The Blood and Hemostasis and Blood Coagulation

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The Blood
◘ The Primary Function of Blood:
• to supply oxygen and nutrients as well as constitutional elements to
tissues and to remove waste products.
• Blood also enables hormones and other substances to be transported
between tissues and organs.
• Problems with blood composition or circulation can lead to downstream
tissue malfunction.
• Blood is also involved in maintaining homeostasis by acting as a
medium for transferring heat to the skin and by acting as a buffer
system for bodily pH.
◘ Gas exchange
♦ Oxygen (O2)
• O2 is the most immediate need of every cell and is carried throughout
the body by the blood circulation.
• Oxygen is used at the cellular level as the final electron acceptor in the
electron transport chain (the primary method of generating ATP for
cellular reactions).
• Oxygen is carried in the blood bound to hemoglobin molecules within
red blood cells.
• Hemoglobin binds oxygen when passing through the alveoli of the
lungs and releases oxygen in the warmer, more acidic environment of
bodily tissues, via simple diffusion.

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♦ Carbon dioxide (CO2)
• CO2 is removed from tissues by blood and released into the air via the
lungs.
• Carbon dioxide is produced by cells as they undergo the processes of
cellular respiration.
• Most of the carbon dioxide combines with water and is carried in the
plasma as bicarbonate ions.
• An excess of carbon dioxide (through exercise, or from holding ones
breath) quickly shifts the blood pH to being more acidic (acidosis).
• Chemoreceptors in the brain and major blood vessels detect this shift
and stimulate the breathing center of the brain.
• Hence, as CO2 levels build up and the blood becomes more acidic, we
involuntarily breathe faster, thus lowering CO2 levels and stabilizing
blood pH.
• In contrast, a person who is hyperventilating (such as during a panic
attack) will expire more CO2 than being produced in the body and the
blood will become too alkaline (alkalosis).
♦ Blood composition
• Blood is a circulating tissue composed of fluid plasma and cells (red
blood cells, white blood cells, platelets).
• Anatomically, blood is considered a connective tissue, due to its origin
in the bones and its function.
• Blood is the means and transport system of the body used in carrying
elements (e.g. nutrition, waste, heat) from one location in the body to
another, by way of blood vessels.

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• Blood is made of two parts:
1. Plasma which makes up 55% of blood volume.
2. Formed cellular elements (red and white blood cells, and platelets) which
combine to make the remaining 45% of blood volume.
♦ Plasma make up of
• 90% water
• 7-8% soluble proteins (albumin maintains bloods osmotic integrity,
others clot, etc)
• 1% electrolytes
• 1% elements
• 1% is salt, which helps with the pH of the blood.

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◘ Plasma Proteins
♦ Albumins
• are the most common group of proteins in plasma and consist of nearly
two-thirds of them (60-80%).
• They are produced in the liver.
• The main function of albumins is to maintain the osmotic balance
between the blood and tissue fluids and is called colloid osmotic
pressure.
• In addition, albumins assist in transport of different materials, such as
vitamins and certain molecules and drugs (e.g. bilirubin, fatty acids, and
penicillin).
♦ Globulins
• are a diverse group of proteins, designated into three groups: gamma,
alpha, and beta.
• Their main function is to transport various substances in the blood.
• Gamma globulins assist the body's immune system in defense against
infections and illness.
♦ Clotting proteins
• are mainly produced in the liver as well.
• There are at least 12 substances, known as "clotting factors" that
participate in the clotting process.
• One important clotting protein that is part of this group is fibrinogen,
one of the main components in the formation of blood clots.

