Physiology 2.02 - Blood and Hemostasis

2.02

PHYSIOLOGY

AUGUST 6, 2013

BLOOD AND HEMOSTASIS Jean Yazmin C. Nulud, M.D.

OUTLINE: I. Overview of blood tissue A. Functions B. Composition C. Erythrocytes D. RBC Genesis E. Regulation of RBC Production F. Iron Metabolism and Storage G. Hemoglobin-Oxygen dissociation curve II. Blood Transfusion A. Blood Typing 1. ABO blood group 2. Rh blood group B. Blood Components/preparations C. Transfusion reactions/complications III. Coagulation and Hemostasis A. Clot Formation B. Clot Lysis or Fibrinolysis C. Tests for blood coagulation

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Hematocrit- percentage of total blood volume that is occupied by packed RBC; cellular portion of blood (normally 40-45%) Pluripotent Stem Cell- ancestral cell from which committed stem cells develop, can remain as a pluripotent, or it can differentiate into lymphoid or myeloid stem cell Myeloid stem cell- cell that ultimately develops into erythrocyte, megakaryocytes, monocytes, macrophages, and granulocytes (neutrophils, eosinophils, basophils) Megakaryocytes- parent cell of platelets; can fragment into thousands of platelets

*Italicized – From recording   

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OVERVIEW OF BLOOD TISSUE Connective tissue with liquid matrix (plasma) Flows within the cardiovascular system Volume = 71ml/kg (5L in a 70 kg adult) 55% is fluid; 45% formed elements

FUNCTIONS: Primary Functions  Transport of molecules  Exchange between tissues and internal/external environment Secondary Functions  Immunity  Thermoregulation * During cold or hot weather, blood vessels undergo vasoconstriction and vasodilation.  Fluid volume homeostasis  pH homeostasis – blood as an ACID-BASE BUFFER

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COMPOSITION       

ERYTHROCYTES undergo change of shape; do not overstretch *Even deformation or excessive stretching doesn’t necessarily rupture the red blood cell. shape: biconcave disc 5.2 million/cubic milliliter (male) 4.7 million/cubic milliliter (female) diameter = 7.8 micrometers thickness = 2.5 micrometers at the thickest point and 1 micrometer or less in the center Life span - 120 days (4months) After 120 days, recognized as senescent cells by reticuloendothelial system and is broken down, iron is transferred back to plasma

FUNCTIONS:  Transport of O2 and CO2  Control of blood pH

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Plasma (55%) is the fluid portion of blood within which the cellular elements are suspended. Cellular elements (45%) include the RBC, WBC and platelets.

TRANSCRIBERS: Maja, Von, Catie, Kenan, Eli, Rissa, Trisha, Eunika

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BLOOD AND HEMOSTASIS

PHYSIOLOGY GENESIS OF RBC

1. Proerythroblast 1st cell identified belonging to line of RBC do not have hemoglobin (Hgb) basophilic (blue) 2. Basophil erythroblast first-generation cells have accumulated little Hgb tint is bluish (basophilic) 3. Reticulocyte nucleus condenses to a small size, and its final remnant is absorbed or extruded from the cell with 34% hemoglobin cells pass from the bone marrow into the blood capillaries by diapedesis (squeezing through the pores of the capillary membrane) 4. Erythrocytes * During the stage of polychromatophil erythroblast up to reticulocyte stage, even up to mature erythrocyte, you can see the tint is becoming reddish. Meaning hemoglobin is being accumulated inside the cell. Where does RBC come from? Mature RBCs come from bone marrow. DEVELOPMENT STAGE Embryonic life Mid-trimester Adult life

Areas of the body that produce RBC Yolk sac Liver, spleen, lymph nodes Bone marrow

REGULATION OF RBC PRODUCTION Tissue oxygenation if tissue oxygenation is low, the body will signal the production of erythropoietin (principal stimulus) EPO- 90% in kidneys; 10% in liver

* Low blood volume  caused by trauma, hemorrhage or massive bleeding * Pulmonary disease  binding of oxygen to hemoglobin is decreased  

Vitamin B12 and Folic Acid – maturation of nucleus of RBC; normal DNA synthesis Deficiency results in MEGALOBLASTIC ANEMIA vitamin B12, folic acid, and intrinsic factor from the stomach mucosa  loss of any one of these can lead to slow reproduction of erythroblasts in the bone marrow erythroblasts cannot proliferate rapidly enough to form normal numbers of red blood cells, those red cells that are formed are mostly oversized, have bizarre shapes, and have fragile membranes IRON METABOLISM AND STORAGE

