Physiology of Blood Lecture Notes PDF

Physiology of Blood Lecture Notes for Physical Therapy Students By Dr. Hany Gamal Professor of Physiology Cairo & MUST Universities  BLOOD Blood is the vital fluid tissue that circulates inside the blood vessels. Blood volume is ab...

Physiology of Blood Lecture Notes for Physical Therapy Students By Dr. Hany Gamal Professor of Physiology Cairo & MUST Universities  BLOOD Blood is the vital fluid tissue that circulates inside the blood vessels. Blood volume is about 8% of body weight. Thus for a 70 kg person blood volume is about 5600 ml. Functions of the Blood: 1- Transport function: Blood is the medium through which many substances such as nutrients, gases, hormones and products of metabolism are transported from one place to another inside the body. 2- Defensive function: White blood cells and antibodies present in plasma defend the body against microorganisms such as bacteria and viruses. 3- Hemostatic function: Platelets and clotting factors of the blood are essential for stoppage of bleeding when a blood vessel is injured. 4- Homeostatic function: Homeostasis is the maintenance of internal environment of the body constant. It is achieved by continuous exchange between interstitial fluid, blood and external environment through lungs, gastrointestinal tract and kidneys. Composition of the Blood: Blood is composed of two main parts: plasma and cellular elements. A- Plasma: It is the fluid part of blood. It constitutes about 55% of blood volume. B- Cellular elements: These represent about 45% of blood volume and include: 1- Red blood corpuscles (RBCs) or Erythrocytes. 2- White blood cells (WBCs) or Leucocytes. 3- Platelets (thrombocytes). __________________ 1  PLASMA Plasma is a pale yellow fluid that constitutes 5% of body weight. Plasma clots on standing, and the remaining fluid is called serum. Composition of Plasma: 1- Water: constitutes 90% of plasma volume. 2- Inorganic substances: constitute 0.9%. - The chief cation is sodium (Na+) - The chief anions are Chloride (Cl-) and bicarbonate (HCO3-) 3- Organic substances: constitutes 9.1% These include many substances such as plasma proteins, lipids, glucose, amino acids, vitamins and waste products (e.g. uric acid and urea) 4- Blood Gases: These include oxygen and carbon dioxide. Plasma Proteins There are three main types of plasma proteins: 1- Albumin. 2- Globulins: which are subdivided into α1, α2 β1, β2 and γ globulins. 3- Fibrinogen. The concentration and molecular weight of the different plasma proteins are shown in the following table: Concentration Molecular weight g/dl Albumin 3.5-5.0 69’000 Globulins 2.5 90’000-156’000 Fibrinogen 0.4 340’000 Concentration and molecular weight of plasma proteins. Site of Formation of Plasma proteins: 2  1- Albumin, fibrinogen, prothrombin and 50% of globulins are formed in the liver. 2- Globulins are also formed mainly by plasma cells. Albumin/Globulins Ratio (A/G ratio): Normally ranges between 1.2 – 1.6. Low A/G ratio can occur in: - Liver diseases: due to decreased formation of albumin. - Kidney diseases: due to loss of albumin in urine. Albumin is lost more than other proteins due to its smaller molecular weight. Functions of Plasma proteins: 1- Osmotic Function: Plasma proteins cannot cross capillary wall due to their high molecular weight. Therefore they exert osmotic pressure on the wall of the capillaries. This pressure is called the colloidal osmotic (or oncotic) pressure which is about 28 mmHg. This pressure tends to pull water into the capillaries. Albumin is responsible for most of this pressure because it has the highest concentration and the smallest molecular weight. 2- Buffer Action: Plasma proteins can act as weak acids or weak bases depending on the pH of the blood. At normal pH of blood, which is 7.4, plasma proteins act as weak acids. They form Sodium salts (Na proteinate). Proteinic acid and Na proteinate constitute buffer system which is responsible for 15% of buffering power of blood. Lactic acid + Na proteinate Na Lactate + proteinic acid (strong acid) weak acid NaOH + proteinic acid Na proteinate + H2O 3- Defensive Function: γ globulins are the immunoglobulins (or antibodies) that defend the body against microorganisms. 4- Blood Clotting: 3  Fibrinogen and prothrombin are essential for coagulation of blood. 