Medical physiology Lec 8

Hemostasis and blood coagulation

The term hemostasis means prevention of blood loss. Whenever a vessel is
severed or ruptured, hemostasis is achieved by several mechanisms:
(1) vascular constriction,
(2) formation of a platelet plug,
(3) formation of a blood clot as a result of blood coagulation, and
(4) eventual growth of fibrous tissue into the blood clot to close the hole
in the vessel permanently.

Vascular spasm:
Vasoconstriction is produced by vascular smooth muscle cells, and is the
blood vessel's first response to injury. When a blood vessel is damaged,
there is an immediate reflex, initiated by local sympathetic pain receptors,
which helps promote vasoconstriction. The damaged vessels will constrict
(vasoconstrict) which reduces the amount of blood flow through the area
and limits the amount of blood loss. Collagen is exposed at the site of
injury; the collagen promotes platelets to adhere to the injury site. Platelets
release cytoplasmic granules which contain serotonin, ADP and
thromboxane A2, all of which increase the effect of vasoconstriction. The
spasm response becomes more effective as the amount of damage is
increased. Vascular spasm is much more effective in smaller blood
vessels.

Formation of the Platelet Plug

Physical and Chemical Characteristics of Platelets
Platelets (also called thrombocytes) are minute discs 1 to 4 micrometers
in diameter. They are formed in the bone marrow from megakaryocytes,
which are extremely large cells of the hematopoietic series in the marrow;
the megakaryocytes fragment into the minute platelets either in the bone

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marrow or soon after entering the blood, especially as they squeeze through
capillaries.
The normal concentration of platelets in the blood is between 150,000 and
300,000 per microliter.

Platelets have many functional characteristics of whole cells, even though

they do not have nuclei and cannot reproduce. In their cytoplasm are such
active factors as
(1) actin and myosin molecules, which are contractile proteins similar to
those found in muscle cells, and still another contractile protein,
thrombasthenia, that can cause the platelets to contract;
(2) residuals of both the endoplasmic reticulum and the Golgi apparatus
that synthesize various enzymes and especially store large quantities of
calcium ions;
(3) mitochondria and enzyme systems that are capable of forming
adenosine triphosphate (ATP) and adenosine diphosphate (ADP);
(4) enzyme systems that synthesize prostaglandins, which are local
hormones that cause many vascular and other local tissue reactions;
(5) an important protein called fibrin-stabilizing factor, which we discuss
later in relation to blood coagulation; and
(6) a growth factor that causes vascular endothelial cells, vascular smooth
muscle cells, and fibroblasts to multiply and grow, thus causing cellular
growth that eventually helps repair damaged vascular walls
The cell membrane of the platelets is also important. On its surface is a
coat of glycoproteins that repulses adherence to normal endothelium and
yet causes adherence to injured areas of the vessel wall, especially to
injured endothelial cells and even more so to any exposed collagen from
deep within the vessel wall.
Mechanism of the Platelet Plug
Platelet repair of vascular openings is based on several important functions
of the platelet itself. When platelets come in contact with a damaged

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vascular surface, especially with collagen fibers in the vascular wall, the
platelets themselves immediately change their own characteristics
drastically. They begin to swell; they assume irregular forms with
numerous irradiating pseudopods protruding from their surfaces;
- Their contractile proteins contract forcefully and cause the release of
granules that contain multiple active factors; they become sticky so that
they adhere to collagen in the tissues and to a protein called von
Willebrand factor that leaks into the traumatized tissue from the plasma;
-they secrete large quantities of ADP; and their enzymes form
thromboxane A2.
-The ADP and thromboxane in turn act on nearby platelets to activate them
as well, and the stickiness of these additional platelets causes them to
adhere to the original activated platelets. Therefore, at the site of any
opening in a blood vessel wall, the damaged vascular wall activates
successively increasing numbers of platelets that themselves attract more
and more additional platelets, thus forming a platelet plug. This is at first a
loose plug, but it is usually successful in blocking blood loss if the vascular
opening is small. Then, during the subsequent process of blood
coagulation, fibrin threads form.
These attach tightly to the platelets, thus constructing unyielding
plug.
Mechanism of Blood Coagulation
Basic Theory. More than 50 important substances that cause or affect blood
coagulation have been found in the blood and in the tissues—some that
promote coagulation, called procoagulants, and others that inhibit
coagulation, called anticoagulants. Whether blood will coagulate depends
on the balance between these two groups of substances. In the blood
stream, the anticoagulants normally predominate, so that the blood does
not coagulate while it is circulating in the blood vessels. But when a vessel
is ruptured, procoagulants from the area of tissue damage become
“activated” and override the anticoagulants, and then a clot does
develop.
Conversion of Prothrombin to Thrombin
First, prothrombin activator is formed as a result of rupture of a blood
vessel or as a result of damage to special substances in the blood

