WINDSOR UNIVERSITYSCHOOL OF MEDICINE

BloodComponent and Function

Dr.Vishal Surender.MD.

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OBJECTIVES: BLOOD COMPOSITION

• Describe the components of blood (cells, ions, proteins, platelets) giving their normal values. Describe the physico-chemical properties of the plasma.

• Define the term hematocrit, state its normal values. Explain the importance of maintaining normal hematocrit level. List the factors that affect the hematocrit value. Understand how dehydration, excessive water intake, RBC count and size affect Htc. Be able to calculate plasma volume for given Htc and total blood volume for given Htc and plasma volume.

• Describe the functions of plasma electrolytes.

• List the main fractions of plasma proteins and describe their properties and functions. Describe the reasons and consequences of hypoproteinemia. Understand why liver diseases with decreased synthesis of albumins cause edema formation. What is the meaning of the term ‘non-protein nitrogen’?

BODY FLUID

Cells Interstitial Fluid (10L)Osmotic

forces

Starlings forces

Transcellular fluid = 1L

BLOOD

Red

cel

l mas

s =

2L

TOTAL BODY WATER = 42L

ICF = 28L ECF = 14L

BLOOD

Pla

sma

= 3

L

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THE COMPONENTS OF BLOOD

Blood is an opaque, red liquid consisting of several types of cells suspended in a complex, amber fluid known as plasma.When blood is allowed to clot or coagulate, the suspending medium is referred to as serum.

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BLOOD FUNCTIONS• Transportation

– Dissolved or chemically bound matter (O2, CO2, nutrients, metabolites)– Heat for heating and cooling (maintenance of the body temperature)

• Protection/Immunity– Defense against foreign agents (specific and non-specific immunity)

• Hemostasis– Prevention of hemorrhage - hemostasis

• Regulation/Hemostasis– Transmission of signals (hormones)– Maintenance of the homeostasis

• Concentration of dissolved substances• Osmotic and oncotic pressure• Acid-base balance - buffering the body fluids (maintenance of pH)

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BLOOD VOLUME AND COMPOSITION

1. Blood plasma - non-cellular portion of the blood - 55%

2. Formed elements (45 %): a. Red blood cells, RBC (erythrocytes) -99% of formd. Elmts. b. Platelets (thrombocytes) c. White blood cells, WBC (leukocytes)

Blood volumeMales: 5000 – 6000 mLFemales: 4000 – 5000 mL 6 - 8% of the total body mass 20% of the ECF.

Composition

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Blood Composition

Blood

Plasma

HEMATOCRIT (Htc)

Note that hematocrit is also known as Packed Cell Volume (PCV) orErythrocyte Volume Fraction (EVF)

Composition of Blood Plasma

Figure 16-1 (1 of 2)

Water

ions

Organic molecules

Trace elements and vitamins

Gases

Amino acids

Proteins

Glucose

Lipids

Nitrogenous wastes

CO2

O2

PLASMAis

composedof

such as

Albumin

Globulin

Fibrinogensuch as

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BLOOD PLASMAPhysico-chemical properties

• Temperature: 38oC• pH: 7.35 (venous) – 7.45 (arterial)• Relative whole blood viscosity - the internal friction of the blood (viscosity of water is

1.0): 3.5 – 5.4 • Relative plasma viscosity: 1.9 – 2.6• Erythrocyte sedimentation rate- 2 to 8 mm/hr.

Composition

• Water: 90-92%– Functions

• Solvent & suspending medium for blood components• Absorbs, transports and release heat

• Electrolytes (Na+, K+, Ca2+, Fe3+, Mg2+, Cl-, HCO3-, HPO4

2-, SO42-etc.) dissolved in water: 1%

– Functions• Create & maintain plasma osmotic pressure - 290 mOsm/L• Essential role in cell functioning (i.e., electrical properties of the blood cells)

• Maintenance of the acid-base homeostasis*-• is a steady state that provides an optimal internal environment for cell function

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BLOOD PLASMA: COMPOSITION (cont.)

• Organic substances (plasma proteins, nutrients, metabolites and waste products, regulatory substances, etc.) – 7-9%– Plasma proteins – proteins confined to the blood– Nutrients - products of digestion (AA, glucose, FA, glycerol, vitamins,

minerals)• Are transported by the blood for distribution in the body

– Waste products – breakdown products of protein metabolism (i.e., urea, uric acid, creatine, creatinine, bilirubin and ammonia)

• Are transported by the blood for excretion from the body– Regulatory substances (i.e., hormones, enzymes, vitamins)

• Dissolved gases: O2, CO2, N2

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PLASMA PROTEINS• 7-9% of the plasma, 65-90 g/L

• Are synthesized by the liver (with the exception of γ-globulins)

• Fractions– Albumins – plasma

concentration – 45 g/L– Globulins (α, β, γ) – 27 g/L– Fibrinogen – 3 g/L

• Albumins have the smallest molecular mass whereas fibrinogen is the largest

The least negatively-charged serum

proteins

The most charged proteins

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GENERAL FUNCTIONS OF PLASMA PROTEINS

• Create colloid osmotic pressure (25 mmHg) → retaining water within the capillaries

• Binding and transport of hormones, enzymes, lipids, vitamins, metals, bilirubin, drugs, etc.

• Contribution to the blood viscosity

• Buffer properties – capability of accepting both H+ and base ions

• Protection of body against microorganisms and toxic substances

• Mediate blood coagulation

• Precursors of some hormones (angiotensinogen, erythropoietin)

• Protein reserve – source of AA for tissues in case of starvation

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PROPERTIES & FUNCTIONS OF VARIOUS FRACTIONS OF PLASMA PROTEINS

• Albumins– 60% of the total plasma protein– High concentration and small size → 80% of the total colloid osmotic

P of the plasma

• 1 globulins (glycoproteins)– Transport of glucose and lipids– Include anti-protease

• 2 globulins– Carriers for different substances (high affinity, low binding capacity)

• Ceruloplasmin – copper• Thyroxin-binding protein• Transcobalamin – Vit B12• Bilirubin binding globulin• Transcortin – cortisol, etc.

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PROPERTIES & FUNCTIONS OF VARIOUS FRACTIONS OF PLASMA PROTEINS (cont.)

• -globulins– Carriers for lipids (lipoproteins), polysaccharides and metals (i.e.,

transferrin – iron and cupper)

• -globulins– Are immunoglobulins (antibodies)– Quantity and composition fluctuate

• ↑ in almost all diseases (inflammation and infections)

• Fibrinogen– A dissolved precursor of fibrin – blood clotting– Serum – plasma without fibrinogen (and clotting factors)

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HYPOPROTEINEMIA• ↓ blood level of proteins

• Results from – Malnutrition– Liver diseases (depression of protein synthesis)– Intestinal disease (malabsorption)– Kidney diseases (lost of albumins in urine)

• Results in – ↓ plasma oncotic pressure (especially due to ↓ albumin

concentration) and edema formation– Depression of specific functions (i.e., ↓ in globulins – ↓ resistance to

infections, ↓ in fibrinogen – defective blood clotting)

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NON-PROTEIN NITROGEN OF THE PLASMA

• Refers to nitrogen-containing substances other than proteins and AA (urea, uric acid, creatinine)

• ↑ in deranged kidney function

Plasma Proteins

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Some components of PlasmaClass Substance Normal Concentration Range

Cations Sodium (Na+) 136 – 145 mEq/L

Potassium (K+) 3.5 – 5.0 mEq/L

Calcium (Ca2+) 4.3 – 5.2 mEq/L

Magnesium (Mg2+) 1.5 – 2.0 mEq/L

Iron (Fe2+) 50 – 170 ug/dL

Copper (Cu2+) 70 – 155ug/dL

Hydrogen (H+) 35 – 45 mmoL/L

Anions Chloride (CL-) 95 – 105 mEq/L

Bicarbonate (HCO-3) 22 – 26 mEq/L

Lactate 0.67 – 1.8 mEq/L

Sulfate (SO42-) 0.9 – 1.1 mEq/L

Phosphate 3.0 – 4.5 mg/dL

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Some components of PlasmaClass Substance Normal Concentration Range

Protein Total 6 – 8 g/dL

Albumin 3.5 – 5.5g/dL

Globulin 2.3 – 3.5 g/dL

Fats Cholesterol 150 -220 mg/dL

Phospholipids 150 – 220 mg/dL

Triglycerides 35 – 160 mg/dL

Carbohydrates Glucose 70 – 110 mg/dL

Vitamins Vitamin B12 200 – 800 ug/mL

Vitamin A 0.15 – 0.6 ug/mL

Vitamin C 0.4 – 1.5 mg/dL

2,3 DPG 3 – 4 mmo/L

Transaminase (SGOT) 9 – 40 U/mL

Alkaline phosphatase 20 – 70 U/mL

Acid phosphatase 0.5 – 2 U/L

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Class Substances Normal Concentration Range

Other Substances Creatinine 0.6 – 1.2 mg/dL

Uric Acid 0.18 – 0.49 mmo/L

Blood Urea Nitrogen (BUN) 7 – 18 mg/dL

Iodine 3.5 – 8.0 ug/dL

CO2 23 – 30 mmol/L

Bilirubin (total) 0.1 – 1.0 mg/dL

Aldosterone 3 – 10nf/dL

Cortisol 5 – 18 ug/dL

Ketones 0.2 – 2.0 mg/dL

Some components of Plasma

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HEMATOCRIT (Htc)-Important Diagnostic Measurement

• Is the fraction of the blood volume made up of the formed elements (mainly RBC)

• Is determined by the centrifuging heparinised/anticoagulated blood in a standard calibrated tube of a small diameter

• When blood is allowed to clot or coagulate, the suspending

medium is referred to as serum

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(Htc)-Important Diagnostic Measurement

Plasma (55% of whole blood)

Erythrocytes (45% of whole blood)

Buffy coat: Leucocytes and Platelets <1% of whole blood

Formed Elements

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HEMATOCRIT

• Values– Males: 40 – 54 vol% (mean – 47%; 0.47)– Females: 38 – 46 vol% (mean – 42%, 0.42)

• ↑ in persons leaving at high altitudes, in dehydrated state, polycythemia, etc.