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• In response to tissue damage, fibrinogen makes fibrin threads, which
serve as adhesive in binding platelets, red blood cells, and other
molecules together, to stop the blood flow.
• Plasma also carries Respiratory gases; CO2 in large amounts (about
97%) and O2 in small amounts (about 3%), various nutrients (glucose,
fats), wastes of metabolic exchange (urea, ammonia), hormones, and
vitamins.
◘ Red blood cell (erythrocyte)
• also known as "RBC's, erythrocytes".
• RBC’s are formed in the myeloid tissue or
most commonly known as red bone marrow,
although when the body is under severe
conditions the yellow bone marrow, which is
also in the fatty places of the marrow in the
body will also make RBC’s.
• The formation of RBC’s is called erythropoiesis ( erythro/ red; poiesis /
formation).
• Red blood cells lose nuclei upon maturation, and take on a biconcave, dimpled,
shape.
• They are about 7-8 mm in diameter, a thickness of 2.5 mm at the thickest point
and 1 mm or less in the center.
• There are about 1000x more red blood cells than white blood cells.
• RBC's live about 120 days and do not self repair.
• RBC's contain hemoglobin which transports oxygen from the lungs to the rest of
the body, such as to the muscles, where it releases the oxygen load.
• The hemoglobin gets it's red color from their respiratory pigments.

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♦ Shape
• RBC'S have a shape of a disk that appears to be “caved in” or almost
flattened in the middle; this is called bi-concave.
• This bi-concave shape allows the RBC to carry oxygen and pass
through even the smallest capillaries in the lungs.
• This shape also allows RBCs to stack like dinner plates and bend as
they flow smoothly through the narrow blood vessels in the body.
• RBC's lack a nucleus (no DNA) and no organelles, meaning that these
cells cannot divide or replicate themselves like the cells in our skin and
muscles.
• RBC’s have a short life span of about 120 days, however, as long as our
myeloid tissue is working correctly, we will produce about 2-3 million
RBC's per second.
• That is about 200 billion a day! This allows us to have more to replace
the ones we lose.
♦ Concentration Of Red Blood Cells In The Blood
• In normal men, the average number of red blood cells per cubic
millimeter is 5,200,000 (±300,000);
• in normal women, it is 4,700,000 (±300,000).
• Persons living at high altitudes have greater numbers of red blood cells.

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♦ Quantity Of Hemoglobin In The Cells
• Red blood cells have the ability to concentrate hemoglobin in the cell
fluid up to about 34 grams in each 100 milliliters of cells.
• The concentration does not rise above this value, because this is the
metabolic limit of the cell’s hemoglobin-forming mechanism.
• Furthermore, in normal people, the percentage of hemoglobin is almost
always near the maximum in each cell.
• However, when hemoglobin formation is deficient, the percentage of
hemoglobin in the cells may fall considerably below this value, and the
volume of the red cell may also decrease because of diminished
hemoglobin to fill the cell.
◘ Production Of Red Blood Cells
♣ Areas of the Body That Produce Red Blood Cells.
• In the early weeks of embryonic life, primitive, nucleated red blood
cells are produced in the yolk sac.
• During the middle trimester of gestation, the liver is the main organ
for production of red blood cells, but reasonable numbers are also
produced in the spleen and lymph nodes.
• Then, during the last month or so of gestation and after birth, red
blood cells are produced exclusively in the bone marrow.

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◘ Stages of Differentiation of RBCs
♣ Erythropoietin Stimulates Red Cell Production, and Its Formation
Increases In Response To Hypoxia
• The principal stimulus for red blood cell production in low oxygen
states is a circulating hormone called erythropoietin, a glycoprotein with
a molecular weight of about 34,000.
• In the absence of erythropoietin, hypoxia has little or no effect in
stimulating red blood cell production.
• But when the erythropoietin system is functional, hypoxia causes a
marked increase in erythropoietin production, and the erythropoietin in
turn enhances red blood cell production until the hypoxia is relieved.