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Second, the prothrombin activator, of ionic Ca++, causes conversion of
prothrombin to thrombin. Third, the thrombin causes polymerization of
fibrinogen molecules into fibrin fibers within another 10 to 15 seconds.
Thus, the rate-limiting factor in causing blood coagulation is usually the
formation of prothrombin activator and not the subsequent reactions
beyond that point, because these terminal steps normally occur rapidly to
form the clot itself. Platelets also play an important role in the conversion
of prothrombin to thrombin because much of the prothrombin first attaches
to prothrombin receptors on the platelets already bound to the damaged
tissue.
Conversion of Fibrinogen to Fibrin— Formation of the Clot Fibrinogen.
Fibrinogen is a high-molecular-weight protein (MW = 340,000) that occurs
in the plasma in quantities of 100 to 700 mg/dl. Fibrinogen is formed in the
liver, and liver disease can decrease the concentration of circulating
fibrinogen, as it does the concentration of prothrombin, pointed out above.
Because of its large molecular size, little fibrinogen normally leaks from
the blood vessels into the interstitial fluids, and because fibrinogen is one
of the essential factors in the coagulation process, interstitial fluids
ordinarily do not coagulate. Yet, when the permeability of the capillaries
becomes pathologically increased, fibrinogen does then leak into the tissue
fluids in sufficient quantities to allow clotting of these fluids in much the
same way that plasma and whole blood can clot.

Extrinsic Pathway for Initiating Clotting

The extrinsic pathway for initiating the formation of prothrombin activator
begins with a traumatized vascular wall or traumatized extravascular
tissues that come in contact with the blood. This leads to the following
steps,
1- Release of tissue factor. Traumatized tissue releases a complex of
several factors called tissue factor or tissue thromboplastin. This factor is
composed especially of phospholipids from the membranes of the tissue
plus a lipoprotein complex that functions mainly as a proteolytic enzyme.
2. Activation of Factor X—role of Factor VII and tissue factor. The
lipoprotein complex of tissue factor further complexes with blood

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coagulation Factor VII and, in the presence of calcium ions, acts

enzymatically on Factor X to form activated Factor X (Xa).
3. Effect of activated Factor X (Xa) to form prothrombin activator—role
of Factor V. The activated Factor X combines immediately with tissue
phospholipids that are part of tissue factor or with additional phospholipids
released from platelets as well as with Factor V to form the complex called
prothrombin activator. Within a few seconds, in the presence of calcium
ions (Ca++), this splits prothrombin to form thrombin, and the clotting
process proceeds as already explained. At first, the Factor V in the
prothrombin activator complex is inactive, but once clotting begins and
thrombin begins to form, the proteolytic action of thrombin activates
Factor V. This then becomes an additional strong accelerator of
prothrombin activation. Thus, in the final prothrombin activator complex,
activated Factor X is the actual protease that causes splitting of
prothrombin to form thrombin; activated Factor V greatly accelerates this
protease activity, and platelet phospholipids act as a vehicle that further
accelerates the process. Note especially the positive feedback effect of
thrombin, acting through Factor V, to accelerate the entire process once it
begins

Intrinsic Pathway for Initiating Clotting

The second mechanism for initiating formation of prothrombin activator,
and therefore for initiating clotting, begins with trauma to the blood itself
or exposure of the blood to collagen from a traumatized blood vessel wall.
Then the process continues through the series of cascading reactions.

1.Blood trauma causes

(1) activation of Factor XII and
(2) release of platelet phospholipids. Trauma to the blood or exposure of
the blood to vascular wall collagen alters two important clotting factors in
the blood: Factor XII and the platelets. When Factor XII is disturbed, such
as by coming into contact with collagen or with a wettable surface such as
glass, it takes on a new molecular configuration that converts it into a
proteolytic enzyme called “activated Factor XII.” Simultaneously, the