• ↓ in anemia, leukemia, bone marrow failure

• Importance– Determines blood viscosity– ↑ Htc → ↑ resistance to blood flow, load on the heart & BP

Determination of hematocrit values is a simple and important screening diagnostic procedure in the evaluation of hematological disease

The contribution of the WBC to hematocrit is only 0.08%. WBCs are lighter than the RBCs, they form a thin whitish layer between the sedimented RBCs and the plasma.

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Circulating Blood Cell LevelsBlood Cell Type Approximate Normal Range

Erythrocytes (cells/uL)

Men 4.3 – 5.9 x 1006

Women 3.5 – 5.5 x 1006

Leukocytes 4,500 – 11,000

Neutrophils 4,000 – 7,000

Lymphocytes 2,500 – 5,000

Monocytes 100 – 1,000

Eosinnophils 0 – 500

Basophils 0 – 100

Platelets 150,000 – 400, 000

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HEMATOCRIT

Normal Anemia Polycythemia

Tube A Tube B Tube C

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Blood composition: summary

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BLOOD 2RED BLOOD CELLS

JAUNDICEANEMIA & POLYCYTHEMIA

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CONTENT• RED BLOOD CELLS (RBC) COUNT, FUNCTIONS, STRUCTURE• HEMOGLOBIN (Hb): CHEMISTRY, REACTIONS, FUNCTIONS, CONCENTRATION• ERYTHROPOIESIS, CONTROL OF ERYTHROPOIESIS• DESTRUCTION OF RBC, METABOLISM OF Hb AND IRON. HEMOSIDEROSIS• JAUNDICE• ERYTHROCYTE SEDIMENTATION RATE• TYPES OF ANEMIA, SICKLE CELL DISEASE• POLYCYTHEMIA

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OBJECTIVES• Describe the functional consequence of the lack of a nucleus, ribosomes, and mitochondria for a)

protein synthesis and b) energy production within the red blood cell.• Relate the three red blood cell concentration estimates, red blood cell count, hematocrit, and

hemoglobin concentration.• Know the importance of MCV and be able to calculate the mean corpuscular volume.• Describe the structure of hemoglobin (Hb). Describe the differences between the major normal types

of Hb (adult A and A2, glucosilated, fetal). Predict the changes in Hb types present in blood when synthesis of beta chains of globin is deficient. Describe the abnormal types of Hb (Hb S, thalassemias). Describe the normal and abnormal Hb reactions (oxyHb, MetHb, carboxyHb). Calculate the mean corpuscular Hb concentration and the mean corpuscular Hb.

• Identify the site of erythropoietin production, the adequate stimulus for erythropoietin release, and the target tissue for erythropoietin action. Describe the role of vitamin B12 & folic acid, and various hormones in regulation of RBC formation. Describe the dietary requirements for RBC production. Relate the rate of red blood cell production and the percentage of immature reticulocytes in the blood.

• Describe the metabolism of iron in the body.• Describe the metabolism of Hb (pre-hepatic, hepatic, post-hepatic). • Describe the three types of jaundice (pre-hepatic, hepatic and post-hepatic). Compare and contrast the

laboratory findings and urine/stool color in the three types of jaundice.• Describe physiological jaundice of the newborn.• Discuss the normal balance of red blood cell synthesis and destruction, including how imbalances in

each lead to anemia or polycythemia. Compare and contrast the main types of anemia (nutritional, hemolytic, aplastic, hemorrhagic). Be able to describe different types of anemia in terms of MCV and MCHC. Describe the main effects of anemia and polycythemia on body functions.

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Blood Cells

red blood cells (erythrocytes)

white blood cells (leucocytes)

platelets (thrombocytes).

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Lymphocytes

Basophils

Eosinophils

Neutrophils

Mnoocytes

Cellular elements

White blood cells

Blood Cells

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RBC: Functions• Transport of O2 from the lungs to the tissues and CO2 in the opposite

direction– Hemoglobin– Carbonic anhydrase

• Catalyses the reaction H2O + CO2 ↔ H2CO3

• Maintenance of pH homeostasis (globin, phosphate and bicarbonate buffers)-hemoglobin in the cells is an excellent acid-base buffer

• Contribution to the blood viscosity

• ↓ blood oncotic P (by keeping Hb-protein inside the cells)

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RBC COUNT

• Normal values– Adult males: 4 600 000 – 6 200 000/mm3 (5.4million/mL)– Adult females: 4 200 000 – 5 400 000/mm3 (4.8million/mL)

• Abnormally high count – polycythemia

• Abnormally low count – anemia

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STRUCTURE OF THE MATURE RBC RBC - biconcave discs with

central depression on each sideSmall size Excess of the plasma

membrane & specific shape

High surface-to-volume ratio

Rapid diffusion of respiratory gases to and from the cell

Deformation of the cells without stretching the plasma membrane

Easy passage through the small capillaries

Figure 16-5

Red Blood Cells

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STRUCTURE OF THE MATURE RBC (cont.)

• Membrane contains special proteins and polysaccharides that differ from person to person – blood groups

• Lack of the nucleus and organelles– Cannot undergo mitosis– Generate ATP anaerobically → do not use oxygen they transport– Can not synthesize new cellular components to replace damaged ones

Life span - 120 days

• Contain a red pigment, hemoglobin (red color of the blood) – Occupies 1/3 of cellular volume– 280 million Hb molecules/RBC

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MEAN CORPUSCULAR VOLUME

• Mean volume of a RBC

• Values– Normal range 82 – 99 femtolitre (fL)– Low volume in microcytic anemia– High volume in macrocytic anemia

• Calculation of the MCVHematocrit x 10RBC count (in millions/mL blood)

•

MCV: 82-99 fL

fL= 10-15 L

Sample calculation: Htc = 40, RBC count = 5 (x 106/mL)

MCV = (40 x 10)/5 = 80 fl

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In a normal individual RBCs show minimal anisocytosis(Excessive variation inthe size of cells )and poikilocytosis(irregularly shaped erythrocytes).

Larger than average RBCs are macrocytic (left), while those smaller than average are microcytic (right).

RBC Morphology

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Pale cells (central pallor >1/3 dia) are referred to as hypochromic

(right), while cells without central pallor are called hyperchromic (left).

Normal peripheral blood RBCs are normochromic normocytic.

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RBC Morphology

Normal peripheral blood RBCs are

normochromic normocytic.

while cells without central pallor are called hyperchromic

Pale cells (central pallor >1/3 dia) are referred to as hypochromic

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Abnormality Associated condition(s)

Target Cells Sickle-cell/ThalassemiaIron-deficient anemiaHyposplenismLiver disease

Tear-drop poikilocytes Myelofibrosis

Spherocytes Hereditary spherocytosisAutoimmune hemolytic anemia

Basophilic stippling Lead poisoningThalassemia

Howell-Jolly bodies Hyposplenism

Heniz bodies G6PD deficiencyAlpha-thalassemia

Schistocytes (“helmet cells”) Intravascular hemolysisMechanical heart valveDisseminated intravascular coagulation

‘Pencil’ poikilocytes Iron deficiency anemia

Burr cells (echinocytes) UremiaPyruvate kinase deficiency

Pappenheimer bodies Hyposplenism

RBC Morphology

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Target cells

• Target cells (codocytes or leptocytes) have a "lump" of hemoglobinized cytoplasm within the area of normal central pallor, causing them to resemble a "bullseye" target

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Tear-drop poikilocytes

An abnormally shaped red blood cell with a single point or elongation.

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Spherocytes

• Spherocytes are red cells which have assumed the form of a sphere rather than the normal discoid shape. As a result, they appear on routine blood films as cells that are smaller and more dense than normal red cells of the species, and have a reduced area of central pallor

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Basophilic stippling

Basophilic stippling of erythrocytes (BSE) represents the spontaneous aggregation of ribosomal RNA in the cytoplasm of erythrocytes

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Howell-Jolly bodies

Howell-Jolly bodies in the blood of a (non-anemic) splenectomized dog. Howell-Jolly bodies (H-J) bodies are small fragments of non-functional nucleus which were not extruded as the cell left the marrow

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Heniz bodies

• Heinz bodies are inclusions within red blood cells composed of denatured hemoglobin.

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Schistocytes

• Schistocytes, or red cell fragments, are generally taken to reflect mechanical injury to erythrocytes. A wide variety of forms may be observed.

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Burr cells (echinocytes)

• Burr cells (echinocytes) are spiculated RBCs. The term crenation is also used to refer to cells of this type. The projections of the cell membrane may be sharp or blunt, are usually numerous, and tend to be evenly spaced around the circumference.

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Pappenheimer bodies

• Pappenheimer bodies are basophilic erythrocytic inclusions that are usually located at the periphery of the cell.