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♣ Role of The Kidneys In Formation of Erythropoietin
• In the normal person, about 90 percent of all erythropoietin is formed in
the kidneys; the remainder is formed mainly in the liver.
• It is not known exactly where in the kidneys the erythropoietin is
formed.
• One likely possibility is that the renal tubular epithelial cells secrete the
erythropoietin, because anemic blood is unable to deliver enough
oxygen from the peritubular capillaries to the highly oxygen-consuming
tubular cells, thus stimulating erythropoietin production.
• At times, hypoxia in other parts of the body, but not in the kidneys,
stimulates kidney erythropoietin secretion, which suggests that there
might be some non renal sensor that sends an additional signal to the
kidneys to produce this hormone.
• In particular, both norepinephrine and epinephrine and several of the
prostaglandins stimulate erythropoietin production.
• When both kidneys are removed from a person or when the kidneys are
destroyed by renal disease, the person invariably becomes very anemic
because the 10 percent of the normal erythropoietin formed in other
tissues (mainly in the liver) is sufficient to cause only one third to one
half the red blood cell formation needed by the body.

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◘ Main Component of RBCs
• The main component of the RBC is hemoglobin protein which is about 250
million per cell.
• The word hemoglobin comes from hemo meaning blood and globin meaning
protein.
• This is the protein substance of four different proteins: polypeptide globin chains
that contain anywhere from 141 to 146 amino acids
• Hemoglobin also is responsible for the cell’s ability to transport oxygen and
carbon dioxide.
• This hemoglobin + iron + oxygen interact with each other forming the RBC's
bright red color.
• You can call this interaction by product oxyhemoglobin.

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♦ Functions
• The main function is the transportation of oxygen throughout the body
and the ability of the blood to carry out carbon dioxide which is called
carbamino – hemoglobin.
• Maintaining the balance of blood is important.
• The balance can be measured by the acid and base levels in the blood.
• This is called pH.
• Normal pH of blood ranges between 7.35-7.45; this normal blood is
called Alkaline (less acidic then water).
• A drop in pH is called Acidic.
• This condition is also called Acidosis.
• A jump in pH higher than 7.45 is called "Alkalosis".
• To maintain the homeostasis (or balance,) the blood has tiny molecules
within the RBC that help prevent drops or increases from happening.
♦ Destruction
• Red blood cells are broken down and hemoglobin is released.
• The globin part of the hemoglobin is broken down into amino acid
components, which in turn are recycled by the body.
• The iron is recovered and returned to the bone marrow to be reused.
• The heme portion of the molecule experiences a chemical change and
then gets excreted as bile pigment (bilirubin) by the liver.
• Heme portion after being broken down contributes to the color of feces
and your skin color changing after being bruised.

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White Blood Cells
♦ Shape
• White blood cells are different from red cells in the fact that they are
usually larger in size 10-14 micrometers in diameter.
• White blood cells do not contain hemoglobin which in turn makes them
translucent.
• Many times in diagrams or pictures white blood cells are represented in
a blue color, mainly because blue is the color of the stain used to see the
cells.
• White blood cells also have nucleii, that are somewhat segmented and
are surrounded by electrons inside the membrane.

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Formation of WBC’s
• White blood cells (leukocytes) are also known as "WBC's".
• White blood cells are made in the bone marrow but they also divide in the blood and
lymphatic systems.
• They are commonly amoeboid (cells that move or feed by means of temporary projections,
called pseudopods (false feet), and escape the circulatory system through the capillary beds.
• They live for about 13-20 days.

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• Classified according to the presence or absence of granules and the staining
characteristics of their cytoplasm.
• Leucocytes appear brightly colored in stained preparations, they have a
nuclei and are generally larger in size than RBC’s.
♣ Granular leukocytes
- Neutrophils, eosinophils, basophils
♣Agranular leukocytes
- Lymphocytes and monocytes
♠ Frequency of WBCs in Human Blood
WBC % number/mm3
• neutrophil 54-69 2700-6900
• lymphocyte 25-33 1250-3300
• monocyte 3-7 150-700
• eosinophil 1-3 50-300
• basophil 0-0.75 0-75