5- Blood Viscosity: Blood is 3 times more viscous than water. Plasma is 1.5 more viscous than water. Fibrinogen is responsible for most of this viscosity due to the elongated shape of its molecules. Viscosity is important for the maintenance of peripheral resistance and arterial blood pressure. 6- Capillary function: Plasma proteins close the pores in capillary wall limiting their permeability 7- Transport and Conservation of important elements: Plasma proteins act as carriers of important substances in blood such as hormones, vitamins and minerals. Binding of these substances to plasma proteins prevent their loss in urine and act as reservoir of these substances. RED BLOOD CORPUSCLES “ERYTHROCYTES” Red blood corpuscles are non-nucleated structures and therefore are not true cells. That is why they are called corpuscles. Erythrocytes (RBCs) count: Count varies with age and sex. Adult males: about 5.4 million/mm3. Adult females: about 4.8 millions/mm3. Infants have higher RBCs count than adults. Children have lower RBCs count than adults. RBCs count decreases in old age. 4 Shape and Size: RBCs are circular biconcave discs. An average erythrocyte has a diameter of 7.2 μm, a thickness of 2.2 μm and a volume of 90 μm3. The biconcave shape of RBCs has the following advantages: - Produces a larger surface area for the same volume. - Enhances cell flexibility allowing RBCs to pass through narrow blood vessels without being ruptured. - Allow change in cell volume with minimal increase in tension on the cell membrane. Hematocrit Value (Packed cell volume PCV): It is the percentage ratio of RBCs volume to total blood volume. It is normally about 46% in males and 42% in females. It is higher in venous blood than arterial blood. - Low hematocrit value is observed in anemia (low RBCs count) and hydration (leading to increased plasma volume). - High hematocrit value is seen in polycythemia (high RBCs count) and dehydration (leading to decreased plasma volume). Structure of Erythrocytes: - Erythrocytes are surrounded by plastic semi-permeable membrane. - Hemoglobin is the main constituent of RBCs. It represents 34% of their weight. - Potassium is the chief intracellular cation. - Erythrocytes contain carbonic anhydrase enzyme, which is important in carbon dioxide transport. - Erythrocytes have no mitochondria. Therefore, they obtain their energy requirements from anaerobic glycolysis. This energy is needed mainly for the operation of Na+-K+ pump. HEMOGLOBIN: It is the red oxygen-carrying pigment. Its molecular weight is 64000. Chemical structure: - Hemoglobin molecule is formed of four subunits. - Each subunit is formed of: 5  - Heme: which is formed of 4 pyrrole rings (protoporphyrin IX) and one ferrous ion. - Polypeptide chain. Therefore, each hemoglobin molecule contains 4 polypeptide chains (called collectively globin) and 4 ferrous ions. Types of Hemoglobin: Differences between different types of hemoglobin arise from differences in the structure of the polypeptide chains. 1- Adult Hemoglobin (HbA): Contains 2α and 2β polypeptide chains. 2- Fetal Hemoglobin (HbF): Contains 2α and 2γ polypeptide chains. The γ chains are formed of the same number of amino acids but have37 different amino acids from β chains. HbF, which is normally present in the fetus, is replaced by HbA after birth. This replacement is almost complete by the age of 4 months. HbF has more affinity to oxygen than HbA. This facilitates the movement of oxygen from maternal to fetal blood. Hemoglobin Concentration: - Adult male blood contains 13-18 grams of Hb /dl. - Adult female blood contains 12-16 grams of Hb/dl. - Infant blood may contain up to 19 grams of Hb /dl. This is due to relative intrauterine hypoxia. Reactions of Hemoglobin: 1- With Oxygen: Oxygen can attach loosely to iron of the hemoglobin molecule forming “oxyhemoglobin”. This process is called oxygenation. Iron remains in the ferrous state. One molecule of hemoglobin can combine with 4 oxygen molecules. One gram hemoglobin can combine with 1.33 ml of oxygen. The affinity of hemoglobin to oxygen is affected by: a- pH of the blood: low pH (high H+ concentration) decreases the affinity of hemoglobin to oxygen. This is because H+ ions compete with oxygen for hemoglobin. 