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HEMOGLOBIN: Chemistry• Protein – globin

– 4 polypeptide chains• Normal adult Hb – HbA,

Hbα2β2– A pair of α chains (141 AA) – A pair of β chains (146 AA)

• Adult Hb – HbA2 (2.5% of Hb), Hbα2δ2 – β chains are replaced by δ chains

• Fetal Hb – HbF, Hbα2γ2 – β chains are replaced by γ chains

(146 AA) • Adult Hb glucosilated – HbA1c

– Has a glucose attached to each β chain (4% - 5.9%, 6.5%)

• Nonprotein pigment bound to each of the 4 chains – hem– Each hem ring has 1 iron ion (Fe2+) that

can combine reversibly to 1 O2 molecule– Each Hb molecule can bind 4 O2 molecules

Hb A 2α 2β

HbA2 2α 2δ

HbF 2α 2γ

Adult Hb – HbA, Hbα2β2

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SICKLE CELL DISEASE• Inherited disease• High prevalence in the

malaria belt • Mutation causes formation

of HbS instead of HbA– HbS precipitates into

long crystals when oxygen tension is low (hypoxia) → cell elongation (sickling) and damage to the cell membrane → hemolysis → hypoxia (vicious cycle)

Rigid sickled RBCs occlude the microvasculature leading to vaso-occlusive crisis.

Negatively charged glutamate is substituted for nonpolar valine at position 6 in the β chain)

HbS – HbαA2βS2

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HEMOGLOBIN: Reactions• Oxyhemoglobin: Hb + 4 O2 (O2 attaches to Fe2+ in hem)

– Is produced in the lungs (oxygen loading)

• Reduced Hb (deoxyHb) – Is produced in tissue capillaries after dissociation of O2 (oxygen unloading)– Combines with H+ - acts as a buffer– Combines with CO2 → Carbaminohemoglobin: Hb + CO2 (CO2 binds to globin, not to

hem)

OxyHb

NH-COO-

CO2 carrying function

Buffering function

COOH/COO-

NH2/NH3+

O2

carrying function

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HEMOGLOBIN (cont)

Oxyhemoglobin (HbO2), the oxygen-saturated form of

hemoglobin, transports oxygen from the lungs to tissues,

where the oxygen is released. When oxygen is released,

HbO2 becomes reduced hemoglobin (Hb). While oxygensaturated hemoglobin is bright red, reduced hemoglobin is bluish-red, accounting for the difference in the color of blood in arteries and veins.

The affinity of hemoglobin for O2 is affected by pH, temperature, and the concentration in the red cells of 2,3-bisphosphoglycerate (2,3-BPG). 2,3-BPG and H+ compete with O2 for binding to deoxygenated hemoglobin, decreasing the affinity of hemoglobin for O2 by shifting the positions of the four peptide chains (quaternary structure).

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HEMOGLOBIN: Reactions (cont.)• Methemoglobin (MetHb):

– Hb iron is oxidized from the ferrous (Fe2+) to the ferric state (Fe3+) – Is incapable of carrying O2 and has a bluish color → cyanosis – Limited amount of metHb can be converted back to Hb by methemoglobin

reductase present in the RBCs– In normal state, 1.5% of Hb is in MetHb state– Methemoglobinemia: Met-Hb > 1.5% (results from oxidation by nitrates,

drugs like phenacetin or sulfonamides and congenital deficiency of methemoglobin reductase).

• Carboxyhemoglobin: Hb + CO(carbon monoxide) → cherry-red color of the skin and mucous membranes– CO has 200-250 times the affinity to Hb as does O2 → HbCO is a very stable

molecule– CO ↓ the functional Hb concentration

• HbCO is unavailable for O2 transport → CO poisoning, acute onset anemia

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HEMOGLOBIN: Reactions (cont.)Certain chemicals readily block the oxygen-transporting function of hemoglobin. For example, carbon monoxide (CO) rapidly replaces oxygen in HbO2, resulting in the formation of the stable compound carboxyhemoglobin (HbCO). The formation of HbCO accounts for the asphyxiating properties of CO. Nitrates and certain other chemicals oxidize the iron in Hb from the ferrous to the ferric state, resulting in the formation of methemoglobin (metHb). MetHb contains oxygen bound tightly to ferric iron; as such, it is useless in respiration. Cyanosis, the darkblue coloration of skin associated with anoxia, becomes evident when the concentration of reduced hemoglobin exceeds 5 g/dL. Cyanosis may be rapidly reversed by oxygen if the condition is caused only by a diminished oxygen supply.However, cyanosis caused by the intestinal absorption of nitrates or other toxins, a condition known as enterogenous

cyanosis, is due to the accumulation of stabilized methemoglobin and is not rapidly reversible by the administration of oxygen alone.

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HEMOGLOBIN: Concentrations

RBC

Plasma

MCHC = Hb amount / Volume of packed RBCHb concentration = Hb amount (g)/Volume of whole blood (dL, L)

Concentration per unit volume of whole blood

Males – 16.0±2.0 g/dLFemales – 14.0±2.0 g/dL

Mean corpuscular Hb concentration - concentration of Hb per unit packed cell volume

Normal range: 31-37 g/dL packed cells↓ value – hypochromia (i.e., Hb deficiency)↑ value – hyperchromia (i.e., spherocytosis)

Calculation:MCHC = Hb concentration x 100

Htc

Sample calculation: [Hb] = 14.5 g/dL, Htc = 45 mL/dLMCHC = (14.5/45) x 100 = 32.2 g/dL packed cells

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Hb CONCENTRATION: Mean corpuscular Hb (MCH)

• Is the total Hb content of a RBC

• Values– Normal range – 27-31 pg– ↓ value – hypochromia (i.e., iron deficiency anemia)– ↑ value – hyperchromia (i.e., vit B12 deficiency)

• CalculationMCH = Hb in grams/100 mL blood x 10

RBC count in million/L blood

• Sample calculation: [Hb] = 12 g/dL, RBC count = 4 x 106/mLMCH = 12/4 x 10 = 30 pg

MCH

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RBC CHARACTERISTICS: SUMMARY

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Erythropoiesis

• Concept: The production of new red blood cells to replace the old and died ones

• In the adult, all the red cells are produced in bone marrow

61CFU：colony-forming unit

Erythropoiesis- Pluripotent stem cells

in the bone marrow can produce any type of

blood cells. is capable of both self-

replication and differentiation to committed precursor cells that can produce only a specific cell line.

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Proerythroblast(Pronormoblast)

BasophilicNormoblast

PolychromatophilicNormoblast

OrthochromatophilicNormoblast

Reticulocyte

Erythrocyte

Early Intermediate Late

Erythropoiesis-CPU-E the committed red cell precursor undergoes several

divisions. The daughter cells becomes progressively smaller, the cytoplasm changes color from blue to pink as

hemoglobin is synthesized, the nucleus becomes small and dense and then extruded.

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Proerythroblast(Pronormoblast)

BasophilicNormoblast

PolychromatophilicNormoblast

OrthochromatophilicNormoblast

Reticulocyte

Erythrocyte

Early Intermediate Late

Erythropoiesis-CPU-E

The resulting non-nucleated cells is termed a reticulocyte since it still contains RNA.

Within a few days of entering the circulation, the reticulocytes lose their RNA and becomes mature red cells

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Regulation of Erythropoiesis

A. Erythropoietin, a glycoprotein released predominantly from the

kidneys in response to tissue hypoxia. also produced by reticuloendothelial system of

the liver and spleen. Effect:

a, Stimulates the proliferation and differentiation of the committed red cell precursor

b, Accelerates hemoglobin synthesis c, Shortens the period of red cell development in the

bone marrow.

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CONTROL OF ERYTHROPOIESIS: Hypoxia

↓

↑

↑

Tissue oxygenation is the most powerful regulator of the RBC production (but not the RBC count in the blood)

Hypoxia stimulates production of EPO by the kidneys - the tubular epithelial cells and juxtaglomerular cells (90% of EPO) & the liver

Biological effects of EPO: 1. ↑ production of proerythroblasts from hematopoietic stem cells2. ↑ speed of erythropoietic stages

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ERYTHROPOIESIS

Appearance of Hb Some Hb is present in the early erythroblastsLate erythroblasts are saturated with Hb

Degeneration of the nucleus Starts in the late erythroblast stage Disappeared by the reticulocyte stage

Degeneration of the cell organelles

Progressive ↓ in the cell size

Reticulocytes enter the blood and within 1-2 days develop into mature RBC.Only mature RBC and reticulocytes are present in the blood

Morpho-functional changes (proerythroblast → RBC)

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RETICULOCYTES & ERYTHROPOIESIS RATE• Normal reticulocytes count in the blood

– 1-4% of the circulating RBC in adults – 2-6% in newborns

• ↑ reticulocytes count – indicator of rapid RBC

production (i.e., hypoxia, hemorrhage, stress, effective therapy of anemia)

• ↓ reticulocytes count - ↓ erythropoiesis (↓ EPO production, ↓ ability of red bone marrow to respond to EPO, nutritional anemia, etc.)