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♣ Granuloctyes
♦ Neutrophils
• Stain light purple with neutral dyes.
• Granules are small and numerous—course
appearance
• Several lobes in nucleus.
• 65% of WBC count.
• Highly mobile/very active
• Diapedesis—Can leave blood vessels and enter tissue space.
• Phagocytosis (eater), contain several lysosomes (janitor)
♦ Eosinophils or Acidophils:
• Large, numerous granules
• Nuclei with two lobes
• 2-5% of WBC count
• Found in lining of respiratory and
digestive tracts
• Important functions involve protections
against infections caused by parasitic
worms and involvement in allergic
reactions
• Secrete anti-inflammatory substances in
allergic reactions
• Eosinophils leave capillaries and enter tissue
fluid
• Release histaminase, phagocytize antigen-
antibody complexes and effective against
certain parasitic worms

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♦ Basophils
• Least numerous-0.5-1%
• Diapedesis—Can leave blood vessels and enter
tissue space
• Contain histamine, serotonin, heparin—
inflammatory chemical
• Basophils leave capillaries and release granules
containing heparin, histamine and serotonin, at
sites of inflammation
• Intensify inflammatory reaction
• Involved in hypersensitivity reactions
(allergies)
♣ Agranulocytes
♦ Lymphocytes
• Smallest WBC
• Large nuclei/small amount of cytoplasm
• Account for 25% of WBC count
• Two types:
- T lymphocytes—attack an infect or cancerous cell,
- B lymphocytes—produce antibodies against specific
antigens (foreign body)
• Lymphocytes are the major soldiers of the immune system
– T cells – attack viruses, fungi, transplanted cells,
cancer cells and some bacteria, it develop in the
thymus.
– B cells – destroying bacteria and inactivating their
toxins, it develop in the bone marrow.
– Natural Killer (NK) cells – attack a wide variety of infectious microbes
and certain tumor cells.
– Lymphocytes originate in the bone marrow, but can proliferate in the
spleen, thymus and other lymphoid tissues.
– Often, large lymphocytes seen in the blood have been activated somewhere
in the body, and are traveling to sites of action.

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♦ Monocytes
• Largest of WBCs
• Dark kidney bean shaped nuclei
• Highly phagocytic
• Monocytes take longer to arrive but arrive
in larger numbers and destroy more
microbes
• Enlarge and differentiate into
macrophages
◘ Leukemia
• Leukemia is a cancer of the blood or bone marrow characterized by an
abnormal proliferation of blood cells, usually white blood cells
(leukocytes).
• It is part of the broad group of diseases called hematological neoplasms.
• Damage to the bone marrow, by way of displacing the normal marrow
cells with increasing numbers of malignant cells, results in a lack of
blood platelets, which are important in the blood clotting process.
• This means people with leukemia may become bruised, bleed
excessively, or develop pin-prick bleeds (petechiae).
• White blood cells, which are involved in fighting pathogens, may be
suppressed or dysfunctional, putting the patient at the risk of developing
infections.
• The red blood cell deficiency leads to anaemia, which may cause
dyspnea.
• All symptoms may also be attributable to other diseases; for diagnosis,
blood tests and a bone marrow biopsy are required.

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◘ Platelets
• Platelets (thrombocytes) are minute discs 1-
4 mm.
• Produced in the bone marrow by
fragmentation of the cytoplasm of
megakaryocytes.
• Normal count: 150,000-400,000/µL (250,000)
• Life span 7-10 days.
• Removed from circulation by tissue macrophage system mainly in
spleen.
• Thrombopoietin: major regulator of platelet production (produced by
liver and kidney).
• It increases no. & rate of maturation of megakaryocytes.
• Platelets do not have nuclei and cannot reproduce.
♣ The cell membrane of platelets contains:
• A coat of glycoprotein (receptors) that cause adherence to injured
endothelial cells and exposed collagen.
• Phospholipids, that plays an important role in blood clotting.
♦ Their cytoplasm contains:
(1) actin and myosin molecules, which are contractile proteins similar to
those found in muscle cells, and still another contractile protein,
thrombosthenin, 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);