6  b- Temperature: high temperature decreases the affinity of hemoglobin to oxygen. c- 2,3 diphosphoglycerate (2,3-DPG): 2,3-DPG combines with globin part and reduce the affinity of hemoglobin to oxygen. 2- With carbon dioxide: Carbon dioxide combines with the globin part of hemoglobin forming “carbamino-hemoglobin”. About 10% of carbon dioxide is carried in this form from tissues to lungs. 3- With carbon monoxide: Carbon monoxide combines with iron of the hemoglobin forming carboxy-hemoglobin. It has 200 times more affinity to hemoglobin than oxygen. Therefore, carbon monoxide can displace oxygen and prevents it from combining with hemoglobin. 4- Oxidation: Exposure of Hemoglobin to strong oxidizing agents leads to oxidation of hemoglobin that is changed into a dark substance called “methemoglobin”. Iron is changed to ferric state. Methemoglobin cannot carry oxygen. An enzyme called NADH- methemoglobin reductase changes methemoglobin back to hemoglobin. Functions of RBCs: 1- Oxygen transport. 2- Carbon dioxide transport: this takes place in the form of carbamino- hemoglobin. In addition, RBCs contain carbonic anhydrase (C.A.) which is important for the rapid hydration of carbon dioxide: C.A. CO2 + H2O H2CO3 H+ + HCO3- 3- Buffering: Buffering power of hemoglobin is six times that of plasma proteins. Hemoglobin is a weaker acid than oxyhemoglobin. Therefore, hemoglobin is a better buffer than oxyhemoglobin. 7  4- The membrane of RBCs keeps hemoglobin inside them. If hemoglobin is free in plasma it will lead to: - Blocking of renal tubules due to filtration of hemoglobin through the glomeruli. - Increase the work of the heart due to increasing the viscosity and osmotic pressure of plasma. Life Span and Fate of RBCs: RBCs live in the circulation for about 120 days. Aged RBCs become more fragile and rupture as they pass through tight spots of the circulation e.g. the spleen. Released hemoglobin is taken by the macrophages and is broken down into globin and heme. Globin is broken down to amino acids. Heme is broken down to iron and protoporphyrin. Iron is carried by a special transport protein called transferrin to the bone marrow for the formation of new RBCs and to the liver to be stored as ferritin. Protoporphyrin is broken down into bilirubin, which is then secreted by the liver in bile. Excessive destruction of RBCs leads to formation of large amount of bilirubin. This causes yellow coloration of skin and mucous membranes. This condition is called “hemolytic jaundice”. SOME PHYSICAL PROPERTIES OF BLOOD: 1- Erythrocyte Sedimentation Rate (ESR): Erythrocytes are heavier than plasma. If blood in a test tube is prevented from coagulation and is left undisturbed, erythrocytes will sediment to the bottom of the tube leaving clear plasma above. The rate at which the erythrocytes sediment is called the “erythrocyte sedimentation rate”. Normal Value of ESR: o Males: 6 mm after one hour, 8 mm after two hours. o Females: 10 mm after one hour, 16 mm after two hours. Clinical Significance of ESR: Increased ESR is observed in: o Physiological conditions e.g. menstruation and pregnancy and after meals. 8  o Pathological conditions: e.g. infections, tissue destruction (such as myocardial infarction) and malignant tumors. Elevation of immunoglobulins concentration in plasma favors sticking of RBCs together forming rouleaux. This leads to more rapid sedimentation rate. o ESR is not a diagnostic test. It cannot tell the type of disease. But ESR is a prognostic test. Repeated measurement of ERS indicates the progress of disease and effect of treatment. 2- Blood Viscosity: The viscosity of blood is 3 times that of Water. The viscosity of plasma is 1.5 times that of water. RBCs and plasma proteins (mainly fibrinogen) are responsible for the viscosity of blood. Blood viscosity increases in polycythemia and decreases in anemia. Blood viscosity is important for normal blood pressure and blood flow. ERYTHROPOIESIS ---------------------------------- Erythropoiesis is the process of formation of new erythrocytes. Site of Erythropoiesis: 1- In Fetus: It takes place in liver and spleen. 2- In Infants and children: It takes place bone marrow of all bones of the body. 3- In Adults: it takes place in membranous bones like the ribs, vertebrae and upper part of humerus and femur. Stages of Erythropoiesis: - Uncommitted stem cells in the bone marrow are changed into committed stem cells. This occurs under the effect of certain chemical substances called “interleukins”. 