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CLINICAL FOCUS: BLOOD DOPING AND EPO

• Beneficial effects of EPO– ↑ RBC count and O2 carrying capacity of the blood → ↑ O2 delivery

to tissues, ↑ muscular performance, ↓ muscular fatigue – Recombinant EPO (rhEPO) is used for treatment of anemias

associated with chronic renal failure, AIDS and cancer chemotherapy

• Dangers of excessive EPO– Genetically engineered EPO (i.e., darbepoetin) has increased life time – ↑ Htc → ↑ blood viscosity, ↑ peripheral resistance, ↑ blood

pressure, ↓ heart rate (secondary to increased blood pressure), ↑ blood clotting

– Genetically engineered EPO often cause production of antibodies against natural EPO and destruction of the RBC

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CONTROL OF ERYTHROPOIESIS:Vitamin B12 and folic acid

• Are required for maturation of the RBC– ↑ Synthesis of DNA

(synthesis of thymidine triphosphate – DNA building block) → rapid proliferation of the erythroblastic cells

• Vitamin B12 (cyanocobolamin)– Is required for action

of folic acid on erythropoiesis

Dietary B12

Parietal/oxyntic cells of gastric mucosa produce intrinsic factor (IF)

B12 binds with the IF – protection from digestion by GIT secretions

Complex of Vit B12 +IF complex binds to the mucosal receptors in the ileum → transport across mucosa

Release of B12 into the portal blood freed of IF

Binding to the plasma globulins (transcobolamin I, II and III) → red bone marrow or storage in the liver (very large quantities – 3-4 years reserve)

B12+IF

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CONTROL OF ERYTHROPOIESIS: Other factors

• Testosterone – Stimulates the release of EPO

• Adrenal cortical steroids and ACTH – In physiological concentrations stimulate EPO production– Large doses are inhibitory

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DESTRUCTION OF THE RBC• Sites of destruction

– Circulating blood (10% of senescent RBCs)– Macrophage system (spleen and liver)

• Senescent RBC– ↓ metabolic rate → ↑ fragility → rupture of the membrane when

RBC pass through tight spots of the circulation (i.e., red pulp of the spleen)

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METABOLISM OF Hb• Prehepatic

– Takes places in the macrophages– Results in formation of bilirubin – a bile pigment

• Hepatic– Takes place in the liver (hepatocytes)– Conjugation of bilirubin with glucuronic acid – bilirubin mono- or bi-

glucuronide and secretion of conjugated bilirubin into the bile

• Posthepatic– Takes place in the GI and kidneys– Formation of urobilinogen and stercobilinogen and excretion

PREHEPATIC METABOLISM OF Hb

Protein pool

RBC or remnant

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HemoglobinCell remnant

Hem Globin

Fe++Pigment

CO Biliverdin

BilirubinExhaled

Albumin

Bilirubin-albumin

Liver

BLOOD

Fe++

Macrophages

74

HEPATIC & POSTHEPATIC METABOLISM OF BILIRUBIN • In the liver

– Replacement of albumin with glucuronic acid – bilirubin mono- or bi-glucuronide (water soluble)

– Excretion of conjugated bilirubin into the small intestine via the bile

• In the small intestine– Conversion of bilirubin to

urobilinogen by the intestinal bacteria

• Conversion to stercobilinogen → oxidation and excretion in the feces as stercobilin

• Absorption from the small intestine & either re-excretion by the liver or oxidation & excretion by the kidneys as urobilin.

Liver

Glucuronic acid Albumin

Bilirubin-glucuronide

Urobilinogen (in the small intestine)

Reabsorption Stercobilinogen

Re-excretion in bile

Excretion as urobilin in urine

Excretion as stercobilin in feces

Transport of bilirubin from plasma into the hepatocytes

75

BILIRUBIN: Concentration in plasma

Bilirubin Concentration in plasma, mg/dL

Free bilirubin = unconjugated bilirubin

0.1 – 1

Conjugated bilirubin 0 – 0.3

Total bilirubin 0.3 – 1.2

76

JAUNDICERefers to the yellow color of the skin, conjunctivae and mucous membranes caused by the presence of excessive bilirubin in the plasma and body fluids (jaune (French) = yellow)

Blood bilirubin level must exceed three times the normal values, for the coloration to be easy visible

Types of jaundice:

Pre-hepatic – the pathology occurs prior to the liverHepatic – the pathology is located in the liverPost-hepatic – the pathology occurs after the conjugation of bilirubin in the liver

PRE-HEPATIC JAUNDICE

77

Cause:Excessive hemolysis of the RBCs – hemolytic jaundice

Capacity of the liver to conjugate bilirubin is exceeded (saturation of enzyme glucuronyl transferase)

↑ unconjugated (indirect) bilirubin

Normal conjugated (direct) bilirubin

urobilinogen formation urobilinogen → dark urine

↑ stercobilinogen → dark feces

Pigment

Biliverdin

Bilirubin

Blood

Albumin

Bilurubin-albumin

Liver

Bilurubin-glucuronide

Urobilinogen (small intestine)

Reabsorption Stercobilinogen

Re-excretion in bile

Excretion as urobilin in urine

Excretion as stercobilin in feces

N

Inc. bilirubin production

78

HEPATIC JAUNDICEResults from infective or toxic damage to the liver cells (hepatocellular damage)

Uptake, conjugation and/or excretion of bilirubin is affected

Normal/decreased conjugated bilirubin

↑ urobilinogen in blood (↓ enterohepatic circulation and hepatic extraction of blood urobilinogen by damaged hepatocytes)

Pale/N stool

↑ unconjugated bilirubin

Dark urine

↑ urobilinogen filtration and excretion in urine

79

POSTHEPATIC JAUNDICE

Results from obstruction of the bile ducts by stones, tumors, etc.

Functioning of the hepatic cells is normal N

plasma level of conjugated bilirubin due to the bile entry into the blood from ruptured congested canaliculi and ↑ total bilirubin

Normal unconjugated bilirubin

urobilinogen formation

↓ stercobilin content in feces → pale feces

↓ or absent urobilin in urine

Conjugated bilirubin in urine (kidney can excrete small quantities of highly soluble conjugated bilirubin) → dark urine

80

PHYSIOLOGICAL JAUNDICE OF THE NEWBORN

Hemolysis of the excess RBC when the infant is suddenly exposed to a high oxygen environment and hence does not need so many RBC as in the uterus

Immaturity of the liver (inability to conjugate significant quantities of bilirubin with glucuronic acid for excretion into the bile) to handle the excess bilirubin (especially in premature babies)

↑ plasma total bilirubin concentration (less than 1 mg/dL → 5 mg/dL)

Mild jaundice (yellowness) of the infant’s skin and the sclerae for 1-2 weeks

81

IRON METABOLISM1. Dissociation of Fe from the hem

→ plasma → binding to transferrin, transport in the blood →

2. Detachment from transferrin & storage in the liver, muscle cells & macrophages attached to ferritin or hemosiderin →

3. Release from the storage sites, transport in the blood by transferrin

4. Transport into the RBC precursor cells by receptor mediated endocytosis → Hem synthesis

↓ quantities of transferrin → ↓ Hb content in the RBC – hypochromic anemia

1

2

3

4

Synthesis of transferrin increases with iron deficiency but decreases with any type of chronic disease.

82

FORMS OF IRON IN THE BODY• Recommended daily intake - 15 – 18 mg (250-330 μmol)• Minimal absorption to balance iron loss

– Adult males - 35 μmol – Adult females - 175 μmol

• Distribution of body iron in an average man – Hb, 2100 mg – Ferritin - water soluble protein-iron complex , 700 mg (in the liver,

spleen, marrow and plasma)– Hemosiderin - water insoluble complex (macrophages of the liver

and bone marrow), 300 mg – Myoglobin - local oxygen reserve, 200 mg– Tissue (heme and nonheme) enzymes, 150 mg– Transport-iron compartment in plasma (transferrin), 3 mg.

83

HEMOCHROMATOSIS• Reasons

– Primary - one of the most common autosomal recessive genetic disorders characterized by excessive absorption of dietary iron resulting in a pathological increase in total body iron stores

• Failure to reduce iron reabsorption in response to increased iron level in the body– Secondary – is not genetic (results from anemia, alcoholism, transfusion iron overload –

hemosiderosis, etc.)

• Consequences– Deposition of iron in the body tissues (liver, heart, pancreas, pituitary, joints, and skin)

initially as ferritin and than as hemosiderin – Toxic action on organs and damage of cells due to action as a pro-oxidant (↑ formation

of free radical formation, i.e., the hydroxyl radical and the superoxide radical) → DNA cleavage, impaired protein synthesis, and impairment of cell integrity and cell proliferation, leading to cell injury and fibrosis.

• Cirrhosis, hyperpigmentation of skin, diabetes mellitus, impotence, joint diseases, etc.

84

ERYTHROCYTE SEDIMENTATION RATE (ESR) • Specific weight of the RBC is higher than that of the

plasma in a stabilized blood, RBC slowly sink towards the bottom of the test tube -sedimentation

• Factors increasing ESR– ↓ Htc, ↓ blood viscosity– ↑ concentration of fibrinogen (i.e., pregnancy,

vascular diseases, heart diseases), haptoglobulin, lipoproteins, immunoglobulins

– Macrocytic RBC– Extreme elevation of WBC count (leukemia)

• Factors decreasing ESR– ↑ Htc – Change in the RBC shape (i.e., sickle-cell anemia,

poikilocytosis – nonuniformity of shape)– ↑ albumin concentration

ESR

Clumps of RBCs

Males – 3-6 mm/hFemales – 8-10 mm/h

85

ANEMIA• Deficiency of blood Hb due to

– ↓ RBC count (too rapid loss or/and too slow production) – ↓ Hb quantity in the RBC

WHO's Hemoglobin thresholds used to define anemia (1 g/dL = 0.6206 mmol/L)

Age or gender group Hb threshold (g/dl)

Hb threshold (mmol/l)

Children (0.5-5.0 yrs) 11,0 6,8

Children (5-12 yrs) 11,5 7,1

Children (12-15 yrs) 12,0 7,4

Women, non-pregnant (>15yrs)

12,0 7,4

Women, pregnant 11,0 6,8

Men (>15yrs) 13,0 8,1

86

ANEMIA: CONSEQUENCES• ↓ oxygen-carrying capacity

of the blood → hypoxia → vasodilation

• ↑ in pulse and respiratory rates (effort to supply sufficient oxygen to tissues)

• ↓ exercise & cold tolerance

• Pale skin (↓ red colored oxyHb)

• ↑ fatigue and lassitude• ↓ blood viscosity → ↓

peripheral vascular resistance → ↑ blood flow, venous return, cardiac output and work load on the heart

Eyes- yellowing

87

ANEMIAS: ClassificationsClassification

according to etiological ground

• Nutritional• Aplastic• Hemorrhagic• Hemolytic

Anemia: classification according to MCV     

Macrocytic anemia (MCV>100) 

Normocytic anemia (80<MCV<100) 

Microcytic anemia (MCV<80)   

Hem synthesis defect (i.e., iron deficiency, chronic diseases) Globin synthesis defect (i.e., thalassemia) Sideroblastic defect

Deficiency of vit B12, folic acid, or IF. Hypothyroidism. Alcoholism. Liver diseases. Drugs that inhibit DNA replication (i.e., methotrexate, zidovudine)

Acute blood loss, chronic diseases, bone marrow failure, hemolysis

88

ANEMIA: NutritionalIron deficiency • Is the most common type• Reasons

– Premenopousal women: Blood loss during menses (20% of all women of childbearing age have iron deficiency anemia, compared with only 2% of adult men)

– Males and postmenopausal females: Excessive iron loss due to chronic occult bleeding (peptic ulcer, tumor, etc.)

