Blood Components and Function

Overview of Blood

• Blood is a fluid connective tissue constituting about 8% of total body weight
• Blood circulates continuously through blood vessels.
• Blood volume is ~5 liters

Components of Blood

• Blood consists of plasma and formed elements

Plasma

• Plasma is the liquid extracellular matrix of blood
• Plasma constitutes ~55% of total blood volume
• Plasma is pale yellow
• Plasma is ~90% water
• Water in plasma determines blood viscosity
• Less water leads to greater blood viscosity and sluggish flow

Formed Elements

• The formed elements are cells and cell fragments suspended in plasma.
• Erythrocytes (Red blood cells or RBCs) carries oxygen
• Leukocytes (White blood cells or WBCs) are part of the immune system
• Platelets are cell fragments involved in hemostasis (blood clotting)

Blood sample layers after centrifugation (separation)

• Spinning a blood sample rapidly in a centrifuge forms three distinct layers
• Top layer: Plasma (~55% of total blood volume)
• Middle layer: Buffy coat (Leukocytes and platelets, ~1% of total blood volume)
• Bottom layer: Erythrocytes (~44% of total blood volume)
• Hematocrit is the percentage of blood volume composed of erythrocytes

Blood functions

• Exchanging gases: Transports oxygen and carbon dioxide
• Distributing solutes: Transports ions, nutrients, hormones, and wastes and also regulates ion concentrations in tissues
• Performing immune functions: Transports leukocytes and immune system proteins
• Maintaining body temperature: Carries away heat generated by chemical reactions
• Clot formation: Seals damaged vessels with blood clots using platelets and proteins, preventing blood loss.
• Preserving acid-base homeostasis: Maintains a stable blood pH in a narrow range (7.35–7.45) via buffering systems
• Stabilizing blood pressure: Blood volume is a major factor

Plasma Proteins

• Plasma contains plasma proteins including Albumin, Immune proteins, Transport proteins, and Clotting proteins
• Plasma proteins form a colloid that makes up about 9% of plasma volume
• Albumin is a large protein synthesized in the liver
• Albumin is responsible for blood's colloid osmotic pressure, drawing water into the blood via osmosis.
• Immune proteins (γ-globulins or antibodies) are produced by leukocytes
• Transport proteins bind to lipid-based molecules, allowing them to be transported in the water-based plasma
• Clotting proteins, along with platelets, stop bleeding from injured vessels by forming blood clots

Plasma miscellaneous

• The remaining 1% of plasma volume contains small molecules dissolved in the water to form a solution
• These molecules are readily exchanged between blood and interstitial fluid in most capillary beds

Erythrocyte (RBC) Structure

• Enables the transport of oxygen and carbon dioxide
• Typical erythrocyte (RBC) is biconcave disc-shaped
• Concave on both sides
• The biconcave shape increases cell surface area for gas exchange
• Mature RBCs are anucleate and lack typical cellular organelles to create space

Hemoglobin (Hb)

• This creates room for enzymes and nearly 1 billion oxygen-binding hemoglobin (Hb) proteins in the cytosol
• Hemoglobin is a large protein
• Hemoglobin has four polypeptide subunits: two alpha (α) and two beta (β) chains
• Each polypeptide chain binds to a heme group containing iron
• Iron binds to oxygen in areas of high concentration, such as the lungs, forming oxyhemoglobin (HbO2)
• This creates a red-colored molecule.
• Hemoglobin releases oxygen in tissues where oxygen concentration is low, such as the systemic capillary beds
• Carbaminohemoglobin: Hemoglobin binds to carbon dioxide when oxygen levels are low which accounts for ~23% of CO2 transport in blood

Erythrocyte Life Span

• Erythrocytes have relatively short lifespan (100–120 days)
• Cells incurs damage due to harsh environment it exists in
• The cells lack the means for repair
• Body has a mechanism for continuously producing new erythrocytes to counter this