9  - The committed stem cells are sensitive to a hormone called “erythropoitin” which stimulates them to develop into erythrocytes. - Committed stem cells undergo successive divisions and develop finally into erythrocytes. FACTORS AFFECTING ERYTHROPOIESIS: 1- Healthy Bone Marrow: Bone marrow is the site where erythropoiesis takes place in adults. Depression or destruction of bone marrow may occur due to exposure to x-ray, atomic radiation and drugs or due to malignant tumors. Under such conditions, erythropoiesis is decreased. This is called aplastic anemia. 2- Liver: Liver is important for erythropoiesis because it is the site of: - Formation of the globin part of hemoglobin. - Formation of 15% of erythropoitin. - Storage of iron and vitamin B12. 3- Hormones: Many hormones are important for erythropoiesis such as erythropoitin, thyroid hormones, glucocorticoids and androgens. 4- Oxygen Supply to Tissues: Erythropoiesis is stimulated by hypoxia. Hypoxia is lack of oxygen at tissues. Hypoxia occurs under many conditions such as: - High altitude. - Athletes (due to increased oxygen demand) - Hemorrhage (due to loss of erythrocytes). - Lung and heart diseases. Hypoxia stimulates erythropoiesis through the secretion of erythropoietin. Erythropoitin: It is a glycoprotein hormone. Its molecular weight is 35000. * Source of Erythropoietin: 10  o In adults: The kidneys form 85% of erythropoietin while the liver forms the remaining 15%. o In Fetus: the liver forms almost all erythropoietin. * Function of Erythropoietin: Erythropoietin combines with receptors on the committed stem cells in bone marrow leading to: o Increase the number of these committed stem cells. o Stimulate the development of committed stem cells. o Accelerate all stages of erythropoiesis. * Regulation of Secretion of Erythropoietin: o Hypoxia, especially of the kidney, is the main stimulus for erythropoietin secretion. o Alkalosis stimulates erythropoietin secretion. o Androgens and cobalt salts stimulate erythropoietin secretion. o -adrenergic stimulation and adenosine stimulate erythropoitin secretion. 5- Diet: Proteins, iron, vitamin B12 and folic acid are especially important for erythropoiesis. A- Proteins: High biological value proteins are needed for the formation of globin part of hemoglobin. B- Iron: Average daily intake of iron is 20 mg. Men lose about 0.6 mg of iron daily, while women lose double this amount due to menstrual blood loss. The total amount of iron in the body is 4 grams. About 70% of this iron is present in hemoglobin, 3% is present in myoglobin and 27% is stored mainly in the liver. Normal iron concentration in plasma is 135 g/dl in males, and 110 g/dl in females. To keep the total body iron stores constant, iron loss should be replaced by absorption of ingested iron. Iron Absorption: 11  o Iron can be absorbed in the ferrous state. Most dietary iron is in the ferric state. Gastric HCl and ascorbic acid present in diet help to reduce ferric to ferrous iron so that it can be absorbed. On the other hand, phytic acid, phosphates and oxalates reduce iron absorption because they form insoluble complexes with iron. o Iron is absorbed mainly in the upper part of the small intestine. Mucosal cells contain intracellular iron carrier. Part of the iron bound to this carrier is delivered to the mitochondria. The remaining part is either combined with apoferritin or is carried in plasma as transferrin. o Apoferritin is found in mucosal cells and other tissues such as the liver. When it combines with iron it is changed to ferritin. o Ferritin is the main storage form of iron. It is present in the liver and intestinal mucosal cells. When the life cycle of the mucosal cells end, they are shed into the lumen of the intestine and pass in stools with their content of ferritin. o Iron is transported in blood bound mainly to transferrin. Transferrin is normally 35% saturated with iron. When transferrin is more saturated with iron, more iron goes to ferritin in mucosal cells and finally lost in stools. The amount of transferrin in plasma is inversely proportional to the amount of iron in the body. ▪ In