– Increased iron demands (i.e., pregnancy and lactation)– Inadequate iron intake or absorption (i.e., vit. C deficiency)– Parasitic infestation (hookworm, amebiasis, schistosomiasis)– Chronic intravascular hemolysis (if the amount of iron released during

hemolysis exceeds the plasma iron-binding capacity)

89

IRON DEFICIENCY ANEMIA: CONSEQUENCES• Low serum ferritin (serum iron) level

– Plasma ferritin concentration is an excellent indicator of the iron stored in the body, because of a dynamic balance between intra- and extracellular ferritin iron

• ↓ bone marrow iron stores (ferritin and hemosiderin)• ↓ saturation of transferrin• ↓ RBC count & Htc • RBC are small and look pale - microcytic hypochromic anemia• Abnormal fissuring of the angular (corner) sections of

the lips (angular stomatitis). • Abnormal craving to eat substances (eg, ice, dirt,

paint).

90

DEFICIENCY OF IRON UTILIZATION: SIDEROBLASTIC ANEMIA

• Inadequate marrow utilization of iron for Hb synthesis despite the presence of adequate or increased amounts of iron

• Reasons: Hereditary or acquired, including lead and ethanol toxicity, pyridoxine deficiency

• Deficient reticulocyte production, intramedullary death of RBCs, and bone marrow erythroid hyperplasia (and dysplasia)

• Presence of polychromatophilic, stippled RBCs (siderocytes)

• Hipochromic, microcytic RBCs, variations in RBC size Ring sideroblasts are erythroid

precursors whose mitochondria (located around the nucleus) are loaded with nonheme iron.

91

ANEMIA: Nutritional (cont.)Deficiency of vitamin B12 and/or folic acid

• Reasons – Inadequate intake (a strict vegetarian diet excluding all meat, fish, dairy products, and eggs;

chronic alcoholism) – Inadequate GI absorption

• Lack of IF - pernicious anemia– Autoimmune destruction of parietal cells (atrophic gastric mucosa) or AB against IF – Removal of the functional portion of the stomach, such as during gastric bypass

surgery• Crohn's disease intestinal malabsorption disorders• Resection (or inflammation) of the ileum (site of B12 reabsorption)

• Consequences– Maturation failure

• Failure of DNA synthesis with preserved RNA synthesis, which result in restricted cell division of the progenitor cells.

• Production of large precursor cells – megaloblasts and larger irregular oval erythrocytes – macrocytes fully saturated with Hb – macrocytic (megaloblastic) anemia

• ↑ fragility of the plasma membrane → ↓ life span → anemia– Vitamin B12 deficiency only results in peripheral neuropathy and spinal

cord degeneration

92

ANEMIA: Hemorrhagic

• Results from abnormal blood loss (mild or severe; acute or chronic)– Replacement of lost fluid within 1 – 3 days (much faster than

the replacement of lost RBC) → dilution of the RBC

• Is normocytic

• Prolonged but mild loss of the blood causes microcytic hypochromic anemia (iron deficiency)

93

ANEMIA: Aplastic

• Results from suppression or destruction of the bone marrow (i.e., overexposure to ionizing radiation, adverse drug reaction, toxic chemicals, severe infections)

• Is usually normocytic

• Panhypoplasia of the marrow is associated with leukopenia and thrombocytopenia

94

ANEMIA: Hemolytic • Is caused by an abnormally high rate of the RBCs destruction

(hemolysis) due to:– Structural abnormalities of the RBC (more fragile cells)

• Hereditary spherocytosis – cells are spherical and can not be compressed

• Sickle cell anemia – cells have sickle shape → hemolysis

– Bacterial toxins, parasitic infections (i.e., malaria)– Adverse drug reactions– Autoimmune reactions

• The bone marrow is unable to compensate for premature destruction of RBC by increasing their production.

Thalassemias (α, β)• Hereditary hemolytic anemia• Abnormal or nonfunctional genes → globin chains are normal in

structure but are produced in reduced amounts• Cells are microcytic and hypochromic

Spherocytosis

95

ANEMIA OF CHRONIC DISEASE• Occurs as part of a chronic disorder (i.e., infection, inflammatory

disease, or cancer)

• Pathophysiologic mechanisms – Shortened RBC survival– ↓ EPO production and marrow responsiveness to EPO– Impaired intracellular iron metabolism

• Is microcytic or marginal normocytic

96

POLYCYTHEMIA• ↑ RBC count, Htc and Hb concentration

• Reasons– Hypoxic erythropoietic drive (i.e., high altitudes, chronic

pulmonary or cardiac disease)– Hemoconcentration - dehydration (i.e., heavy sweating,

vomiting or diarrhea)– Polycythemia vera or erythremia – uncontrolled RBC

production (i.e., neoplastic disease condition of hemocytoblastic cells)

• Results in– ↑ blood viscosity– ↑ peripheral resistance → ↓ venous return to the heart– ↑ blood volume tends to ↑ venous return – ↑ arterial BP– Ruddy skin and mucosa membranes with cyanotic tint

(sluggish blood flow → ↑ blood deoxygenation in the skin circulation)

97

CLINICAL CASEA 14-year-old girl complained of fatigue and loss of stamina. Her appetite was marginal,

as she was very conscious of maintaining her body weight at 96 pounds. Her monthly menstrual flow was always heavy and long from its onset at twelve years of age. Relevant laboratory findings included the following: – Hematocrit (Hct) - 28%– Hemoglobin (Hgb) - 9 g/dL– Iron 16 µg/dL– Bone marrow iron - absent– Erythrocytes - small and pale

Suggested treatment included ferrous sulfate or ferrous gluconate for six months orally between meals, since food may reduce absorption. A well-balanced diet was also suggested, as well as a gynecological examination.

Questions.1. What is the primary disorder of this individual?2. What does the ferrous sulfate or ferrous gluconate provide? Why is it necessary?3. What dietary inclusions would you suggest?4. Why is the gynecological examination important?5. Why is bone marrow iron an important clinical indicator in this individual?

98

PAST EXAMS QUESTION

A 51-year old male complains of generalized weakness and weight loss over the past 6 months. His blood pressure and pulse rate are elevated. Laboratory values revealed a hematocrit of 35% and hemoglobin level of 10.9 g/dL. A blood smear shows hypochromic and microcytic cells. A stool test for occult blood is positive. Which of the following would be the most likely cause of the findings?a. Acute blood lossb. Iron deficiencyc. Spherocytosisd. Folic acid deficiency Be. Autoimmune reactions

• Normal hemostasis is a consequence of tightly regulated process that maintain blood in a fluid state in normal vessels, yet also permit the rapid formation of a hemostasis clot at the site of a vascular injury.

HEMOSTASIS

Endothelium and Hemostasis

• Endothelia cells are key players in regulation of hemostasis. Before injury, they exhibit:– Antiplatelets – Production of Prostacyclin (PGI2)

and Nitric Oxide – Anticoagulant and– Fibrinolytic propertiesThey however acquire numerous pro coagulant

activities as a result of injury.

Endothelium Antiplatelets effects

• Endothelium produce– Prostacyclin (PGI2) and Nitric oxide which

impede platelets adhension– It also elaborate adenosine diphosphatase which

degrades adenosine diphosphate (ADP) and further inhibits platelets aggregation

Endothelium Anticoagulant effects

Endothelium inhibit coagulation via production of:• Thombomodulin which binds to thrombin and

converts it from procoagulant to an anticoagulant via its ability to activated Protein C which inhibits clotting by inactivating factors Va and VIIIa

• Protein S a cofactor for protein C, and tissue factor pathway inhibit (TFPI) a cell surface protein that directly inhibits tissue factor VIIa and factor Xa activities

Endothelium Fibrinolytic effects

• Endothelia cells synthesize tissue type plasminogen factor (t-PA), a protease that cleaves plaminogen to form plasmin, plasmin in turn cleaves fibrin to degrade thrombi

Endothelium Prothrombotic Properties

• As a result of trauma, inflammation and other factor, endothelia cells can induce a prothrombotic state via:

• Secretion of von Willebrand factor (vWF)• Secretion of tissue factor (the major activator

of the extrinsic clotting cascade)• Section of inhibitor or plasminogen activator

(PAIs) which limit fibrinolysis and tend to favor thrombosis

105

HEMOSTASISThe process of hemostasis can be divided into two distinct stages namely:1. Primary hemostasis – platelet plug formation2. Secondary hemostasis – coagulation cascade

• The goal of both primary and secondary hemostasis is to arrest bleeding from damaged blood vessels (hemo = blood, stasis = standing)

• Is counter-balanced by reactions, which prevent blood coagulation in uninjured vessels and maintain the blood in a fluid state– Balance between procoagulants and anticoagulants

• 4 overlapping processes or stages– Local vasoconstriction– Formation of a platelet plug– Formation of a web of fibrin proteins that penetrate and surround the platelet plug –

blood coagulation or clotting – Clot retraction.