Hematopoiesis

• The process in red bone marrow
• Formed elements in blood are produced by hematopoietic stem cells (HSCs)

Erythropoiesis (Red Blood Cell Formation)

• Erythropoiesis is the process that produces erythrocytes from hematopoietic stem cells (HSCs)
• This entire process takes ~5–7 days.
• Erythropoiesis begins when HSCs differentiate into erythrocyte colony-forming units (CFUs) that form only one cell type
• Erythropoietin (EPO) hormone, secreted by kidneys, causes erythrocyte CFUs to differentiate into proerythroblasts
• Proerythroblasts develop into erythroblasts
• Erythroblasts synthesize hemoglobin and other proteins.
• Nucleus shrinks in the erythroblast, is ejected, and becomes a reticulocyte
• Reticulocytes enter bloodstream
• After ejected from bone marrow their remaining organelles exit through pores in sinusoidal capillaries

Erythropoiesis Regulation

• Erythropoietin regulates the hematocrit within a normal range through a negative feedback loop.
• Stimulus: Blood oxygen levels fall below normal.
• Receptor: Kidney cells detect falling oxygen levels.
• Control center: Kidneys increase erythropoietin production and release into the bloodstream.
• Effector/Response: Erythrocyte production increases.
• Homeostasis: Blood oxygen levels rise to normal.

Erythrocyte Death

• Erythrocytes become less flexible and get trapped in spleen sinusoids
• Spleen macrophages digest erythrocytes
• Hemoglobin breaks down into amino acids, iron ions, and bilirubin.
• Heme converts to biliverdin and then to bilirubin.
• Iron ions and amino acids are recycled to make new hemoglobin in red bone marrow.
• Iron ions are transported back to red bone marrow in the bloodstream via the protein transferrin.
• Bilirubin is sent to the liver for excretion.

Anemia

• A condition with decreased oxygen-carrying capacity of blood.
• Three primary causes:
• Decreased hemoglobin
• Decreased hematocrit
• Abnormal hemoglobin
• Variety of events/conditions is associated with anemia.
• Symptoms includes pallor, fatigue, weakness, and shortness of breath.
• Elevated reticulocytes in blood.
• Severe anemia can elevate heart rate.

Decreased Hemoglobin (Anemia)

• Iron deficiency anemia is the most common which is caused by:
• Inadequate dietary iron intake
• Reduced intestinal absorption of iron
• Slow blood loss
• Vitamin B6 deficiency, malnutrition, poisoning with drugs or heavy metals like lead, and pregnancy can all decrease hemoglobin levels

Decreased Hematocrit (Anemia)

• The number of erythrocytes in the blood can be reduced by several factors which lowers hematocrit
• Blood loss from:
• Acute injury
• Stomach ulcers
• Pernicious anemia results from vitamin B12 deficiency
• Erythrocyte destruction can be caused by:
• Bacterial infections
• Diseases of the immune system or liver
• Lead poisoning
• Hemolytic anemia
• Aplastic anemia is caused by:
• Certain medications
• Exposure to ionizing radiation It can also inhibit or stop the production of erythrocytes in the red bone marrow

Abnormal Hemoglobin (Anemia)

• The most common cause of abnormal hemoglobin is sickle-cell disease

Leukocytes (White Blood Cells/WBCs)

• Larger than erythrocytes with a prominent nucleus
• Use bloodstream for transportation
• Do not generally perform their functions within the blood.
• Adhere to blood vessel walls, then squeeze between endothelial cells to enter surrounding tissue

Leukocytes categories

• Granulocytes have cytoplasmic granules released when activated
• Agranulocytes lack visible granules.