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blood trauma also damages the platelets because of adherence to either
collagen or a wettable surface (or by damage in other ways), and this
releases platelet phospholipids that contain the lipoprotein called platelet
factor 3, which also plays a role in subsequent clotting reactions.
2. Activation of Factor XI. The activated Factor XII acts enzymatically on
Factor XI to activate this factor as well, which is the second step in the
intrinsic pathway. This reaction also requires HMW (high-molecular-
weight) kininogen and is accelerated by prekallikrein.
3. Activation of Factor IX by activated Factor XI. The activated Factor XI
then acts enzymatically on Factor IX to activate this factor also.
4. Activation of Factor X—role of Factor VIII. The activated Factor IX,
acting in concert with activated Factor VIII and with the platelet
phospholipids and factor 3 from the traumatized platelets, activates Factor
X. It is clear that when either Factor VIII or platelets are in short supply,
this step is deficient. Factor VIII is the factor that is missing in a person
who has classic hemophilia, for which reason it is called antihemophilic
factor. Platelets are the clotting factor that is lacking in the bleeding disease
called thrombocytopenia.
5. Action of activated Factor X to form prothrombin activator—role of
Factor V. This step in the intrinsic pathway is the same as the last step in
the extrinsic pathway. That is, activated Factor X combines with Factor V
and platelet or tissue phospholipids to form the complex called
prothrombin activator. The prothrombin activator in turn initiates within
seconds the cleavage of prothrombin to form thrombin, thereby setting into
motion the final clotting process, as described
earlier.
Prevention of Blood Clotting in the Normal Vascular System—
Intravascular Anticoagulants Endothelial Surface Factors.
Probably the most important factors for preventing clotting in the normal
vascular system are
(1) the smoothness of the endothelial cell surface, which prevents contact
activation of the intrinsic clotting system;
(2) a layer of glycocalyx on the endothelium (glycocalyx is a
mucopolysaccharide adsorbed to the surfaces of the endothelial cells),

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which repels clotting factors and platelets, thereby preventing activation of
clotting; and
(3) a protein bound with the endothelial membrane, thrombomodulin,
which binds thrombin. Not only does the binding of thrombin with
thrombomodulin slow the clotting process by removing thrombin, but the
thrombomodulin-thrombin complex also activates a plasma protein,
protein C, that acts as an anticoagulant by inactivating activated Factors V
and VIII. When the endothelial wall is damaged, its smoothness and its
glycocalyx-thrombomodulin layer are lost, which activates both Factor XII
and the platelets, thus setting off the intrinsic pathway of clotting. If Factor
XII and platelets come in contact with the subendothelial collagen, the
activation is even more powerful.
Antithrombin Action of Fibrin and Antithrombin III.
Among the most important anticoagulants in the blood itself are those that
remove thrombin from the blood. The most powerful of these are
(1) the fibrin fibers that themselves are formed during the process of
clotting and (2) an alpha-globulin called antithrombin III or antithrombin-
heparin cofactor. While a clot is forming, about 85 to 90 per cent of the
thrombin formed from the prothrombin becomes adsorbed to the fibrin
fibers as they develop. This helps prevent the spread of thrombin into the
remaining blood and, therefore, prevents excessive spread of the
clot.
The thrombin that does not adsorb to the fibrin fibers soon combines with
antithrombin III, which further blocks the effect of the thrombin on the
fibrinogen and then also inactivates the thrombin itself during the next 12
to 20 minutes.
Heparin. Heparin is another powerful anticoagulant, but its concentration
in the blood is normally low, so that only under special physiologic
conditions does it have significant anticoagulant effects. However, heparin
is used widely as a pharmacological agent in medical practice in much
higher concentrations to prevent intravascular clotting.

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Prevention of Blood Coagulation Outside the Body

Although blood removed from the body and held in a glass test tube
normally clots in about 6 minutes, blood collected in siliconized containers
often does not clot for 1 hour or more. The reason for this delay is that
preparing the surfaces of the containers with silicone prevents contact
activation of platelets and Factor XII, the two principal factors that initiate
the intrinsic clotting mechanism. Conversely, untreated glass containers
allow contact activation of the platelets and Factor XII, with rapid
development of clots. Heparin can be used for preventing coagulation of
blood outside the body as well as in the body. Heparin is especially used in
surgical procedures in which the blood must be passed through a heart-lung
machine or artificial kidney machine and then back into the person.
Various substances that decrease the concentration of calcium ions in the
blood can also be used for preventing blood coagulation outside the body.
For instance, a soluble oxalate compound mixed in a very small quantity
with a sample of blood causes precipitation of calcium oxalate from the
plasma and thereby decreases the ionic calcium level so much that blood
coagulation is blocked. Any substance that deionizes the blood calcium
will prevent coagulation. The negatively charged citrate ion is especially
valuable for this purpose, mixed with blood usually in the form of sodium,
ammonium, or potassium citrate. The citrate ion combines with calcium in
the blood to cause an un-ionized calcium compound, and the lack of ionic
calcium prevents coagulation. Citrate anticoagulants have an important
advantage over the oxalate anticoagulants because oxalate is toxic to the
body, whereas moderate quantities of citrate can be injected intravenously.
After injection, the citrate ion is removed from the blood within a few
minutes by the liver and is polymerized into glucose or metabolized
directly for energy. Consequently, 500 milliliters of blood that has been
rendered incoagulable by citrate can ordinarily be transfused into a
recipient within a few minutes without dire consequences. But if the liver
is damaged or if large quantities of citrated blood or plasma are given too
rapidly (within fractions of a minute), the citrate ion may not be removed
quickly enough, and the citrate can, under these conditions, greatly depress
the level of calcium ion in the blood, which can result in tetany and
convulsive death.