106

LOCAL VASOCONSTRICTION• Results from

– Release of vasoconstrictor substances (paracrine & autocrine agents) from

• Platelets (i.e., serotonin & thromboxane A2) • Traumatized tissue

– Local myogenic spasm initiated by direct tissue damage– Reflex vasoconstriction initiated by activation of nociceptors and

other sensory endings

• Effects– ↓ blood flow and Pressure in the damaged area

Last for many minutes or even hours, during this time the ensuing processes of platelet plugging and blood

coagulation can take place

FORMATION OF A PLATELET PLUG (temporary hemostatic plug, white plug)

Intact blood vessel wall

Adhesion of the platelets

Platelet release reaction & activation

Platelet aggregation & plug retraction

Secretion of prostacyclin & nitric oxide

Local vasoconstriction

Collagen fibers are exposed to the blood and coated with WF*

Damaged blood vessel wall

-

-

-

Temporary hemostatic (platelet) plug

Factors that prevent/limit formation of a plug

1. Prostacyclin (prostaglandin I2).Inhibits platelet aggregation; vasodilator

2. Nitric oxide* (NO).Inhibits platelet adhesion, activation and aggregation and stimulates local vasodilation

* Von Willebrand factor, a protein synthesized by endothelialcells and megakaryocytes, enhances platelet adherence by forming a bridge between cell surface receptors and collagen in the subendothelial matrix.

+ stimulation- inhibition

+

+

108

FORMATION OF A PLATELET PLUG (cont.)Stage 1. Platelets adhesion

vWF binds to glucoprotein Ib receptors of platelets and to collagen

Failure of this step may be due to:- Absence of von Willebrand factor - Malfunction of collagen -

A. vWF - von Willebrand factor (soluble plasma protein) binds to collagen of subendothelial matrix

B. vWF exposes multiple intrinsic binding sites for the platelet specific membrane glycoprotein Ib (GPIb)

Scurvy

109

FORMATION OF A PLATELET PLUG (cont.)Stage 2-3 Platelets release reaction and activation. Binding of the platelets to the collagen → Release of agents from secretory granules (degranulation) – serotonin, adrenaline, several clotting factors, thromboxane A2, tissue factor and ADP Serotonin, adrenaline and ADP act locally → changes in the metabolism, shape, and surface proteins of the platelets.

Serotonin and thromboxane A2 stimulate local vasoconstriction

Primary Hemostatsis

Subendothelium

Endothelium

Von willebrand factor

GpIb

Deficiency: Glanzmann thombasthenia

GpIIb-IIIa complex

Fibrinogen

Platelet

Step 1: Transient vasoconstriction (endothelin)

Step 2: Platelet adhesion - von

willebrand factor bind to the disrupted blood vessel via GpIb

Step 3: Platelet release ADP and thromboxane A2 which stimulate adhesion of the next layers of platelets (recruitment) through a positive feedback mechanism and formation of a platelet plug inside the vessel ADP induces platelet to express GpIIB-IIIa which is needed to platelets aggregation via fibrinogen

Step 4: Platelet aggregation (Platelt plug)

GpIb

Deficiency: Bernard-Soulier syndrome

111

FORMATION OF A PLATELET PLUG (cont.)

• ADP and thromboxane A2 stimulate adhesion of the next layers of platelets (recruitment) through a positive feedback mechanism and formation of a platelet plug inside the vessel

Failure of this step:- Insufficient number of platelets - Dysfunctional platelets (prior

activation occurs during cardiopulmonary bypass, storage, exposure to aspirin, uraemia and acute and chronic alcohol exposure)

Stage 4: Recruitment and loose platelets aggregation

GpIIb-IIIa complex

Platelet

111

112

FORMATION OF A PLATELET PLUG (cont.)

Stage 5- irreversible platelet aggregation

• Destruction of the platelets membrane (stimulated by thrombin) → release of BAS from thrombocytes → secondary vasoconstriction

• Release of factor 3 (platelet thromboplastin) facilitates activation of blood coagulation

Stage 6. Plug retraction

• Contraction of actin and myosin in the aggregated platelets → compression and strengthening of the platelet plug

Normal Blood Vessel

Injured blood vessel

Exposed collagen binds and

activates platelets

Release of Platelet factors

Attracts more platelets

Aggregate into platelet plug

Secondary Hemostasis

This involve the conversion of the fibrinogen (solube) in the platelet plug to fibrin (insoluble).Fibrin is then cross-linked to yield a stable platelet-fibrin thrombus.

Secondary hemostasis involve the activation of coagulation cascade factors in both intrinsic and extrinsic pathways

116

BLOOD COAGULATION (CLOTTING)• Is the transformation of the blood into a solid gel (a clot or thrombus)• Occurs locally around the platelet plug; supports and reinforces the

plug• Requires 12 plasma clotting factors and platelets • Involves a cascade of biochemical reactions in which each factor that

is activated in turn activates the next factor • The fundamental reaction is conversion a soluble protein, fibrinogen

to an insoluble protein, fibrin

In coagulation a series of plasma proteins called blood-clotting factors play major roles.

Most of these are inactive forms of proteolytic enzymes. When converted to the active forms, their enzymatic actions cause the successive, cascading

reactions of the clotting process.

The two pathways• Two separate coagulation cascades result in blood clotting in

different circumstances. The two systems are:

1 The Intrinsic coagulation pathway (Contact activation pathway)

2 The Extrinsic coagulation pathway (tissue factor pathway)

However, the final steps in fibrin formation are common to both pathways.Phospholipids are required for activation of both coagulation pathways – provide a surface for the efficient interaction of several factors.

Clotting pathways• In the intrinsic pathway, all the factors required for

coagulation are present in the circulation.• For the initiation of extrinsic pathway, a factor

extrinsic to blood but released from injured tissue, called tissue thromboplastin or tissue factor (factor III), is required.

• The common pathway is initiated by the conversion of inactive clotting factor X to its active form factor Xa and results in the conversion of prothrombin to thrombin – calalysing the generation of fibrin

Scientific Name Common Name Main Function

Factor I Fibrinogen Converted to fibrin

Factor II Prothrombin Enzyme

Factor III Tissue thromboplasm Cofactor

Factor IV Calcium Cofactor

Factor V Proaccelerin Cofactor

Factor VII Proconvertin Enzyme

Factor VIII Antihemophilic factor Cofactor

Factor IX Christmas factor Enzyme

Factor X Stuart factor Enzyme

Factor XI Plasma thromboplatin antecedent Enzyme

Factor XII Hageman factor Enzyme

Factor XIII Fibrin stabilizing factor Enzyme

PLASMA CLOTTING FACTORS

120

3 PHASES OF BLOOD COAGULATION• Formation of a complex of activated substances - prothrombinase

(prothrombin activator)

• Formation of active thrombin from prothrombin– Is catalyzed by prothrombin activator

• Formation of insoluble fibrin from soluble fibrinogen– Is catalyzed by thrombin

PHASE 1 – FORMATION OF PROTHROMBINASE

XII

XI

II

XIa

XIIIa

XIII

IX

X

XIIa

Ca2+IXa

PF-3Ca2+VIII Xa

PF-3Ca2+

VX

Thrombin

Fibrinogen Fibrin Stable fibrin polymer

INTRINSIC PATHWAY EXTRINSIC PATHWAY

IIICa2+

VIIa VII

122

PHASE 3 – FORMATION OF FIBRIN• Thrombin catalyses release of 2

pairs of polypeptides from each fibrinogen molecule and formation of fibrin monomers– Ca++ and platelet factors are

also required

• Monomers join together to form insoluble fibrin polymers – a loose mesh of stands

• Stabilization of fibrin – formation of covalent cross-bridges, which is catalyzed by factor XIII (+ Ca++)

123

FINAL EVENTS OF HEMOSTASIS• Fibrin forms a meshwork, which supports

the platelet plug

• Clot occludes the damaged blood vessel and ↓ or stops bleeding

• Retraction of the clot due to contraction of fibrin fibers and contractile proteins of the platelets – ↑ clot density– Occlusion of the damaged vessel– Bringing the edges of wound together

→ facilitation of wound heeling

• Fate of the blood clot– Invasion by fibroblasts → formation of

connective tissue through the clot– Fibrinolysis and destruction of the clot

Overview of Hemostasis and Tissue RepairDamage to

wall of blood vessel

Vasoconstriction

Temporary hemostasis

Collagen exposed

Platelets aggregate into loose platelet plug

Intact blood vessel wall

Cell growth and tissue repair

Clot dissolves

Fibrin slowly dissolved by plasmin

Thrombin formation

Tissue factor exposed

Coagulation cascade

Converts fibrinogen to

fibrin

Platelets adhere and release

platelet factors

Reinforced platelet plug (clot)

125

ROLE OF VITAMIN K IN CLOTTING • Vitamin K acts as a cofactor of the enzyme γ-glutamyl carboxylase

• Is required for γ carboxylation in the liver of – Prothrombin and factors VII, IX and X – Proteins S and C (natural anticoagilants)

Vit K is activated by epoxide reductase in the liver

• γ carboxylation (introduction of a carboxylic acid group) of certain glutamate residues in target clotting factors → binding sites for Ca++ and PF3

• most of clotting factors are synthesized by the liver. Therefore, liver diseases (i.e., hepatitis, cirrhoses, atrophy) depress the clotting system. Decreased dietary intake of vit K has limited consequences on blood clotting because Vit K is continuously synthesized by the intestinal flora. Note that Vit K is fat soluble and requires fats for absorption. Lack of the bile decreases fat digestion and absorption.

