing.
Vascularity of the area:Wound in the area good vascular supply heals faster than
wounds in the vascular area. Poor oxygenation interferes with collagen synthesis.

Immobilization of the wound: wound under constant movement cause delayed healing.
Other Factors:
a) Severe trauma the area of the wound results in the delayed healing.
b) Slightly increased temperature of the area will accelerate the wound healing
whereas environmental hypothermia will lead to delayed healing.
c) Low does of X-radiation to the area tends to stimulate wound healing.
Infection in the area:
In the oral cavity, it is one of the most important factor affecting wound healing.severe or
established infection results in delayed healing or non healing.
Foreign bodies:
Foreign bodies at the site lead to delayed or non healing.
Approximation of wound margins:
Close approximation leads to proper and faster healing healing.

Systemic Factors:
1) Age of the patient:
Wounds in elderly patients heal slowly due to decrease in tissue metabolism and
poor blood supply.
2) Nutrition:
Delayed healing occurs in patients who are deficient in any of the essential
nutrients.
3) Proteins:
Patient with hypoproteinmia shows decreased formation and multiplication of
fibroblasts leading to delayed wound healing.
4) Vitamins and Zinc:
Vitamin C and zinc is important for formation of collagen. Similarly vitamin A
and D also play a role in normal healing.

Circulating Factors:
Anaemia, neutropenia and dehydration reslts in delay in wound healing.

Hormonal Factors:
Both ACTH and cortisone have been shown to affect wound healing.
In patients taking ACTH or cortisone, growth of granulation tissue is inhibited.
Diabetes mellitus also results in delay I wound healing.
Corticosteroids decrease the wound strength if given within first 3 days of injury.
Drugs like indomethacin delay the wound healing.
Antineoplastic agents have cytotoxic effect and retard healing.
Miscellaneous:
- Wound sutured with non resorbable sutures tends to be weaker than wounds with
absorbable one.
-There is lower incidence of infection with monofilaments sutures
-Tissue adhesives like butyl cyanoacrylates not only permit uncomplicated but also
hasten the healing process.

Wound Healing Can Be Categorized Into:

Healing By Primary Intention –
Occurs when a sharply made cut is accurately reapproximated with in hours and heals
with minimum space between the edges eg surface incision closed by sutures or fractures
accurately reduced using compression plates.

Healing By Secondary Intention –

Involves filling the tissue defect through formation of connective tissue eg seen after
excisional or avulsive injury that leaves a residual defect and in spontaneous healing of
compound fractures.

Healing By Third Intention –

Occurs when the wound is surgically closed after healing by secondary intention has
begun or when tissue grafts are used to assist in healing of wounds. This type of closure
is also known as delayed primary closure.

Healing by primary intention –

Events that occur during this type of healing are
First few hours:
- Filling of the space by small amount of clotted blood
- Dehydration of surface clot forming scab that covers the wound and seal it from
the environment.
First 24 hours:
- Acute neutrophillic response at the wound margins
- Thickening of the epidermis at wound margins because of the mitotic activity of
the basal cells
24 – 48 hours:
- Growth of epidermal cells along the cut margins of epidemis
- Migration of the epithelial cells from the margins
- Fusion of the epithelial cells from both the margins of the wound in midline
beneath the surface scab.
Third Day:
- Replacement of nutrophils by macrophages
- Hypertrophy of subepithelial fibroblasts and their replication
- Sprouting of capillary buds from the wound margins towards the midline
- Invasion of wound space by granulation tissue
- Collagen fibres present but are vertically oriented and therefore no bridging the
gap.
5th day:
Neovascularisation is maximum. Epidermis recovers in normal thickness.
End of First Week:
- Filling of wound space by granulation tissue
- Fusion of capillary buds from both the sides to form continuous channels
- Collagen fibres start bridging the gap
-Epidermis attain its normal thickness
End of Second Week:
 -Continuous collagen deposition and proliferation of fibroblast within the
connective tissue
- Decrease in degree of vascularisation
End of First Month:
- No inflammation
- Scar composed of cellular connective tissue
- Further there is more and more decrease in vascularisation. It almost takes a year
to become avascular, acellular, pale and collagenous.
- Dermal appendages that have been destroyed due to injury are permanently lost,
those partially destroyed along the wound margins may regenerate.
- Little or no wound contraction takes place.