Granulocytes Characteristics

• Unusual nuclei consisting of multiple connected lobes
• Contain general lysosomal granules along with specialized granules
• Neutrophils, Eosinophils, and Basophils are three categories based on the color the granules appear (dark purple/red), after staining

Neutrophils

• Neutrophils are the most common leukocyte
• Cytoplasmic granules absorb both dyes
• Cytoplasm is light lilac
• Active phagocytes that ingest and destroy bacterial cells
• Often called polymorphonucleocytes (polys/PMNs)
• Have uniquely shaped nuclei with three to five lobes
• Chemicals released by injured cells attracts neutrophils (chemotaxis)
• Neutrophils exit bloodstream and release granules in the damaged tissue.
• Granules directly kill bacterial cells and attract more neutrophils and leukocytes, enhancing inflammation

Eosinophils

• Eosinophils have bilobed nucleus
• Appear red due to uptake of eosin dye
• Ingest foreign molecules
• Respond to infections with parasitic worms and allergic reactions
• Their granules contain enzymes and toxins that enhance those activities

Basophils

• Basophils are the least common leukocytes
• Have an S-shaped nucleus
• Appear dark purple due to uptake of methylene blue dye
• Mediate inflammation

Agranulocytes

• Includes Lymphocytes and Monocytes
• Lack visible cytoplasmic granules.
• Contain lysosomes, like granulocytes

Lymphocytes

• Lymphocytes are the second most common leukocyte in blood
• Large, spherical nuclei and light blue rim of cytoplasm when stained
• Two basic types of lymphocytes that have similar appearances but have different functions.
• Both cell types are activated by cellular markers (antigens)
• B lymphocytes (B cells) produce antibodies
• The antibodies bind to and remove antigens from tissues.
• Each population of B lymphocytes secretes antibody that binds only to a specific unique antigen
• T lymphocytes (T cells) do not produce antibodies
• Have membrane-bound receptors for individual antigens.
• Activate other immune system components.
• Directly destroy abnormal body cells, e.g., cancer cells or virally infected cells
• Antibodies and T cell receptor cells both bind to only one unique antigen based on their structure

Monocytes

• The largest cells in the leukocyte family
• Have large U-shaped nuclei surrounded by light blue or purple stained cytoplasm.
• The monocytes circulate briefly in the blood
• The monocytes exit the capillaries and enter tissues
• In the tissues, monocytes mature into macrophages
• Macrophages are phagocytic cells that ingest:
  • Dead & dying cells
  • Bacteria
  • Antigens
  • Other cellular debris
• Macrophages activate other components of the immune system by displaying phagocytosed antigens to other leukocytes

Leukopoiesis

• In bone marrow, hematopoietic stem cells form new leukocytes
• HSCs divide and split into two cell lines: myeloid and lymphoid cells line
• The myeloid cell line produces most of the formed elements (erythrocytes and platelets)
  • The myeloid cell line differentiates into blast cells committed to becoming monocytes called monoblasts
  • Granulocytes are derived from myleoblasts that differentiate into precursor cells called promyelocytes
  • Band cells or stab cells are the final stage before mature granulocytes migrate to bloodstream
• The lymphoid cell line produces lymphoblasts
  • These are committed to becoming B and T lymphocytes
  • B and T lymphocytes mature in different locations
  • B lymphocytes mature in bone marrow
  • T lymphocytes migrate from the bone marrow to thymus gland to the mediastinum

Platelets

• Small cell fragments surrounded by a plasma membrane
• The smallest of the formed elements, involved in hemostasis (stops blood loss from injured vessels)
• Platelets lack nuclei and other organelles, but contain several types of granules
• Granules contain clotting factors, enzymes, mitochondria, and glycogen deposits
• Platelets contain cytoskeletal elements (microtubules, actin, and myosin filaments)
• Platelet lifespan is 7-10 days
• These are removed from circulation by the liver and spleen

Platelet Formation

• Platelet formation begins when HSCs differentiate into megakaryoblasts from the myeloid cell line
• Megakaryoblasts develop into megakaryocytes that undergo repeated cycles of mitosis without cytokinesis.
• Megakaryocytes are massive
• Mature megakaryocytes send cytoplasmic extension through clefts in bone marrow sinusoids, stimulated by hormones like thrombopoietin
• Pieces break off into the bloodstream, forming thousands of platelets