126

ROLE OF Ca++ IN COAGULATION

• Ca++ is required for all steps of coagulation (except first 2 steps of the intrinsic pathway)

• ↓ in the plasma [Ca++] below the threshold level for clotting → ↓ blood clotting by both pathways

127

ROLE OF THE PLATELETS IN COAGULATION

Activated platelets

• Display specific plasma membrane receptors that bind several of the clotting factors → several cascade reactions take place on the surface of activated platelets

• Display phospholipids (platelet factors), which act as cofactors of the bound clotting factors

128

ROLE OF THE LIVER IN BLOOD COAGULATION

• Synthesis of the plasma clotting factors

• Synthesis of the bile salts, which are required for intestinal absorption of lipid soluble vitamin K

129

FIBRINOLYTIC SYSTEM

• Fibrin is digested by an enzyme, plasmin (fibrinolysin) into fibrin degradation products

• Plasmin also degrades factors Va, VIIIa and GPIb

• In the blood, plasmin is present as an inactive precursor, plasminogen

• Plasminogen is activated by plasminogen activators– Adrenaline, urokinase,

thrombomodulin-thrombin complex, kallikrein, tissue plasminogen activators (t-PAs)

• t-PAs are secreted by the endothelial cells

• urokinase is produced by kidney.

Fibrinolysis - dissolution or disposal of blood clots

Plasminogen activation

Plasminogen

Plasmin

Fibrin Soluble fibrin fragment

130

Fibrinolysis Clinical application-

Human t-PA is produced by recombinant DNA technology and available for clinical use. lyses clots in the coronary arteries if given to

patients soon after the onset of myocardial infarction.

Streptokinase (from bacteria-streptococcci) and urokinase are also fibrinolytic enzymes used in the treatment of early myocardial infarction

131

ANTICLOTTING MECHANISMS

• Removal of activated clotting factors from the blood by the liver

• Factors that reduce the adhesiveness of platelets– The smooth lining of the intact vessel walls– Mucopolysaccharides on the surface of endothelial cells

(glycocalyx) – repulsion of clotting factors and platelets– Circulation of the blood– Antiplatelet-aggregation effect of the prostacyclin by the intact

endothelial cells

ANTICLOTTING MECHANISMS: Natural anticoagulants

• Antithrombin III (antithrombin-heparin cofactors)– Is a plasma α globulin – its binding to heparin increases its activity.– Inactivates thrombin and some other clotting factors (IX, X, XI,

XII)

• Heparin– Is produced by the mast cells and blood basophils

By itself, it has little or no anticoagulant property, but when it combines with antithrombin III, it increases a hundred-fold the

effectiveness of antithrombin III

• Activated protein C – Inactivates factors Va and VIIIa and activates plasminogen

133

NATURAL ANTICOAGULANTS (cont.)

Endothelial cell

Thrombomodulin

Thrombin

Protein C

Activated Prot C

Protein S

Inactivation of inhibitors of plasminogen activator

Plasminogen

Fibrinolysis

Thrombin/thrombo-modulin/protein C pathway

Thrombomodulin is a thrombin-binding endothelial cell receptor

Binds thrombin and inactivates it

Complex of thrombin+thrombo-modulin binds protein C and activates it

Protein C in collaboration with protein S inactivates factors Va and VIIIa and activates plasminogen and fibrinolysis

VVaVIIIViIIa

Plasmin

Thrombin

Note: Mutated factor V cannot be inactivated (switched off) by activated protein C, and this will lead to hypercoagulable state

134

ANTICLOTTING MECHANISMS: SUMMARY

Tissue factor pathway inhibitor

AT III-Heparin

Proteins C & S

135

DRUGS THAT INHIBIT BLOOD CLOTTING (ANTICOAGULANTS)

• Heparin: Heparin binds to the enzyme inhibitor antithrombin III (AT), causing a conformational change that results in its activation. The activated AT then inactivates thrombin and other proteases involved in blood clotting such as XIIa, XIa, Xa and IXa

• Coumarin derivatives (i.e., warfarin) – Block stimulatory effects of vitamin K on synthesis of clotting

factors II, VII, IX, and X by the liver (inhibit epoxide reductase which activates vit K in the liver: K → K1)

• Aspirin – Low doses inhibit prostaglandins and thromboxanes synthesis by

the platelets → inhibition of platelet release reaction and platelet aggregation

– Is effective in preventing of heart attack and reduction of the incidence of sudden death

136

IN VITRO INHIBITION OF BLOOD CLOTTING• Keeping of blood in seliconized containers – prevention of contact activation of

platelets and factor XII

• Substances that bind ionized calcium to produce un-ionized calcium compound or to form insoluble salts with calcium – Sodium citrate or oxalate– Ammonium or potassium citrate– EDTA (ethylenediaminetetraacetic acid)

• Is ability to "sequester" di- and tricationic ions (Ca2+ & Fe3+)• Is widely used as an anticoagulant for blood samples for complete blood

count/full blood examination

• Heparin

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PROTHROMBIN TIME (protime, PT test)• Measures the clotting time of

plasma from the activation of factor VII, through the formation of fibrin clot

• Assesses the integrity of the extrinsic/tissue factor pathway and common pathways of coagulation (factors VII, X, V, II, I)

• The PT test is widely used to monitor patients taking anticoagulants as well as to help diagnose clotting disorders

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PROTHROMBIN TIME (cont.)• Depends on [prothrombin] in the blood

• Normal range 12 – 14 sec

• Increased– ↓ prothrombin (less than 10% of normal)– Deficiency of fibrinogen or factors V, VII, or X – Therapeutic anticoagulants (i.e., heparin,

warfarin, aspirin), some drugs (i.e., antibiotics, anabolic steroids, estrogens, etc.)

– Liver diseases– Vit K deficiency – Disseminated intravascular coagulation

• Decreased– Vit K supplementation– Thrombophlebitis

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ACTIVATED PARTIAL THROMBOPLASTIN TIME

(aPTT) • Assesses the integrity of the intrinsic and common pathways

of coagulation

• Measures the clotting time of plasma, from the activation of factor XII by a reagent through the formation of fibrin clot

• Normal range 25 – 38 sec

• Prolonged time– Use of heparin– Antiphospholipids antibodies– Coagulation factors deficiency (intrinsic and common pathways; i.e.,

hemophilias)

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2 TYPES OF ABNORMALITIES OF HEMOSTASIS

• Excessive bleeding (hemorrhagic disease) caused by deficiency of a clotting factor/s or platelets

• Excessive clotting: thrombosis, embolism, disseminated intravascular coagulation

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CONDITIONS THAT CAUSE EXCESSIVE BLEEDING

• Vitamin K deficiency

• Deficiency of clotting factors (i.e., hemophilia)

• Deficiency of thrombocytes – thrombocytopenia

• Deficiency of von Willebrand factor

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VITAMIN K DEFICIENCY• Results from

– ↓ intestinal absorption of fats due to ↓ bile secretion (i.e., liver disease or obstruction of the bile ducts)

– ↓ dietary intake of vit K (limited importance)

• Results in – ↓ hepatic gamma carboxylation of

• Prothrombin (II)• Factors VII, IX and X• Protein C and S

– Bleeding tendency• Prolonged prothrombin time and partial thromboplastin time• Normal platelets count and serum fibrinogen split products

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HEMOPHILIA

• Is a hemorrhagic disease that results from deficiency of – Factor VIII (the smaller component) - hemophilia A

or classical – Factor IX – hemophilia B, Christmas disease – Factor XI – hemophilia C

• Is a genetic disease– Hemophilia A and B are sex linked (X chromosome)

• Occur in males• Females are hemophilia carriers

• Results in ↑ aPTT (PT, thrombocytes count, fibrin split products are normal)

Deficiency Factor

Clinical Syndrome Cause

Factor I Afibrinogenemia Depletion during pregnancy with premature separation of placenta: Congenital

Factor II Hypoprothrombinemia (Hemorrhagic tendency in liver diseases

Decreased hepatic synthesis (secondary to vitamin K deficiency)

Factor V Parahemophila Congenital

Factor VII Hypoconvertinemia Congenital

Factor VIII

Hemophilia A (classical hemophilia)

Congenital recessive sex-linked defect due to abnormalities of the gene that codes for factor VIII (X chromosome)

Factor IX Hemophilia B (Christmas disease)

Congenital recessive trait carried on X chromosome

Factor X Stuart-Prower factor deficiency

Congenital

Hemophilia C (PTA deficiency

Congenital

Hageman trait Congenital

HEMOPHILIA

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THROMBOCYTOPENIA• Low thrombocytes count (below 50 000/m l) → poor plug formation,

deficient clot retraction, deficient platelet phospholipids, poor constriction of ruptured vessels → bleeding tendency from many small venules and capillaries

• Multiple hemorrhages in the skin and mucous membranes – thrombocytopenic purpura– Petechiae – small punctate hemorrhages(1-3 mm)– Echymoses - large hemorrhages (bruises)

• Other causes of purpura– ↓ plasma level of 1 or more clotting factors– ↑ fragility of capillary walls (congenital, Vit C deficiency,

adrenal failure, toxins, drugs, allergic reactions)

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von Willebrand’s disease

• Is the most common genetic bleeding disorder

• Results from defect in vWF ( quantitative or functional)

• Results in combination of– Platelet function abnormality (vWF) - impaired adhesion– Clotting factor deficiency (factor VIII) - ↑ aPTT (PT is normal)

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THROMBO-EMBOLIC CONDITIONS• Thrombosis - blood clotting within the CVS which obstruct the blood flow through

the CVS (Should be distinguished from extravascular clotting, clotting in wounds and clotting that occurs in the CVS after death). Thrombosis is rather a pathological condition.

• Common causes– Roughened endothelial surface (i.e., atherosclerosis, infections, traumas)– Slow blood flow– Hypercoagulobility– Acquired refers to transient or acquired conditions that increase the tendency

to clot. This might include antiphospholipid antibodies or a temporary hypercoagulable state such as pregnancy. Also, advanced carcinomas of the pancreas or lung may produce a hypercoagulable state.