* RBCs produced in the bone marrow. Releases circulating erythrocytes. As RBCs age, it becomes senescent RBCs and hemoglobin is being degraded. Divides into heme and globin components. Globin becomes part of the amino acid pool. Heme releases iron. Iron bound to transferrin, will either go to the liver to be stored or to be transferrred to bone marrow for utilization. This is what you call recycling of iron. 1. Dietary iron absorbed in small intestine 2. Iron binds with apotransferrin (protein responsible for iron transport in plasma) 3. Apotransferrin becomes transferrin when it goes to plasma 4. Plasma transferrin excess will be deposited to liver 5. Iron will unbind from transferrin 6. Iron binds with apoferritin 7. Ferritin is stored in liver BODY IRON CONTENT Adult Male (80 Adult Female kg) (60 kg) Hemoglobin 2500 mg 1700 mg Myoglobin and 500 mg 300 mg enzymes Serum iron 3 mg 3 mg Iron stores 500 -1000 mg 0 – 200 mg

TRANSCRIBERS: Maja, Von, Catie, Kenan, Eli, Rissa, Trisha, Eunika

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PHYSIOLOGY

BLOOD AND HEMOSTASIS

HEMOGLOBIN MOLECULE Alpha1 and alpha2 chains Beta1 and beta2 chains Heme moiety 4 iron in ferrous state

* Normal adult: Hemoglobin A  4 globin chains (α1,

α2,

β1, β2) Heme group contains protoporphyrin ring, which binds iron. Iron is responsible for binding oxygen. 1 Heme group = 8 molecules of Oxygen (4 O2) Notes from Ganong: In normal adult human hemoglobin (hemoglobin A), the two polypeptides are called α-chains, each of which contains 141 amino acid residues, and β-chains, each of which contains 146 amino acid residues. Thus, hemoglobin A is designated α2β2. Hemoglobin binds O2 to form oxyhemoglobin When blood is exposed to various drugs and other oxidizing agents in vitro or in vivo, the ferrous iron (Fe2+) that is normally in the molecule is converted to ferric iron (Fe3+), forming methemoglobin Carbon monoxide reacts with hemoglobin to form carbon monoxyhemoglobin (carboxyhemoglobin) The blood of the human fetus normally contains fetal hemoglobin (hemoglobin F). Its structure is similar to that of hemoglobin A except that the β-chains are replaced by γ-chains; that is, hemoglobin F is α2γ2. HEMOGLOBIN-OXYGEN DISSOCIATION CURVE

1. Hgb molecule has conformational shapes Relaxed phase- O2 is bound to Hgb Taut Phase- involved in deoxyhemoglobin (no O2 attached to Hgb); have salt bridges 2. In tissues with decreased O2 = decreased pH If decreased pH, by Bohr Effect = Hgb delivers more oxygen in tissues [with decreased pH] In decreased pH, beta chains release O2 readily; Alpha chains bind O2 more tightly RBC produce 2, 3 biphosphoglycerate (BPG) affects the affinity of O2 to Hgb 3. 2, 3 BPG- binds both beta chains and stimulates the O2 from alpha chains to be released 2, 3 BPG is important for it decreases the affinity of Hgb to O2 thereby releasing more O2 to tissues One Hgb molecule contains 4 oxygen molecules More tissue oxygenation ↑ pH, ↓ 2, 3 BPG, ↓ tissue oxygenation ↓ pH, ↑ 2, 3 BPG, ↑ tissue oxygenation Increasing temperature decreases the affinity of hemoglobin for oxygen, ie. increases the P50 value. Increasing acidity, or pH, does the same thing. This behavior linked to pH is known as the Bohr Effect. Unlike hemoglobin, myoglobin displays a hyperbolic curve of loading and unloading oxygen, not sigmoidal. The curve for hemoglobin is shifted far to the left (lowered P50) when all diphosphoglyceric acid (DPG) is removed. This is called "stripping". The addition of DPG moves the curve to right, decreasing affinity for oxygen and increasing the P50 value. BLOOD TRANSFUSION Procedure done before Blood Transfusion: A-B-O Blood and RH typing Cross Matching

Blood Type Genotype RBC Surface Antigen Plasma Antibodies Can receive blood types:

ABO BLOOD GROUP A B AB AA, AO BB, BO AB

O OO

A

B

A, B

None

Anti-B

Anti-A

None

Anti-A, Anti-B

A, O

B, O

All types

O

The membranes of human red cells contain a variety of blood group antigens, which are also called agglutinogens. Antibodies against red cell agglutinogens are called agglutinins. Blood Typing Agglutination Blood type AB- Universal recipient (no antibody in serum) Blood type O- Universal donor (no antigen in RBC surface) IgG - 2 binding sites; can bind 2 RBCs Theoretically, we can transfuse any blood type to newborns ABO Antibodies are not produced in the womb No agglutinins found in the plasma of newborn Antibodies reaches a peak at 10 years old and continuously decrease

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BLOOD AND HEMOSTASIS

PHYSIOLOGY

FFP (Fresh Frozen Plasma) blood clotting disorders, coagulopathies

Cryoprecipitate

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RH BLOOD GROUP D-antigen = Rhesus (+); DD, Dd d-antigen = Rhesus (-); dd Filipinos: mostly Rh (+)

200-250

Plasma proteins – coagulation factors, proteins C and S, antithrombin

↑ by 2%

10-15

Cold insoluble plasma proteins, fibrinogen, factor VIII, vWF

Topical fibrin glue Given to volsensitive px (with cardiac and renal prob)

HEMOGLOBIN Normal Hb (Hemoglobin) = 12-15 mg/dl Hb = 7 mg/dl → PRBC TRANSFUSION Hb = 10 mg/dl → give Fe supplements (oral ferrous sulfate) Hb = 8-9 mg/dl → Iron Sucrose IV

RH INCOMPATIBILITY

Situation: Rh-negative mother (dd) Rh-positive father (DD) Rh-positive fetus (Dd) 1st pregnancy Sensitization phase Normal pregnancy, baby is alive Mother develops antibodies against Rh+ antigens Subsequent pregnancies evidence of hemolysis (2nd pregnancy) mother develops more antibodies → anti-Rh cross the placenta to an Rh-positive fetus attacks the baby → hemolytic disease of the newborn → fatal ERYTHROBLASTOSIS FETALIS Rhogam given to Rh-negative pregnant women during 28 weeks of pregnancy, 2nd dose may be given before delivery Characteristics of BLOOD COMPONENTS COMPONENT PRBC (Packed Red Blood Cells) given when blood loss is more than 25%

VOL (ml)

CONTENT

180-200

RBC with variable WBC content and small amount of plasma

50-70 Platelet 200-400

5.5 x 1010 / RD unit >3.0 x 1011/ SDAP product

CLINICAL RESPONSE ↑ Hb: +10g/L or 1mg/dl ↑ Hct: +3%

↑ Platelet: +500010000/ml

TRANSCRIBERS: Maja, Von, Catie, Kenan, Eli, Rissa, Trisha, Eunika

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PHYSIOLOGY

BLOOD AND HEMOSTASIS

TRANSFUSION REACTIONS FREQUENCY OF REACTION EPISODES Febrile 1-4: 100 Allergic 1-4: 100 Acute lung injury (DOB) 1: 5,000 Acute Hemolytic 1:12, 00 Fatal Hemolytic 1: 100, 000 Anaphylactic 1: 150, 000 Infections - Hep B 1: 63, 000 - Hep V 1: 1, 600, 000 - HIV 1 1: 1960 - HIV 2 None Reported Most common: febrile and allergic reactions (1-4 per 100 cases of transfusion) Febrile reaction: nonhemolytic transfusion reaction, occurs in ↑ rate of transfusion Normal rate of transfusion o Whole blood: should not exceed 4 hrs o PRBC: should not exceed 2 hrs Before transfusion: THAW the blood first

3. Coagulation Phase Activation of clotting system and clot formation Activate prothrombin (rate limiting factor) to thrombin Thrombin activates fibrinogen to fibrin → Fibrin clot Clot formation 2 Pathways: INTRINSIC and EXTRINSIC Both pathways will not occur without Ca INTRINSIC PATHWAY blood trauma or contact with collagen Platelet Phospholipid Factors VII, IX, X, XI, XII

BLOOD COAGULATION and HEMOSTASIS HEMOSTASIS Balance between clot formation and clot breakdown Normal conditions: blood is antithrombotic, doesn’t promote coagulation BLOOD COAGULATION

EXTRINSIC PATHWAY tissue trauma Factor VII

Severed/injured BV wall ↓ Local vasoconstriction to stop bleeding ↓ Platelet agglutination ↓ Fibrin appears ↓ Fibrin clot formation ↓ Clot retraction PHASES OF COAGULATION 1. Vascular Phase Vasospasm or vasoconstriction 2. Platelet Phase Platelet Aggregation and Adhesion Seals off the break in BV Platelets are attracted to the exposed collagen