iron deficiency anemia: transferrin concentration increases and its saturation with iron decreases and less ferritin is formed. Thus less iron is lost in stools. ▪ In iron overload: Transferrin concentration decreases and its saturation with iron increases. More ferritin is formed in mucosal cells and is finally lost in stools. HCl , vitamin C (in stomach) Fe+++ Fe++ Blood vessel Intestinal Mucosal Cell Intracellular 12  carrier Apoferritin Transferrin mitochondria Ferritin Iron Absorption C. Vitamins: Most vitamins are important for erythropoiesis. Vitamin B12 and folic acid are particularly essential. They are needed for DNA synthesis. Bone marrow cells grow and divide very rapidly. This process of growth and division needs DNA synthesis for nuclear maturation. Deficiency of B12 and folic acid leads to failure of nuclear maturation and cell division. However, RNA is formed normally and the cells continue to synthesize hemoglobin and they increase in size. The resulting RBCs are therefore larger in size but fragile and irregular in shape. * Vitamin B12 (Cyanocobalamine, Extrinsic Factor): Source: food of animal origin e.g. meat and liver. Requirements: 5 g/day. Storage: Mainly in the liver. Amount stored is about 5 mg. Absorption: Parietal cells of the gastric glands secrete a glycoprotein called “intrinsic factor”. This glycoprotein combines with vitamin B12 and serves two functions. First, it protects it from being digested. Second, it binds vitamin B 12 to a specific receptor on mucosal cells of the lower ileum (terminal part of small intestine). Both of vitamin B12 and the intrinsic factor enter the mucosal cells by pinocytosis. Inside the cells vitamin B12 is set free and is absorbed to the blood. In plasma, vitamin B12 is bound to a transport protein called “transcobalamine II”. 13 Deficiency of vitamin B12 is usually due to failure of absorption. It leads to Macrocytic anemia. It also causes neurological manifestation because vitamin B 12 is needed for myelination of nerves. B12 in diet Parietal Cells (stomach) Intrinsic Factor Liver Receptor Transcobolamine II B12 Mucosal epithelial cell in terminal ileum Blood vessels Vitamin B12 Absorption * Folic Acid: Source: green vegetables, fruits, meat and liver. It is easily destroyed by cooking. Deficiency: may be due to deficiency in diet or failure of absorption due to gastro- intestinal tract disease. Deficiency of folic acid causes macrocytic anemia. D. Trace Elements: ▪ Copper: acts as cofactor for hemoglobin formation. 14  ▪ Cobalt: is also a cofactor for hemoglobin formation. It is a part of vitamin B 12 and it stimulates erythropoitin secretion by the kidneys. ANEMIAS Anemia is the decrease in oxygen carrying power of blood due to decreased number of RBCs or a decrease in their hemoglobin content. Anemia is present when: RBCs count is less than: 4.5 millions/mm3 in males. 3.9 millions/mm3 in females. Hemoglobin Concentration less than: 13.0 g/dl in males. 12.0 g/dl in females. In order to determine the type of anemia, we use indices that describe the size an appearance of the red blood corpuscles: 1- Mean Corpuscular Hemoglobin (MCH): It is the average amount of Hb in a single red blood corpuscle. Normal value: 25 – 32 pg (picogram) Values less than 25 pg are called hypochromia. 2- Mean Corpuscular Volume (MCV): It is the average volume of single red blood corpuscle. Normal value: 80 – 95 3. Values less than 80 indicate microcyte Values greater than 95 indicate macrocyte. Types of Anemia: Anemia is classified according to the size and Hb content of the red blood corpuscle into three types: 1- Normocytic Normochromic anemia. 2- Microcytic Hypochromic Anemia. 3- Macrocytic Anemia. 1- Normocytic Normochromic Anemia: This type of anemia is characterized by normal MCV and MCH. Causes: 15  o Acute Blood Loss: If blood is lost (hemorrhage) the liver can replace plasma volume in 2 two days while the bone marrow replaces RBCs in a longer time. Therefore, RBCs become diluted in plasma. o Excessive Hemolysis (Hemolytic Anemia): This can be due to intrinsic disorders or extrinsic factors. Intrinsic disorders include hereditary spherocytosis, G-6-PD deficiency and sickle cell anemia. Extrinsic factors include Rh incompatibility, exposure to some chemicals as benzene derivatives, and some infections like malaria and streptococcal infection. o Bone marrow depression (Aplastic Anemia): Bone marrow may be destroyed by x-rays, atomic radiation, drugs and malignant tumors. 