– Congenital refers to hereditary conditions that increase the tendency to clot. These include Factor V Leiden, prothrombin ,protein C, protein S and antithrombin deficiencies

ConsequencesFormation of emboli (thromboembolism) – braking down of the thrombus and spreading of its particles particles throughout the CVS

Thrombosis in the left side of the heart and large arteries → emboli in the brain, kidneys, etcThrombosis in the venous system and in the right side of the heart → emboli in the pulmonary circulation

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DESSIMINATED INTRAVASCULAR COAGULATION

• Reasons– Large areas of necrotic tissue (release of tissue factors into the blood)– Septicemia (activation of clotting by circulation bacteria and bacterial

toxins)

• Consequences– Consumption coagulopathy

• ↓ fibrinogen, thrombocytopenia• ↑ fibrin split products• ↑ PT and PTT

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CHALENGE YOURSELF 1/5A baby is born prematurely at 28 weeks gestational age with a birth weight of 1200 g. A few weeks after birth his mother noticed a bleeding tendency in the infant. Blood test revealed a low prothrombin level. Which vitamin can be given to the baby to reduce or to prevent the bleeding tendency?a. Vitamin B12b. Vitamin B6c. Vitamin Kd. Folic acid e. Vitamin A

Answer is C

CHALENGE YOURSELF 2/5

• A 72-year-old African-American man undergoes hip surgery. On his third hospital day he experiences chest pain, tachycardia, dyspnea, and a low-grade fever. The man goes into cardiac arrest, and efforts to resuscitate him are unsuccessful. On autopsy a massive pulmonary embolus is discovered. Which of the following, if present, would most likely predispose the patient to this event?

(A) Factor VIII defi ciency(B) Low serum homocysteine levels(C) Mutation in the Factor V gene(D) Overproduction of protein C(E) von Willebrand factor defi ciency

Answer is C

CHALENGE YOURSELF 3/5

Taking aspirin every day can reduce the risk of heart disease becausea.it is a powerful vasodilator b it stimulates fibrinolysis c. it prevents atherosclerosis d.it loosens atherosclerotic plaque on arterial walls e. it prevents platelet aggregation

Answer is E

CHALENGE YOURSELF 4/5

• A one month old Caucasian with a history of persistent jaundice experiences muscle rigidity, lethargy and seizure. Which of the following causes of hyperbilirubenimia is most likely to produce this patient’s neurological abnormalities.

A. Absent liver onjugation enzymesB. Deficient bilirubin excretion into bile canaliculiC. Impaired canalicular bile transitD. Increased gut deconjugation of bilirubinE. Impaired gut reabsorption of bilurubin

The answer is A

CHALENGE YOURSELF 5/5

• A 65-year-old man presented with history of acute chest pain that radiates to his left arm. Coronary angiography demonstrates more than 75% occlusion of his coronary artery. He was administered a thrombolytic agent for reestablishment of blood flow to the dying myocardium. The thrombolytic agent activates:

A. HeparinB. ThrobinC. Plasminogen D. KininogenE. Prothrombin

The Answer is C

Fill in the gap please

PT PTT Platelet Count

Bleeding Time

Reticulocyte

Megakyryocyte

D-Dimer Schicocyte

Hemophilia A N N N N N No No

Hemophilia B

vWF disease

Vit K Def

Liver Disease

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Section 3 Blood Group and Transfusion

Agglutination, hemolysis and the transfusion reaction

157

Blood Groups

Based on the types of special antigens on the surface of the red blood cells

Two particular groups of antigens are more likely than the others to cause blood transfusion reactions.ABO system Rh system,

158

ABO Blood

classifications depend on the presence or absence of the agglutinogens (antigens) on the surface of the red cell membrane.

159

Agglutinogens

Two antigens- type A and type B occur on the surface of the red blood cells.

These antigens are called agglutinogens because they often cause blood cell agglutination, causing blood transfusion reactions

Group OGroup B Group ABGroup A

160

Agglutinins The agglutinins (antibodies) are gamma globulin in the plasma. When type A agglutinogen is not present in a person’s red

blood cells, antibodies known as anti-A agglutinins develop in the plasma.

when type B agglutinogen is not present in the blood cells, antibodies known as anti-B agglutinins develop in the plasma.

• type O blood, although containing no agglutinogens,does contain both anti-A and anti-B agglutinins.

• Finally, type AB blood contains both A and B agglutinogens but no agglutinins.

161

Major types of ABO blood group

The bloods are normally classified into four major types, depending on the presence of absence of the two

agglutinogens, A and B

162

ABO blood group• But why are these agglutinins produced in people

who do not have the respective agglutinogens in their red blood cells?

• Small amounts of type A and B antigens enter the body in food, in bacteria, and in other ways, and these substances initiate the development of the anti-A and anti-B agglutinins.

163

Blood Type Agglutinogen Agglutinin

O -- Anti-A, Anti-B

A A Anti - B

B B Anti-A

AB A and B --

164

Agglutination and hemolysis in transfusion reaction

When bloods are mismatched so anti-A or anti-B plasma agglutinins are mixed with red blood cells that contain A or B agglutinogens respectively, agglutinins will make the red cells adhere to each

other – agglutination. The clumps of red cells will plug small blood

vessels throughout the circulatory.

165

Group B Group B

Group BGroup B anti-B

anti-B

anti

-B

anti

-B

166

167

Agglutination and hemolysis in transfusion reaction

During the ensuring few hours to a few days, either physical distortion of the cells or

attack by phagocytic white cells destroys the agglutinated cells,

releasing hemoglobin into the plasma, which is called “hemolysis” of the red blood

cells. Type II hypersensitivity reaction

168

agglutination

During severe hemolytic reaction, fever, chills and shock may occur.

One of the most lethal effects of transfusion reactions is kidney shutdown, which can begin within few minutes to a few hours.

If the shutdown is complete and fails to open up, the patient die of renal failure.

169

Blood typing

Before giving a blood transfusion to a person, it is necessary to determine the blood type of the recipient’s blood and the blood type of the donor blood so that thebloods can be appropriately matched. This is calledblood typing and blood matching.

170

Rh Blood Groups

Blood Typing, Showing Agglutination of Cells of the Different Blood Types with Anti-A or Anti-B Agglutininsin the SeraRed Blood Cell Types Sera Anti-A Anti-BO - -A + -B - +AB + +

Blood typing and blood matching

Serum fromContains

Serum fromContains

Serum fromContains

Serum fromContains

A

Agglutinin B

B

Agglutinin A

O

Agglutinin A &BAB

No Agglutinin

N = No Agglutination A = Agglutination

A B O AB

Blood typing and blood matching

ANSerum from A

Agglutinin B

B

Agglutinin A

O

Agglutinin A &BAB

No Agglutinin

N

N

N

N

A

A

A

A

N N N

N

A A

Contains

Serum fromContains

Serum fromContains

Serum fromContains

N = No Agglutination A = Agglutination

A B O AB

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Rh BLOOD GROUP SYSTEM• Rh (rhesus factor)

– Is named after the rhesus monkey in which it was found– Rh is an antigen present on the RBC membrane – transmembrane proteins,

which may act as ionic channels– 6 types of Rh AG

• C, D, E, c, d*, and e (+ 43 other Rh antigens, which are less common)

• Antigen D is the most common and potent– Presence of D antigen - Rh+ (RhD+)– Lack of D antigen - Rh- (RhD-)

• Population data on RhD+– 85% of Caucasians– 90% of African Americans– 99% of Asians– 100% of Africans

174

Rh BLOOD GROUP SYSTEM: Antibodies

• Blood of normal individual does not contain anti-Rh antibodies (with exception of anti-E)

• Production of anti-Rh antibodies can be evoked by– Transfusion of Rh- individual with Rh+ blood (only 0.5 ml may

suffice)– The presence of Rh+ fetus in a Rh- mother

175

Rh BLOOD GROUP SYSTEM: Typing

• Mixing of the blood sample with anti-Rh serum (D AB)– Agglutination – RhD+– No agglutination – RhD-

176

Rh IMMUNE RESPONSE

• Transfusion of RhD+ blood to un-immunized RhD- recipient– No immediate reaction– Sensitization of recipient’s blood to RhD AG - slow formation of

anti-RhD AB– Delayed transfusion reaction – hemolysis of the donor’s RBC

that still circulate in the recipient’s blood

• Transfusion of RhD+ blood to immunized RhD-recipient – Enhanced transfusion reaction – acute hemolysis

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ERYTHROBLASTOSIS FETALIS

↑ risk starting from gravida 2 or para 2.

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ERYTHROBLASTOSIS FETALIS (cont.) Rh+ fetus/RH- mother

Some fetal RBC enter the maternal circulation

Formation of anti-Rh AB in the maternal blood

Hemolysis of fetal Rh+ RBC

Some anti-Rh antibodies enter the fetal circulation

Gross edema (hydrops fetalis)Hemolytic jaundice, anemia, high reticulocyte count, erythroblastsBile pigments pass into the brain and are deposited in the basal ganglia → impairment of motor functions - Kernicterus

Gross edema

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ERYTHROBLASTOSIS FETALIS (cont.)• Treatment

– Replacement of the neonate’s blood with Rh- blood – removal of Rh AB and damaged RBC

• Prevention– Administration of anti-D AB to the Rh- pregnant woman who

expects RhD+ baby• Destruction of Rh+ RBC, which cross the placenta and

enter the maternal circulation• Inhibition of AG-induced AB production by B

lymphocytes in the pregnant woman

180

Past exams questionFred's blood type is A, Rh- and Linda is B, Rh+. Fred and Linda have a 4-year-

old son who is AB, Rh+. Which one of the following conclusions is correct?a. During the second pregnancy Ginger needs to take anti-D antibodies b. There is no risk to a second child to develop erythroblastosis fetalis c. If the son needs a blood transfusion Fred could provide it safely, but

not Ginger d. Fred is not the boy’s father

B