Suture tracks – each suture track is a separate wound and incites same phenomenon as
in healing of primary wound. When sutures are removed around seventh day, much of
epithelised suture track is avulsed and remaining epithelial tissue in the tract is absorbed.
However, sometimes suture track gets infected (stitch abscess) or epithelial cells may
persist in the track (implantation and epidermal cysts)

Healing By Secondary Intention

- Occurs in cases where there is extensive loss of cells and tissues creating large
defects accompanied by intense inflammatory response.
- Defect is filled by growth of granulation tissue.
It differs from healing by first intention in the following ways:-
1) More loss of normal tissue architecture
2) More necrotic tissue and exudates
3) More intense inflammatory reaction.
4) Absolute dependence on growth of granulation tissue for filling of defect.
5) More wound contraction due to action of myofibroblast in granulation tissue.
Wound contracts to ½ to ¼ of its original size.
6) Larger scar
7) Slow healing response

Chronology Of Events
First few hours:
- Clot and scab formation
- Acute inflammatory reaction at junction of living tissue and clot
24-48 hours:
- Epithelial continuity restored because of mitotic activity of cells at the wound margins
- Wound contraction starts
- Sprouting of capillary buds
- Macrophages start replacing neutrophills
End of first week:
- Wound contraction continues
- Growth of epithelium
- Shedding of surface debris
- Formation of small granulation tissue formation

End of second week:

- Wound contraction continues
- Epithelial covering completed
- Collagen arrangement starts in the transverse direction, bridging the gap
- Vascularity of wound decrease
End of the first month
- Full thickness of epithelial restored
- Varying degree of surface depression remains
- Thick collagen scar present in underlying connective tissue

Wound Contraction
It is a complex process that produces a dramatic spatial decrease in wound area (1/3 to ¼
of original size) and together with re-epitheliazation, accounts for significant proportion
of wound closure
Two factors that are most important in the wound contraction are:
- The formation of significant quantity of granulation tissue
- Presence of movable wound edges

Wound contraction is believed to be due to young specialized cells called myofibroblast.

In fresh wound, most of the tensile strength is due to sutures placed and later on collagen
formation is responsible for wound strength. At the end of first week, wound strength is
approximately 10% of uninjured skin.

Scar Remodelling
- Results in reduction of scar size and reorganization of collagen fibres.
- Regulated to a large extent by tissue colagenases that carry out collagenolysis and
prevent excessive build up of collagen
- Initially during repair, collagen fibres are deposited in a haphazard manner. Scar
remodeling results in gradual orientation and with reorganization, results in closely
aligned collagen fibres that allow for increased intermolecular bonding and stimulation
increase in tensile strength.

Complication Of Wound Healing

1) Contracture:
- Specially seen in cases where healing is by secondary intention
- Generally due to thickening an shortening of collagen bundles
- Cause distortion, may lead to cosmetic and functional deformity
2) Proud Flesh:
- Excessive formation of granulation tissue formation
- May protrude above margins and block re-epitheliasation
- May be treated by chemical cauterization or surgical excision
3) Keloid:
- Seen most commonly in blacks
- Genetically related
-Abnormal amount of collagen formation in connective tissue leading to bulging
tumorous scar
- margins ar tenser, red, vascular, erythematous and spread to normal tissue” claw like
process”
- Most common site is sternum
- More in women
- Cause is unknown but it may be due to deficient polymerization of fibrin( inadequate
lysis of collagen by collagenase and )
Purists: Keloids send little processes into the surrounding connective tissue, like the
serrations on a crab's claw. If these are absent, an exuberant scar is merely called
"hypertrophic". A keloid is likely to grow over time. A hypertrophic scar is likely to
regress over time. Both can show glassy fibers.
4) Infection
5) Implantation Cyst*
6) Deficient scar
7) Sq Cell carcinoma

Treatment
- Intrakeloid injections steroids (inj triamcinolone), hyaluronidase, vitamin
methotrexate have been tried with some success
- Deep X ray and ultrasonic therapy
- Surgical excision of excess tissue and resurfacing of area with thick skin grafts.
But in some cases recurrence can occur.
Newer remedies that show promise are tamoxifen (changes the milieu of fibroblast
growth factors) and 5-fluorouracil.

Healing In Special Situations

Healing of fracture:
Healing of a fracture can be considered as regeneration rather than repair because
characteristic structure of bone is restored

Osseous healing can be described as:-

Indirect: follows the routine histologic cascade involving granulation tissue,
fibrocartilage and bone. It is characterized by formation of callus.
Direct: takes place by intracortical remodeling without formation of intervening callus.