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(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 (factor XIII); and
(6) a growth factor that causes vascular endothelial cells, vascular smooth
muscle cells, and fibroblasts to multiply and grow.
♣ Platelets secrete factors that:
• increase local platelet aggregation (e.g., Thromboxane A),
• enhance vasoconstriction (e.g., Serotonin),
• and promote blood coagulation (e.g., Thromboplastin).
♦ Hemostasis (coagulation or clotting)
• Hemostasis is the natural process of stopping blood flow or
loss of blood following an injury.
• (hemo = blood; stasis = standing).
♠ 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. Growth of fibrous tissue into the blood clot to close the hole in the
vessel permanently.
◘ 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.

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• 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.
1. Vascular spasm or vasoconsriction:
○ The trauma of the ruptured vessel wall causes the smooth muscle in the
wall to contract; this reduces the flow of blood from the ruptured vessel.
○ The contraction results from:
1. Local myogenic spasm,
2. Release of vasoconstrictors (TXA2 and serotonin) from platelets that
adhere to the walls of damaged vessels, and
3. Nervous reflexes.
2. Formation of a platelet plug:
• If the cut in the blood vessel is very small, the cut is often sealed by a
platelet plug, rather than by a blood clot.
♣ Mechanism:
- Platelet adherence
- Platelet activation
- Platelet aggregation
• This process results in a platelet plug that seals the injured area.
• If the injury is small, a platelet plug may be able to form and close it within
several seconds.

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• If the damage is more serious, the next step of blood clotting will take
place.
• Platelets contain secretory granules.
• When they stick to the proteins in the vessel walls, they degranulate, thus
releasing their products, which include ADP (adenosine diphosphate),
serotonin, and thromboxane A2.
3. A blood clot forms:
• If the platelet plug is not enough to stop the bleeding, the third stage of
hemostasis begins:
♣ the formation of a blood clot.
• First, blood changes from a liquid to a gel.
• At least 12 substances called clotting factors take part in a series of
chemical reactions that eventually create a mesh of protein fibers within
the blood.
• Each of the clotting factors has a very specific function.
• We will discuss just three of the substances here: prothrombin,
thrombin, and fibrin.
• Prothrombin and fibrinogen are proteins that are produced and
deposited in the blood by the liver.
• Prothrombin: When blood vessels are damaged, vessels and nearby
platelets are stimulated to release a substance called prothrombin
activator, which in turn activates the conversion of prothrombin, a
plasma protein, into an enzyme called thrombin.
• This reaction requires calcium ions.

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• Thrombin: Thrombin facilitates the conversion of a soluble plasma
protein called fibrinogen into long insoluble fibers or threads of the
protein fibrin.
• Fibrin: Fibrin threads wind around the platelet plug at the damaged
area of the blood vessel, forming an interlocking network of fibers and a
framework for the clot.
• This net of fibers traps and helps hold platelets, blood cells and other
molecules tight to the site of injury, functioning as the initial clot.
• This temporary fibrin clot can form in less than a minute, and usually
does a good job of reducing the blood flow.
• Next, platelets in the clot begin to shrink, tightening the clot and
drawing together the vessel walls.
• Usually, this whole process of clot formation and tightening takes less
than a half hour.
◘ Basic Mechanism Of Clotting

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♣ Prothrombin
• Is a plasma protein, an alpha2-globulin,
• Its molecular weight of 68,700.
• It is normal plasma concentration about 15 mg/dl.
• It is an unstable protein that can split easily into smaller compounds,
one of which is thrombin, which has a molecular weight of 33,700,
• Prothrombin is formed continually by the liver,
• Vitamin K is required by the liver for normal formation of prothrombin
as well as for formation of a few other clotting factors.
• Therefore, either lack of vitamin K or the presence of liver disease that
prevents normal prothrombin formation can decrease the prothrombin
level so low that a bleeding tendency results.
♣ 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.
• Liver diseases →↓fibrinogen → bleeding tendency.
• Large molecule, does not leak into interstitial fluid which has very poor
clotting property.
• When it leaks into interstitial space in pathological conditions.
Interstitial fluid clots.
• The use of adsorbent chemicals, such as zeolites, and other hemostatic
agents, are also being explored for use in sealing severe injuries
quickly.

