Lecture 14: Blood And Hemostasis

Question 1

Give the characteristics of erythrocytes

Answer 1

• 4.3 ─ 5 x 103/μL in males
• 3.5 ─ 5 x 103/μL females
• Numbers increase under the influence of erythropoietin:
• Produced by kidney
• Devoid of granules and organelles
• Major contents:
• Lipids
• ATP
• Carbonic anhydrase
• Hemoglobin
• Proteins:
• About 50% are integral membrane proteins
• Peripheral proteins
• Spectrin
• Actin (bound via ankyrin)

Question 2

Give the characteristics of erythrocyte membrane proteins

Answer 2

• Red blood cells (erythrocytes) have been useful for studies of the cortical cytoskeleton.
• They have no nucleus or organelles, so the plasma membranes and associated proteins are easily isolated.
• They also lack other cytoskeletal components, so the cortical cytoskeleton is the principal determinant of cell shape.
• The major structural protein is spectrin, a member of the calponin family of actin-binding proteins.
• It is a tetramer of two polypeptide chains, α and β. The ends of the spectrin tetramers associate with short actin filaments, resulting in the spectrin-actin network.
• Ankyrin links the spectrin-actin network and the plasma membrane by binding to spectrin and a transmembrane protein (band 3).
• Protein 4.1 is another link that binds spectrin-actin junctions and the transmembrane protein glycophorin.

Question 3

Give the characteristics of neutrophils

Answer 3

• Neutrophils are also referred to as polymorphonuclear leukocytes (PMNs):
• Characteristics (Refer to Figure 6-4):
• 7-9 μm
• 3-5 nuclear lobes with connecting strands
• Active amoeboid phagocytes
• Small, numerous specific granules: Contain Lysozyme and other Proteases
• Larger, less numerous azurophilic granules:
• Remain in circulation for 10 –12 hours.
• Live for 1 -2 days after leaving circulation.
• Secrete a class of enzymes capable of destroying certain bacteria by formation of free radicals (superoxide) as well as the release of lysozyme and lactoferrin, which destroy bacterial walls.

Question 4

Give the characteristics of a basophil

Answer 4

• 7-9 μm
• Lobulated nucleus (bilobed)
• Large, membrane-bound basophilic granules:
• Contain:
• Vasoactive substances:
• Serotonin
• Heparin (anticoagulant)
• Kallikrein (attracts eosinophils)
• Can produce leukotrienes:
• Increases vascular permeability
• Slow contraction of smooth muscles
• (Refer to Figure 6-6 Slide 21)

Question 5

Give the characteristics of an eosinophil

Answer 5

• 9-10 μm
• Bilobed nucleus
• Specific granules:
• Major basic protein (MBP):
• Disrupts parasite membranes
• Causes basophils to release histamine
• Peroxidase:
• Cationic protein:
• Neutralizes heparin and is anti-parasitic
• Respond in allergic diseases and parasitic infections.
• Phagocytize antibody-antigen complexes and parasites.
  (Refer to Figure 6-5 Slide 23)

Question 6

Give the characteristics of a lymphocyte

Answer 6

• Large round, sometimes slightly indented nucleus; fills most of cell
• Variation in cell size:
• Small lymphocytes: 6-8 μm
• Medium lymphocyte: 10-12 μm
• Large lymphocyte: up to 18 μm
• B lymphocytes:
• Precursor of plasma cell
• T lymphocyte:
• Precursor of T lymphocytes 25
  (Refer to Fig. 6-7 Slide 27)

Question 7

Give the characteristics of a monocyte

Answer 7

• 9-12 μM
• Largest leukocytes
• Eccentrically located, kidney-shaped nucleus
• Granular cytoplasm due to small lysosomes
• Precursor of macrophages and osteoclasts
• (Refer to Fig. 6-8)
• Slide 30

Question 8

Give the characteristics of platelets

Answer 8

• Derived from megakaryocytes •
• 2 μM
• 200,000 –400,000 per microliter of blood
• Enhance aggregation by release of factors, and they promote clot formation, retraction, and dissolution.
• Repair damage to endothelium by forming platelet plug.
• Adhesion of platelets involves integrins
• Platelets release thromboxane which increases platelet aggregation.
• Endothelial cells release prostacyclin which decreases platelet aggregation.