Indirect Fracture Healing

First 4-5 Days:
- Hemorrhage from harvesian and periosteal vessels into the adjacent area.
- Clotting of blood and loose fibrin meshwork formation
- Necrosis at the ends of fractured bone
1-3 Weeks:
- Fibroblastic activity and capillary buds sprouting
- Removal of necrotic material by granulation tissue formation
- Soft tissue callus formation
-Formation of new cartilage and bone matrix by chondroblast and osteoblast respectively
- Dispersion of bone and cartilage spicules in soft tissue callus
- Procallus formation, temporary bone formation is achieved. It is also called provisional
callus and is of three types
External
Intermediate
Internal
- Increased bone formation in procallus leading to primary bony callus formation
Months:
- Bone maturation in fracture area and secondary callus formation
- Functional remodeling of fractured bone

Osseous callus formation:

The procallus acts as scaffolding on which callus composed of lamellar bone is formed.
The woven bone is cleared away by incoming osteoblasts. In this place, newly formed
blood vessels and osteoblasts invade; laying down osteoid which is calcified and lamellar
bone is formed by developing haversian system concentrically around blood vessel.
Cartilaginous callus (soft callus) - internal and external
Soft callus formed around # site increase the strength and stiffness of the bone during
healing period.
Hard (bony callus) – cartilaginous callus undergo calcification into woven bone.

Remodelling
The external callus is cleared away. Compact bone is formed in place of intermediate
callus and bone marrow cavity develops in internal callus.
Remodeling is a slow process in accordance with Wolff’s law which states that changes
in functional state of bone causes structured and architectural changes in tissue through
bioelectric field production.
If gap is < 0.3mm, lamellar bone formed directly and if 0.5 – 1mm then filled with woven
bone and subsequently lamellar bone is laid down.
Depending upon location and function, primary callus can be divided into 4 types:

Uniting callus: formed between fractured ends of bone by direct ossification giving a
well united fracture
Anchoring callus: develops near the periosteum on the external surface of the bone away
from the site of fracture
Bridging Callus: forms on external surface between the two fractured ends and
anchoring callus. It is primarily cartilaginous
Sealing Callus: develops on the endosteal surface near uniting callus.

Direct Bone Healing

Follows excellent anatomic apposition and rigid maintainance of fracture has
consolidated.
Events that occur during healing are:
- Accurate adaption of fractured ends
- No space for hematoma formation
- Osteoclast formation from cutting cone at the ends of harvesian canal close to
the fracture site
-Tunneling by osteoclast across the fracture site to form enlarged harvesian canals
- Migration of vessels and osteoblast in the tunnel forming a new osteon across
the fracture line that replaces the old bone
- Deposition of new osteoid in the canals that eventually matures into lamellar
bone

The process described above is contact healing. However in clinical setting,

interfragmentary compression is not always homogenously distributed with minute gaps
remaining between the fractured ends.Fibrous bone fills the gaps that is replaced by
lamellar bone, which is gradually restructured by harvesian remodeling. This type of
healing is called gap healing. Osteoclasts cut bone at the rate of 50- 80 micron m/day.
Osteon forms at the rate of 1-2 micron m/day hence radiolucency in compact bone is seen
up to 3months and complete reconstruction of cortex is in 6 months.

Complications Of The Fracture Healing

Infection: Especially in cases of compound fracture
Fat embolism: especially in fracture of long bones
Malunion: occurs when accurate anatomic approximation of fracture ends is not there at
the onset or the approximation lost during initial period of healing or in cases where no
treatment is given
Pseudoarthosis: occurs when there is inadequate immobilization of fracture due to
fibrous union.
Non Union: occurs when callus for both the ends fail to fuse in midline
Osteomylitis: occurs due to infection in the area of healing, usually begins in the marrow
spaces and spread to cortical bone.

Healing Of Extraction Wound

Specialized example of wound healing by secondary intention

Various Events:

First 24 hours:
- Filling of the socket with blood
- Coagulation of blood and fibrin meshwork formation
- Inflammatory reaction follows
24-48 hours:
-Vasodilatation and engorgement of blood vesels in periodontal ligament remants
-Mobilization of leukocytes in the immediate area around the clot
First week:
-Fibroblastic proliferation into the clot and periphery
-Alveolar crestal bone of socket margin starts to show osteoclastic activity
-Epithelial cell proliferation
-Organization of blood clot
-Formation of capillary buds and their sprouting occurs
Second Week:
-Penetration of clot by newly formed capillaries
-Degeneration of periodontal ligament remnants
-Osteiod trabeculae can be seen
 -Epithelial proliferation over the wound margin continues
Third Week:
- Organization of blood clot by maturing granulation tissue
-Increased osteoid formation around the periphery of wound
-Complete epithelialization of the wound surface
Fourth Week onwards:
-Continuous formation and remodeling of bone