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Hemostasis and Blood Coagulation
◘ Normal Coagulation Pathways Summary
♣ Intrinsic pathway clotting factors
Factor XII Factor IX
Factor VIII Factor XI
♣ Extrinsic pathway clotting factors
Tissue factor (TF)*
Factor VII
♣ Common pathway clotting factors
Factor X
Factor V
Factor II Prothrombin
Factor I Fibrinogen
◘ Prevention Of Intravascular Clotting
Endothelial surface factors
• The most important factor to prevent clotting in the normal vascular system is:
– Smoothness; prevents contact activation of the intrinsic clotting factor
– Glycocalyx a mucopolysaccarides adsorbed to the surfaces of the
endothelial cells which repels clotting factors & platelets
– Thrombomodulin, a protein bound to endothelial membrane binds with
thrombin; slow the clotting
– (Thrombomodulin + thrombin) complex activates a plasma protein C →
inactivate factor V and VIII

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♣ 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.
♣ Intravascular Anticoagulants
♦ Heparin
• is a powerful anticoagulant, but its concentration in the blood is
normally low
• A heteropolysaccharide
• Heparin is produced in small quantities by
- Mast cells in the pericapillary CT throughout the body,
specially abundant in lungs and liver
- Basophilic polymorphonuclear leucocytes in the blood
• Combines with antithrombin III → 100 – 1000 times increase in
antithrombin activity of antithrombin III
• (Antithrombin + heparin) complex → inactivation or removal of factor
IX, X, XI and XII
♦ Plasmin (fibrinolysis)
• Plasminogen (profibrinolysis), a plasma protein contain euglobulin,
when activated, becomes a substance called plasmin (or fibrinolysin).
• Plasmin is a proteolytic enzyme resembles trypsin, causes digestion of
fibrin, Fibrinolysis.
• Removes extra or unwanted minute clots in the blood vessels.
• Plasmin also causes lysis of other clotting factors like prothrombin,
factor V, VIII & XII
• So it acts as anticoagulant as well.

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♦ Clot Dissolution
1. Plasmin is formed from plasminogen - enzyme called activator (e.g.
enzymes from urine, tears, saliva or bacterial enzyme streptokinase)
2. Plasmin as an enzyme is involved in breaking down fibrin into soluble
fragments (fibrinolysis)
♦ Natural Anticoagulants
• Antithrombin III – inhibits factor X and thrombin
• Heparin from basophils and mast cells potentiates effects of
antithrombin III (together they inhibit IX, X, XI, XII and thrombin)
• Antithromboplastin (inhibits “tissue factors” – tissue thromboplastins)
• Protein C and S – activated by thrombin; degrade factor Va and VIIIa
♦ Conditions That Cause Excessive Bleeding In Human Beings
• Excessive bleeding can result from deficiency of any one of the many
blood-clotting factors.
• Three particular types of bleeding tendencies caused by
1. Vitamin K deficiency,
2. Hemophilia, and
3. Thrombocytopenia (platelet deficiency).

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◘ Diseases Of The Liver
• With few exceptions, almost all of the 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.
♦ 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.
• Vitamin K is continually synthesized in the intestinal tract by bacteria,
except in neonates before they establish their intestinal bacterial flora.
• However, in gastrointestinal disease, vitamin K deficiency often occurs
as a result of poor absorption of fats from the gastrointestinal tract.
• The reason is that vitamin K is fat-soluble and ordinarily is absorbed
into the blood along with the fats.
• One of the most prevalent causes of vitamin K deficiency is failure of
the liver to secrete bile into the gastrointestinal tract.
• Lack of bile prevents adequate fat digestion and absorption and,
therefore, depresses vitamin K absorption as well.