Question 9

Give the characteristics of hemostasis

Answer 9

• Hemostasis is the elimination of bleeding.
• The most effective mechanisms for hemostasis occur in small vessels such as capillaries, arterioles, and venules.
• Accumulation of blood in tissues is a hematoma.
• Hemostatic sequence of events (in small vessels):
• Constriction of smooth muscles around vessels
• Constriction of vessels
• Slowing of blood
• Formation of platelet plug
• Blood clotting (coagulation)

Question 10

Describe the formation of a platelet plug

Answer 10

• Platelets do not normally adhere to the endothelial cells that line the blood vessel walls.
• In an injury, the endothelial lining is disrupted, exposing the underlying collagen fibers.
• Platelets adhere to the collagen and release the contents of their secretory vesicles, including ADP, and also cause the conversion of arachidonic acid in the platelet plasma membrane to thromboxane A2, which further stimulates platelet aggregation.
• ADP and other factors cause the platelets to aggregate, forming a plug.
• Von Willebrand factor is a plasma protein, released from Weibel-Palade bodies in endothelial cells, that facilitates the adherence of platelets to the walls of the damaged blood vessel.

Question 11

Describe the first step of blood clotting

Answer 11

Activation of prothrombin

• Prothrombinis always found in the blood of normal individuals. It is an inactive form of an enzyme that is activated by Factor XII.
• Factor XII is activated when it contacts collagen in the damaged vessel wall.

Question 12

Describe the second step of blood clotting

Answer 12

Conversion of prothrombin to thrombin

• Thrombin is the active form of prothrombin.
• Thrombin catalyzes the conversion of fibrinogen to fibrin.

Question 13

Describe the third step of blood clotting

Answer 13

Conversion of fibrinogen to fibrin by thrombin

• (Fibrin is a meshwork in which platelets, blood cells, and plasma become entrapped to form the actual clot.)
• Fibrinogen is always present in the blood of normal individuals. It is formed by the liver.
• The fibrin meshwork forms in the presence of Factor XIII, which is also activated by thrombin. Note that many of the clotting components are referred to as “factors.” this is an old term that basically refers to proteins. Also note that factors are not numbered in the order in which they are activated. This is because they were numbered in the order they were discovered.

Question 14

Describe the fourth step of blood clotting

Answer 14

Reshaping of the clot by polymerization of fibrin:

• Fibrinogen is split into a number of polypeptides by thrombin.
• These polypeptides are then chemically linked by the enzymatic action of Factor XIII.
• Erythrocytes and other cells are trapped in this mesh and become part of the clot.

Question 15

Describe the fifth step of blood clotting

Answer 15

Dissolution of fibrin clots through activation of the plasminogen activator system and the action of plasmin:

• This is referred to as fibrinolysis.
• A cascade of protein plasminogen activators convert inactive plasminogen to its enzymatic form, plasmin.
• One of the plasminogenactivators is tissue plasminogen activator (t-PA) which is produced by endothelial cells and circulates in the blood.
• t-PA is a weak enzyme in the absence of fibrin, so fibrin actually initiates its own destruction.
• Plasmin and t-PA dissolve the clot.

Question 16

What are the intrinsic and extrinsic pathways?

Answer 16

• The activation of prothrombin to thrombin, which initiates the clotting process is actually the end product of a number of cascading reactions that involve the sequential activation of circulating inactive proteins (clotting factors) into active enzymes. Each sequence activates the next step in the sequence, hence, the term “cascade.”
• There are two different cascade sequences, each of which leads to the same common pathway. The two initial sequences are referred to as the intrinsic and the extrinsic pathways.
• *Note that this division into two pathways is somewhat artificial, because the two pathways have many interconnections.

Question 17

What initiates the intrinsic pathway

Answer 17

The intrinsic pathway typically is initiated by Injury to the endothelium of the blood vessel exposing collagen fibers. Everything necessary for it to occur is already within the blood, including calcium, required as a cofactor for many of the sequential steps in the clotting cascades.

Question 18

What is the purpose of the extrinsic pathway?

Answer 18

• The extrinsic pathway involves the formation of tissue factor (thromboplastin or Factor III).
• Thromboplastin is a membrane-bound lipoprotein expressed at sites of cell injury; it is derived from the plasma or organelle membranes of damaged cells in the disrupted tissue and enters into the circulating blood.