2- Microcytic Hypochromic Anemia: This type of anemia is characterized by MCV less than 80 3 and MCH less than 25 pg. The main cause of microcytic hypochromic anemia is iron deficiency. Causes of iron deficiency: a. Deficiency of iron in diet: This may be due to decreased intake of iron in diet or increased demand for iron as during pregnancy. b. Deficient iron Absorption: This may be due to loss of gastric HCl (as in partial gastrectomy) excess intake of phytic acid and phosphates and oxalates, or diseases of the small intestine. c. Chronic blood loss: This can occur for example in ankylostoma infestation or from peptic ulcers, piles or excess menstrual blood loss. 3- Macrocytic Anemia: This type of anemia is characterized by MCV greater than 95 3. Causes of Macrocytic anemia (megaloblastic anemia): a. Folic acid deficiency: This can be due to deficiency in diet, increased requirements as in pregnancy or failure of absorption due to small intestinal diseases. 16  b. Vitamin B12 deficiency: This is usually due to defective absorption as after gastrectomy, lack of intrinsic factor or diseases of the small intestine. c. Pernicious anemia is a familial disease that is more common in elderly women. It is due to immune reaction leading to destruction of gastric parietal cells. There is deficiency of both HCl and intrinsic factor. Effects of anemia: 1- Hypoxia: decreased oxygen transport capacity of blood leads to oxygen deficiency (hypoxia) at different parts of the body. Manifestations of hypoxia are many e.g. easy fatigue, dyspnea, lack of concentration, poor memory and retarded growth in children. 2- Increased cardiac work. Polycythemia: Polycythemia means increased number of RBCs above normal. Polycythemia causes increased blood viscosity. HEMOSTASIS Hemostasis means the prevention of blood loss after injury. It involves the following three mechanisms: 1- Vasoconstriction of blood vessels at the site of injury. 2- Formation of temporary hemostatic plug: this is the function of platelets. 3- Formation of definitive blood clot: this is the function of blood clotting factors. Local Vasoconstriction Constriction of blood vessels at the site of injury aims at limiting bleeding. The following mechanisms contribute to vasoconstriction: 1- Local myogenic contraction: trauma to blood vessels causes them to constrict, independent of nerve supply or humoral factors. 17  2- Nervous reflex: initiated by pain of the injury. 3- Vasoconstrictor substances (e.g. thromboxane A2 and serotonin) released locally at the site of injury. Formation of Temporary Hemostatic Plug When a blood vessel is injured, platelets form a plug to seal the injury. This plug may be sufficient to stop bleeding if the injury is small. However, larger injuries require clotting of blood to stop the bleeding. Platelets: - Platelets are non-nucleated granulated bodies. They are round or oval in shape and their diameter is about 2-4 m. - Normal platelet count in blood is about 150’000 – 500’000 / mm3. - Platelets are formed in the bone marrow from megakaryocytes. - 70% of the platelets extruded from the bone marrow remain the blood while 30% reside in the spleen. Structure of platelets: 1- Platelet Membrane: phospholipids of the membrane contain “Platelet Factor3 (PF3)” which is important for blood clotting. The membrane contains receptors for collagen fibers and other substances. The membrane has inward invaginations called the open canalicular system. The membrane is surrounded by a glycoprotein layer. 2- Cytoplasmic Structures: the cytoplasm contains the following important structures: a- Contractile proteins: myosin, actin and thrombasthenin. b- Microtubules: form a skeleton of platelet. c- Mitochondria, glycogen granules, cytoplasmic reticulum, golgi apparatus, and lysosomes. d- Enzymes for the synthesis of prostaglandins. e- Two types of granules: i- Dense granules: contain non-protein substances e.g. serotonin, ADP and calcium ions. 18  ii- Alpha granules: contain protein substances e.g. clotting factors, platelet-derived growth factor (PDGF) and platelet activating factor (PAF). Platelet Reactions in Hemostasis: 1- Platelet Adhesion: Injury of blood vessels leads to exposure of subendothelial tissues. Platelets adhere to subendothelial collagen fibers. This adhesion depends on: - Glycoprotein layer of the platelet. - Von-Willebrand factor which is present in plasma and in subendothelial tissues. 