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Lysis of Blood Clots—Plasmin The plasma proteins contain a euglobulin

called plasminogen (or profibrinolysin) that, when activated, becomes a
substance called plasmin (or fibrinolysin). Plasmin is a proteolytic enzyme
that resembles trypsin, the most important proteolytic digestive enzyme of
pancreatic secretion. Plasmin digests fibrin fibers and some other protein
coagulants such as fibrinogen, Factor V, Factor VIII, prothrombin, and
Factor XII. Therefore, whenever plasmin is formed, it can cause lysis of a
clot by destroying many of the clotting factors, thereby sometimes even
causing hypocoagulability of the blood.
Activation of Plasminogen to Form Plasmin: Then Lysis of Clots. When
a clot is formed, a large amount of plasminogen is trapped in the clot along
with other plasma proteins. This will not become plasmin or cause lysis of
the clot until it is activated. The injured tissues and vascular endothelium
very slowly release a powerful activator called tissue plasminogen
activator (t-PA) that a few days later, after the clot has stopped the
bleeding, eventually converts plasminogen to plasmin, which in turn
removes the remaining unnecessary blood clot. In fact, many small blood
vessels in which blood flow has been blocked by clots are reopened by this
mechanism. Thus, an especially important function of the plasmin system
is to remove minute clots from millions of tiny peripheral vessels that
eventually would become occluded were there no way to clear them,
Blood disease
1.Decreased Prothrombin, Factor VII, Factor IX, and Factor X Caused by
Vitamin K Deficiency.
With few exceptions, almost all the blood-clotting factors are formed by
the liver. Therefore, diseases of the liver such as hepatitis, cirrhosis, and
acute yellow atrophy can sometimes depress the clotting system so greatly
that the patient develops a severe tendency to bleed. Another cause of
depressed formation of clotting factors by the liver is vitamin K deficiency.
Vitamin K is necessary for liver formation of five of the important clotting
factors: prothrombin, Factor VII, Factor IX, Factor X, and protein C. In the
absence of vitamin K, subsequent insufficiency of these coagulation factors
in the blood can lead to serious bleeding tendencies.

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2. Hemophilia
Hemophilia is a group of sex-linked inherited blood disorders
characterized by prolonged clotting time. In this disorder, males are
affected and the females are the carriers. Because of prolonged clotting
time, even a mild
Types of hemophilia
Depending upon the deficiency of the factor involved, hemophilia is
classified into three types:
i. Hemophilia A or classic hemophilia that is due to the deficiency of factor
VIII. 85 percent of people with hemophilia are affected by hemophilia A.
ii. Hemophilia B or Christmas disease which is due to the deficiency of
factor IX. 15 percent of people with hemophilia are affected by hemophilia
B.
iii. Hemophilia C which is due to the deficiency of factor XI. It is a very
rare blood disorder.

3. Purpura
It is a disorder characterized by prolonged bleeding time. However, the
clotting time is normal. The characteristic feature of this disease is
spontaneous bleeding under the skin from ruptured capillaries. It causes
small tiny hemorrhagic spots under the skin which are called purpuric spots
(purple colored patch like appearance). That is why this disease is called
purpura.

Types and causes of purpura

The purpura is classified into different types depending upon the causes.
Thrombocytopenic purpura
Thrombocytopenic purpura is due to the deficiency of platelets
(thrombocytopenia). In bone marrow disease, platelet production is
affected leading to deficiency of platelets.
Idiopathic thrombocytopenic purpura Purpura due to some unknown cause
is called idiopathic thrombocytopenic purpura. It is believed that platelet
count decreases due to the development of antibodies against platelets,
which occurs after blood transfusion.

Thrombasthenia purpura
It is due to structural or functional abnormality of platelets. However, the
platelet count is normal. It is characterized by normal clotting time, normal
or prolonged bleeding time but defective clot retraction.

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4. von Willebrand Disease

von Willebrand disease is a bleeding disorder characterized by excess
bleeding even with a mild injury. It is due to inherited deficiency of von
Willebrand factor which is a protein secreted by endothelium of damaged
blood vessels and platelets. This protein is responsible for adherence of
platelets to endothelium of blood vessels during hemostasis after an injury.
It is also responsible for the survival and maintenance of factor VIII in
plasma.
The deficiency of von Willebrand factor suppresses platelet adhesion. It
also causes deficiency of factor VIII. This results in excess bleeding which
resembles the bleeding that occurs during platelet dysfunction or
hemophilia.

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