Complication
Alveolar osteitis (Dry Socket)
Most important cause of post operative pain
Focal osteomyelitic reaction because of loss of blood clot
There is production of foul odor and severe radiating pain but no suppuration

Causes
Pre-existing or post extraction infection
Trauma during extraction
Decreased bleeding due to vasoconstrictor present in local anesthetic solution
Loss of blood clot because of month rinsing
General debilitation

Symptoms start on 3rd to 5th post operative day and may last upto 14 days if left untreated

Treatment
Primarily aimed at relief of pain
Irrigation of socket with saline
Placement of obtundent or topical anesthetic dressing or placement of zinc oxide pack is
advised
Analgesic can be prescribed for relief of pain
Antibiotics are not recommended unless there is suppuration or any other systemic signs
of infection

Fibrous Healing
Normally occurs when tooth healing is followed by loss of both labial and lingual cortical
plates
Generally is asymptomatic
Histologically consist of dense bundles of collagen with few fibrocytes and blood vessels
Radiographically well circumscribed radiolucent area at site of previous extraction seen

Healing Of Biopsy Wound

Biopsy can be defined as the removal of tissue from the living organism for the purpose
of microscopic examination and diagnosis
It can be excisional (total excision of a small lesion for microscopic study) incisional
(removal of small section of a large lesion along with the part of normal tissue)
Healing of biopsy wound of oral cavity is identical with the healing of similar wounds in
other parts of the body and thus it may occur by primary or secondary intentions.

Healing of Nerves

Peripheral nerve injury occurs due to trauma either accidental or iatrogenic

Seddon’s Classification (1943)
Neuropraxia – simple contusion of nerve, mildest, continuity of both epineural sheath
and axon is maintained. Disintegration of myelin, proximal to injury responds within few
days to weeks usually up to 4 weeks)
Axontemesis – destruction of both axon and myelin sheath, endoneurium is intact. It is
seen in crushing injuries and regeneration is within 2-6 months.
Neurotemesis – complete transaction of the nerve trunk. Little chances of repair without
microsurgery.

Sunderland Classification (1978)

First degree- equivalent to neuropraxia
Second, third and fourth degree-equivalent to axontemesis
Fifth degree-equivalent to neurotemesis
First degree – conduction block bcoz of anoxia from interruption of segmental or
epineural blood vessel, but no axonal degeneration of myelination .
Second degree – axon damaged, intact endoneural sheath, peri and endoneurium.
Third degree – axon damaged, endoneural sheath breached but peri and epineurium is
intact.
Fourth degree – disruption of axon, endo and perineurium but epineurium is intact.
Fifth degree – damage to all components of nerve trunk.

Following transaction or crush injury, myelin sheath of nerve segment distal to site of
injury starts breaking down. The severed axonal segment undergoes wallerian
degeneration upto peripheral nerve endings. Similar degeneration is also seen in
proximal nerve segment upto first node of Ranvier. Schwann cells close to site of injury
starts to proliferate within first week post injury and grow towards each other resulting in
formation of Hanke Bunger Bands which bridges the gap formerly occupied by
transected axon.As the axon buds advances at the rate of 1-3 mm per day, investing
Schwann cells cause a new myelin sheath to form around it. The regeneration phase last
upto 3 months and ends on contract with ends on contact with end organ by a thin
myelinated axon.

Superior functional results are obtained by microsurgical repair carried out by direct
reanastomosis of severed ends or by interposition o sural or great auricular nerve graft.

Free Skin Graft Healing

Skin grafts may be either full thickness or split thickness
Nutritional support for free skin graft is initially provided by plasma that exudes from
dilated capillaries of host bed.
A fibrin clot forms the graft host interface fixing the graft to host interface
Graft survival depends on in growth of blood vessels from the host into the graft and
direct anastomosis between graft and host vasculature
The endothelial capillary buds from the host invade the graft at 48 hours and vascular
connections are also established between them.
Fibrin clot is slowly resorbed. By 9 th day, new blood vessels and fibroblast achieved firm
union with host bed
Re-innervation of graft occurs by nerve fibres entering the graft from its base. Recovery
of sensation usually begins within 2 months.
References
1) Pathological basis of disease by Kumar, Cortran and Collins
2) Principles of oral and maxillofacial surgery by L.J.Peterson
3) Essential pathology by Harshmohan
4) Oral and maxiallofacial trauma by Raymond J Fonseca
5) Textbook of oral pathology by William G Shafer