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◘ Why Vitamin K is injected before performing the surgical procedure?
• Thus, liver disease often causes decreased production of prothrombin
and some other clotting factors both because of poor vitamin K
absorption and because of the diseased liver cells.
• Because of this, vitamin K is injected into all surgical patients with liver
disease or with obstructed bile ducts before performing the surgical
procedure.
• Ordinarily, if vitamin K is given to a deficient patient 4 to 8 hours
before the operation and the liver parenchymal cells are at least one-half
normal in function, sufficient clotting factors will be produced to
prevent excessive bleeding during the operation.
◘ Hemophilia
• Hemophilia is a bleeding disease that occurs almost exclusively in
males.
• In 85% of cases, it is caused by an abnormality or deficiency of Factor
VIII; this type of hemophilia is called hemophilia A or classic
hemophilia.
• In the other 15% of hemophilia patients, the bleeding tendency is
caused by deficiency of Factor IX.
• Both of these factors are transmitted genetically by way of the female
chromosome (XX).
• Factor VIII has two active components, a large component with a
molecular weight in the millions and a smaller component with a
molecular weight of about 230,000.

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• The smaller component is most important in the intrinsic pathway for
clotting, and it is deficiency of this part of Factor VIII that causes
classic hemophilia.
• Another bleeding disease with somewhat different characteristics, called
von Willebrand’s disease, results from loss of the large component.
♣ Hemophilia A (lack of F VIII; 85%)
• Spontaneous or traumatic subcutaneous
bleeding
• Blood in the urine
• Bleeding in the mouth, lips, tongue
• Bleeding to the joints, CNS, gastrointestinal
tract
♣ Thrombocytopenia
• Severe reduction in the number of PLTs
• this causes spontaneous bleeding as a reaction to minor
trauma, the bleeding is usually from many small
venules or capillaries, rather than from larger vessels
as in hemophilia.
• in the skin-reddish-purple blotchy rash giving the disease the name
thrombocytopenic purpura.
• Ordinarily, bleeding will not occur until the number of platelets in the
blood falls below 50,000/ml, rather than the normal 150,000 to
300,000.
• Levels <10,000/ml are frequently lethal.
◘ Anticoagulants For Clinical Use
In some thromboembolic conditions, it is desirable to delay the
coagulation process.
- Various anticoagulants have been developed for this purpose.
The ones most useful clinically are heparin and the
coumarins.

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Heparin
• Is extracted from several different animal tissues and prepared in almost
pure form.
• Causes the blood-clotting time to increase from a normal of about 6
minutes to 30 or more minutes.
• Furthermore, this change in clotting time occurs instantaneously,
thereby immediately preventing or slowing further development of a
thromboembolic condition.
• The action of heparin lasts about 1.5 to 4 hours. The injected heparin is
destroyed by an enzyme in the blood known as heparinase.
• Given intravenously or subcutaneously, and can be used in pregnant
women.
◘ Mechanism Of Action Heparin
• Heparin binds to AT-III and causes a conformational change thereby
activating AT-III
• Heparin enhances the action of Antithrombin III (AT-III) (plasma
protease inhibitor) 1000 fold ↑ activity.
• Antithrombin III inhibits clotting factor proteases, Thrombin (IIa), IXa,
Xa, XIa and XIIa, by forming stable complexes.
• LMWH predominantly inhibit factor Xa.
• Heparins do not affect thrombin bound to fibrin or Xa bound to
platelets.