2- Platelet Activation: Platelets swell, change in shape and put out pseudopodia. Activation is initiated by binding of platelets to collagen. Activation is stimulated by ADP and thrombin. 3- Platelet Release: This means the release of the contents of platelet’s dense and alpha granules. The important released substances are: - serotonin: which produces vasoconstriction of blood vessels at the site of injury. - ADP: which stimulates platelets aggregation. 4- Platelet Aggregation: Released ADP and thromboxane A2 stimulate platelets to stick to each other. Aggregated platelets release more ADP and thromboxane A 2 which stimulate more aggregation. This self-propagating process continues leading to the formation of platelet plug that closes the site of injury. 5- Platelet Procoagulant Activity: After platelets aggregation, the membrane phospholipid “platelet factor 3 (PF3)” is exposed. PF3 is an ideal surface for the concentration of clotting factors and to start coagulation process. 19  6- Platelet Fusion: High concentration of ADP and other substances released from platelets stimulate irreversible fusion of aggregated platelets. N.B. Platelet-derived growth factor (PDGF) stimulates growth and multiplication of endothelium, smooth muscle and fibroblasts of injured vessels enhancing their repair. ------------------------------------------------------------ BLOOD COAGULATION Blood coagulation means the formation of fibrin clot. Coagulation depends on a number of coagulation factors present in plasma. These factors interact to finally form blood clot. Coagulation factors: Blood coagulation factors are listed in the following table: Factor Name I Fibrinogen II Prothrombin III Thromboplastin IV Calcium 20  V Proaccelerin, labile factor VII Proconvertin, stable factor VIII Antihemophilic globulin IX Plasma thromboplastic component, Christmas factor X Stuart-Power factor XI Plasma thromboplastin antecedent XII Hageman factor XIII Fibrin stabilizing factor HMW-K High molecular weight kininogen Pre-K Pre-kallikrein Ka Kallikrein PL Platelet phospholipids The Clotting Mechanism: Many of the clotting factors are actually proteolytic enzymes that are present in plasma in an inactive form. Clotting mechanism involves activation of these inactive enzymes. Activated enzymes activate other clotting enzymes in a series of cascade reactions. These cascade reactions finally lead to the formation of fibrin clot. 1- The main reaction in clotting mechanism is the conversion of soluble fibrinogen to insoluble fibrin. This reaction is catalyzed by thrombin which is an active proteolytic enzyme that splits 2 pairs of polypeptides from each fibrinogen molecule leaving fibrin monomer: Thrombin Fibrinogen Fibrin monomer + 2 polypeptides pairs 2- Thrombin is present in plasma as inactive precursor prothrombin. Prothrombin is activated by active factor X. Factor X is again present in plasma as inactive enzyme. Activation of factor X can proceed in two pathways: intrinsic pathway and intrinsic pathway. Extrinsic pathway: 21  Injured tissues release thromboplastin (tissue phospholipid TPL). Thromboplastin activates factor VII. Active factor VII activates factors IX and X. Active factor X converts prothrombin to thrombin. This reaction requires Ca2+, PL and factor V. Xa Prothrombin Thrombin Ca2+, PL, V Intrinsic Pathway: Factor XII is activated by contact with collagen fibers in subendothelial tissues. This activation is catalyzed by high molecular weight kininogen (HMWK) and plasma kallikrein. Factor XII can also be activated in vitro by contact with negatively charged wet surfaces such as glass. Active factor XII activates factor XI. Active factor XI activates factor IX. Factor VIII is activated by thrombin. Active factor VIII and IX form a complex. This complex activates factor X. This reaction requires Ca2+ and PL. IXa VIIIa Factor X Active factor X Ca2+ PL Active factor X converts prothrombin to thrombin in a reaction that requires Ca2+, PL and actor V. Xa Prothrombin Thrombin