4. Clot Retraction Broken edges of the vessel are being pulled together Growth factors seal off permanently the BV

TRANSCRIBERS: Maja, Von, Catie, Kenan, Eli, Rissa, Trisha, Eunika

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PHYSIOLOGY

BLOOD AND HEMOSTASIS

5. Clot Destruction Brought about by plasmin PLASMIN: lyses clot BV is repaired -

PLATELETS Minute flat disks, ~1-4 μm Anucleated Very active Capable of contracting, have actin and myosin, and THROMBOSTHENIN Secretes: o FIBRIN STABILIZING FACTOR: stabilizes clot o THROMBOXANE: for vasoconstriction

PLATELET PLUG FORMATION 1. Adhesion Exposure to collagen Binding to von Willebrand Factor 2. Accumulation and Aggregation Thromboxane synthesis 3. Surface coagulation Facilitated by platelet phospholipids 4. Consolidation 5. Contraction of the clot via actin-myosin CLOT FORMATION 1. Formation of Prothrombin Activator Activated Factors X and V, and Phospholipids Factor X (prothrombin activator) has a protease activity splitting prothrombin Factor V will accelerate the rate of activity of Factor X, together with phospholipids Intrinsic and Extrinsic pathway occur simultaneously 2. Conversion of Prothrombin to Thrombin 3. Conversion of Fibrinogen to Fibrin Thrombin- removes low molecular weight peptides from fibrinogen to form fibrin monomers Fibrin fibers- influenced by platelet thru fibrinstabilizing factor (XIII) Fibrin fibers produces fibrin clot Fibrin fibers entrap blood cells, plasma, platelets Fibrin clots- blocks the break in the wall Partial Thromboplastin Time (PTT) Isolated prolongation of PTT is indicative of abnormalities of intrinsic pathway and for hemophilia Hemophilia – factor VIII deficiency Prothrombin Time (PT) Isolated prolongation of PT is indicative of abnormalities in extrinsic pathway *Prolonged PT and PTT: problem in prothrombin, thrombin, or fibrinogen -

CLOT LYSIS or FIBRINOLYSIS Conversion of Plasminogen to Plasmin Serum is plasma without clotting factors tPA activates plasminogen Plasmin lysis the fibrin fibers forming fibrin degradation products Inhibitors of Blood Coagulation o Anti-Thrombin III o Heparin co factor II o Activated Protein C o Tissue Factor Pathway Inhibitor

TESTS FOR BLOOD COAGULATION Clotting Time Time of formation of a clot Time for the thrombin to be generated Indicative of problems with clotting factors Bleeding Time Measures the time when there would be casospasm and platelet plug formation Platelet phase Indicative of ↓ platelets; problems with coagulation factors Prothrombin Time and Partial Thromboplastin Time Reflects the clotting factors Elevated in cases of warfarin and heparin administration and oral contraceptive pills intake (hormone therapy) PT: extrinsic pathway PTT: intrinsic pathway and factor VIII *Acquired diseases of the liver will cause bleeding problems BLOOD DONATION and TRANSFUSION Calcium Citrate or Calcium Oxalate Added to blood bags Prevent clotting of the donated blood Calcium Gluconate Given to patient transfused with 3 or more blood bags Activate clotting factors

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PHYSIOLOGY

BLOOD AND HEMOSTASIS

CLOTTING FACTOR I

Fibrinogen

II

Prothrombin

III IV V VII

Tissue thromboplastin Ionized Calcium Proaccelerin Proconvertin

VIII

Antihemophilic Factor A

IX

Antihemophilic Factor B Plasma Thromboplastin Component (PTC); Christmas Factor

Hemophilia B (Christmas disease)

X

Stuart-Prower Factor

Stuart-Prower factor deficiency

XI XII XIII

NAME

Plasma Thromboplastin Antecedent (PTA) Hageman Factor Fibrin-Stabilizing Factor (FSF) Prekallikrein (PKK); Fletcher factor High-molecular-weight kininogen (HMK); Fitzgerald factor

DEFICIENCY (Clinical Syndrome) Afibrinogenemia Hypoprothrombinemia (hemorrhagic tendency in liver disease)

Parahemophilia Hypoconvertinemia Hemophilia A (classic hemophilia)

PTA deficiency Hageman trait

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