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◘ Anticoagulants For Clinical Use
Coumarin
• When given to a patient, the plasma
levels of prothrombin and Factors VII,
IX, and X, all formed by the liver,
begin to fall by blocking the action of
vitamin K.
• After administration, the coagulant
activity of the blood decreases to about 50% of normal by the end of 12
hours and to about 20% of normal by the end of 24 hours.
• The coagulation process is not blocked immediately but must await the
natural consumption of the prothrombin and the other affected
coagulation factors already present in the plasma.
• Normal coagulation usually returns 1 to 3 days after discontinuing
coumarin therapy.
◘ Prevention Of Blood Coagulation Outside The Body
• 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.
• Heparin can be used for preventing coagulation of blood outside the
body as well as in the body.

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• Various substances that decrease the concentration of calcium ions in
the blood can also be used for preventing blood coagulation outside the
body. e.g. calcium oxalate.
• 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.
• 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.
• 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 convulsive death.

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Blood Coagulation Tests
♦ Bleeding time
• This is a test that measures the speed in which small blood
vessels close off (the condition of the blood vessels and
platelet function).
• This test is useful for detecting bleeding tendencies.
• The bleeding stops within 1 to 6 minutes.
• The time depends largely on the depth of the wound and the degree of
hyperemia in the finger or ear lobe at the time of the test.
• Lack of any one of several of the clotting factors can prolong the bleeding
time, but it is especially prolonged by lack of platelets.
♦ Abnormal Bleeding Time
• Prolonged bleeding time may indicate:
 A vascular (blood vessel) defect
 A platelet function defect (see platelet aggregation)
 Platelets count defect (low platelets)
• Drugs that may increase times include dextran, indomethacin, and
salicylates (including aspirin).

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♦ Bleeding time
♣ Procedure:
• Clean the earlobe with an alcohol
• Prick the earlobe with an automatic lancet
• Note the time when blood first appears on the
skin
• After half a minute (30sec) place the edge of the filter paper on the
top of the drop of blood.
• Perform the operation at half minute (30 sec) interval
• The end point or bleeding time is the first half minute when no blood
is seen on the filter paper.
♦ Clotting Time
• The time taken for blood to clot mainly reflects
the time required for the generation of
thrombin.
• The surface of the glass tube initiates the
clotting process.
• This test is sensitive to the factors involved in
the intrinsic pathway.
• The expected range for clotting time is 6-10 mins.
♣ Procedure:
• Collect blood in a chemically clean glass test tube and then to tip the
tube back and forth about every 30 seconds until the blood has
clotted.
• By this method, the normal clotting time is 6 to 10 minutes.
• Procedures using multiple test tubes have also been devised for
determining clotting time more accurately.

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♦ Prothrombin Time
• It gives an indication of the concentration of prothrombin in the
blood.
• Blood removed from the patient is immediately oxalated (or citrated)
so that none of the prothrombin can change into thrombin.
• Then, a large excess of calcium ion and tissue factor is quickly mixed
with the oxalated blood.
• The excess calcium nullifies the effect of the oxalate, and the tissue
factor activates the prothrombin-thrombin reaction by means of the
extrinsic clotting pathway.
• The time required for coagulation to take place is known as the
prothrombin time.
• The shortness of the time is determined mainly by prothrombin
concentration.
• The normal prothrombin time is about 12 seconds.
♠ Prothrombin Time (PT)
• Normal 11 -15 Sec
• Evaluates Extrinsic System (VII, X, V, II,
Fibrinogen)
• Prolonged PT indicates a deficiency in
any of factors VII, X, V, prothrombin
(factor II), or fibrinogen (factor I).
• Prolonged PT:
- A vitamin K deficiency (vitamin
K is a co-factor in the synthesis
of functional factors II (prothrombin), VII, IX and X)
• liver disease
• Warfarin therapy
• DIC
• excesive heparin

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♦ Thrombin Time (TT)
• Normal: 14-15 Sec
• Prolonged TT:
• Heparin (much more sensitive to heparin than aPTT)
• Hypofibrinogenemia
Tests of clotting factors