Ca2+, PL, V * Thrombin formed in either intrinsic or extrinsic pathways will convert fibrinogen to fibrin monomer as mentioned before. *Fibrin monomers spontaneously polymerize forming loose mesh. * Loose fibrin mesh is changed to tight mesh by the action of factor XIII (fibrin stabilizing factor). This tight mesh entangles RBCs, platelets and plasma forming a blood clot, *Factor XIII is activated by thrombin in presence of Ca2+. *contraction of actin, myosin and thromboasthenin of platelets causes clot retraction that makes the clot stronger. N.B. 22  1- Thrombin that is formed during blood clotting enhances more blood clotting in the following ways: - Thrombin can activate prothrombin. - Thrombin accelerates the action of factors V, VIII, IX, X, XI and XIII. - Thrombin accelerates platelet aggregation. 2- Intrinsic and extrinsic pathways are not two separate mechanisms for blood clotting. When a blood vessel is injured blood coagulates by both pathways. However, the extrinsic pathway is much faster (about 15 seconds) than intrinsic pathway (about 6 minutes). In addition, there is an interaction between the two pathways: active factor VII of the extrinsic pathway can activate factor IX of the intrinsic pathway. CLOTTING MECHANISM Intrinsic Pathway HMW kininogen Kallikrein XII XIIa HMW kininogen Extrinsic Pathway XI XIa TPL IX IXa VIIa VII PL, Ca++ PL, Ca++ VIIIa TPL 23  X Xa PL Ca++ V Va Prothromin Thrombin Fibrinogen Fibrin monomer XIII XIIIa Fibrin Polymer (threads) Anticoagulants: Anticoagulants are substances used to prevent blood clotting. These substances can be used inside the body (in vivo) or outside the body (in vitro). In vitro Anticoagulants: 1- Oxalate salts: precipitate calcium ions. Citrate salts: de-ionize calcium. 2- Silicon-coated tubes: silicon cannot activate factor XII. 3- Heparin. In vivo anticoagulants: 1- Heparin: Heparin is a naturally occurring anticoagulant. It is formed by mast cells and basophils. Chemically, it is a sulfated polysaccharide with acidic properties. 2- Coumarin derivatives: Examples of these drugs are dicumarol and warfarin. 24  Heparin Dicumarol Origin Mast cells and basophils Plant Mode of action - Facilitates the action of Competitive inhibition of anti-thrombin III. vitamin - Anti-prothrombin. K in the liver, so it inhibits the - Anti-thrombin. formation of factors II, VII, IX, - Prevent activation of factor IX and X, protein C and S Site of action: Both in vivo and in vitro Only in vivo Onset of Rapid onset (minutes) Slow onset (days) action: Duration of Short duration Long duration action: Administration Intravenous/ Intramuscular Oral Antidote: - Protamine sulfate 1% - Vitamin K. - Fresh blood transfusion. - Fresh blood transfusion. Abnormalities of Hemostasis: I. Conditions that cause excessive bleeding: 1- Thrombocytopenic Purpura: it is a disease characterized by excessive subcutaneous small hemorrhages (petechiae). It is due to deficiency of platelets (count below 50’000/cmm) 2- Vitamin K deficiency: Vitamin K is needed for the synthesis of many coagulation factors by the liver. Vitamin K deficiency can occur in: - Obstructive jaundice: in this condition bile cannot reach the intestine. Fat absorption is decreased because it requires bile. Vitamin K is a fat soluble vitamin and its absorption is also decreased. - Newly born infants: the main source of vitamin K is the intestinal bacterial flora. Newly born infant do not have intestinal bacteria yet, so they may have vitamin K deficiency. Surgical operations in infants should better be postponed for one month till bacterial flora is formed. 3- Hemophilia: 25  - Hemophilia is a congenital sex-linked recessive disease. It is carried by females and manifested almost always in males. - It is characterized by bleeding tendency after mild trauma. - There are 3 types of hemophilia: Hemophilia A: due to deficiency of factor VIII. It is the classic hemophilia and the most common type (85% of all cases) Hemophilia B: due to deficiency of factor IX. Hemophilia C: due to deficiency of factor XI II. Conditions that cause excessive intravascular clotting (thrombosis): - Slowing of blood flow in leg veins e.g. during pregnancy, prolonged bed rest after surgical operations, varicose vein. - Roughness of vascular endothelium in atherosclerosis. 26