Question 19

Describe the intrinsic pathway process

Answer 19

• Injury to endothelium of blood vessel exposing collagen fibers leads to the activation of Factor XII (Hageman factor):
• Activated Factor XII activates Factor XI.
• Note also that activated factor XII also converts prekallikrein to kallikrein.
• Kallikrein is involved in the formation of bradykinin (increases vascular permeability) in the kinin cascade and in the conversion of plasminogen to plasmin in the fibrinolytic system.
• Kallikrein can also feedback and activate more Hageman factor
• Activated Factor XI activates Factor IX.
• *Note that thrombin is also involved in the activation of Factor XI and Factor VIII.
• Activated Factor IX combines with activated Factor VIII and calcium to activate factor X.
• **Note that factor VIII is activated by thrombin.

Question 20

Describe the extrinsic pathway

Answer 20

• Damaged cell membranes from injured tissues release thromboplastin into the blood:
• Thromboplastin leads to the activation of Factor VII.
• Activated Factor VII and calcium activate Factor X.
• See Slide 50

Question 21

Describe the common pathway (in blood clotting)

Answer 21

• Either the intrinsic or extrinsic pathways lead to the common pathway:
• Activated Factor X (from either the intrinsic or the extrinsic pathways) combines with activated factor V and calcium to activate prothrombin.
• Note that thrombin activates inactive factor V to active factor V.
• Prothrombin (inactive factor II) thrombin (activated factor II).
• Thrombin with calcium converts fibrinogen fibrin
• Thrombin with calcium also activates factor XIII.
• Activated factor XIII is necessary in the cross-linking of fibrin polymers to stabilize the fibrin gel.

Question 22

What are some additional things to remember regarding hemostasis?

Answer 22

• Remember that most of the clotting factors are synthesized in the liver.
• Liver dysfunction may affect the clotting mechanism.
• Vitamin K is necessary in the synthesis of factors VII, IX and X.

Question 23

Describe leukocyte extravasation

Answer 23

• Homing mechanism is activated by various cytokines released by mast cells, platelets, and damaged tissue cells.
• NO is released by endothelial cells and increases vascular permeability
• Leukocytes (i.e., neutrophils) leave the laminar flow and move toward the endothelium of the vessel wall.
• Two phases involving cellular adhesion molecules:
• Selectin Phase
• Integrin Phase

Question 24

Describe the selectin phase

Answer 24

• Sialyl Lewis-x antigens are oligosaccharide ligands for P-selectin binding found on leukocyte membranes.
• P-selectin appears on the cell surface when endothelial cells are activated by inflammatory signaling.
• Oligosaccharide ligands on leukocytes bind to carbohydrate recognition domains (CRDs) on the P-selectins.
• P-selectins are from Weibel-Palade bodies
• Binding of ligands to the P-selectins causes leukocytes to roll along the endothelium.

Question 25

Describe the integrin phase

Answer 25

• Integrin receptors are activated on leukocyte membrane.
• Bind to ICAM-1 and ICAM-2 (Igsuperfamily) on endothelial cells
• Integrins β1 and β2 are activated on leukocyte membrane and:
• Bind to VCAM and ICAM on endothelial cell membranes.
• Integrins interacting with endothelial ligands promote the transendothelial migration of leukocytes.

Question 26

Describe erythroblastosis fetalis

Answer 26

• This is an antibody-induced hemolytic disease in the newborn that is caused by blood group incompatibility between mother and fetus.
• This incompatibility occurs when the fetus inherits RBC antigenic determinants that are foreign to the mother. ABO and Rh blood group antigens are of particular interest.
• D antigen is the major cause of Rh incompatibility.
• The initial exposure to the Rh antigen during the first pregnancy does not cause erythroblastosis fetalis because IgM is produced and these are too large to cross the placenta. Subsequent exposure to D antigen during the second or third pregnancy leads to a strong IgG response, and IgGs can cross the placenta.
• Rh negmothers are given anti-D globulin soon after delivery of an Rh+ baby. Anti-D antibodies mask the antigenic sites on the fetal RBCs that may have leaked into the maternal circulation during childbirth.
• Hemolysis in erythroblastosis fetalis results in:
• Hemolytic anemia which causes hypoxic injury to the heart and liver leading to generalized edema (hydrops fetalis)
• Jaundice which causes damage to the CNS
• Hyperbilirubinemia