Hemostasis

• Hemostasis involves five events that form a gelatinous blood clot, plugging broken vessel.
• Mechanisms limit significant blood loss

Hemostasis - Vascular Spasm

• Begins immediately when a blood vessel is injured
• Blood leaks into extracellular fluid
• Vasoconstriction and increased tissue pressure decrease the blood vessel diameter
• Minimizes blood loss via reducing blood pressure and blood flow locally

Hemostasis - Platelet Plug Formation

• Platelets form a plug/patch that adheres only to injury site to reduce blood loss.
• Injured endothelial cells release von Willebrand factor (vWF)
• vWF is a glycoprotein that binds to receptors on platelet plasma membranes
• Exposed collagen plus von Willebrand factor causes platelets to become sticky
• Platelets adhere to each other and surrounding vessel walls.
• Binding of vWF and collagen to platelets triggers platelet activation
• Granule contents attract and activate nearby platelets, causing them to clump together or aggregate
• Platelet aggregation forms the platelet plug and seals the injured vessel temporarily

Hemostasis - Coagulation

• Forms molecular glue that binds platelets, endothelial cells, and other formed elements
• Coagulation occurs through a cascade of events including:
  • Intrinsic pathway
  • Extrinsic pathway
• The intrinsic and extrinsic pathways converge at a common pathway activating fibrin

Fibrin

• Fibrin is a threadlike protein
• Fibrin converts a soft, liquid platelet plug into a substantial solid mass
• Fibrinogen is an inactive form that is found circulating in plasma and platelets
• Fibrinogen is converted into fibrin
• Fibrin conversion is a series of reactions at the surface of platelets and/or damaged endothelial cells called the coagulation cascade.
• Depends on clotting factors
• Clotting factors are mostly inactive enzymes synthesized in liver and circulate in the blood
• Clotting factors are labeled with Roman numerals in the order of their discovery
• Clotting factors II, VII, IX, and X depend on vitamin K for synthesis

Intrinsic/Contact Activation Pathway

• Named because all involved clotting factors are found in the blood, Pathway activation occurs
• Following sequence of the events
• Inactive protein clotting factor XII comes in contact with exposed collagen fibers
• Exposed collagen fibers activate XII to XIIa
• Factors XI and IX become activated
• XIIa is an enzyme that activates XI
• XIa subsequently activates IX
• In a cascade of events, each factor activated activates another in the pathway
• Factors IXa and VIIIa along w/ calcium ions form an enzyme complex
• This complex activates factor X to Xa to complete the pathway

Extrinsic Tissue Factor Pathway

• Named because it is initiated by a factor outside of the blood.
• Damaged subendothelial cells display a protein called tissue factor.
• tissue factor activates factor from VII to VIIa.
• factor VIIa, tissue factor, and calcium ions form an enzyme complex
• the enzyme complex activates factor X to Xa.

Common Pathway

• Both the intrinsic and extrinsic pathways form factor Xa.
• Factors Xa and Va along with calcium ions form prothrombin activator
• Activator converts; prothrombin into the active form (thrombin)
• Thrombin turns fibrinogen into fibrin
• Fibrin glues the platelet plug together

Hemostasis - Clot retraction

• Actin and myosin fibers in involved platelets contract, bringing edges of wounded vessels closer
• Serum is forced out of the clot
• Serum consists of plasma but without the clotting proteins

Hemostasis - Thrombolysis

• The process that begins after injury has healed and blood clotting is no longer necessary
• Fibrinolysis is the first step in thrombolysis and it breaks down the fibrin glue

Regulation of clotting

• Requires strict regulation
• Endothelial cells regulate the first and second stages of clot formation by producing and secreting chemicals
• Endothelial cells and hepatocytes produce anticoagulants to inhibit coagulation:
• Antithrombin III (AT-III) protein binds and inhibits activity of factor Xa and thrombin.
• Antithrombin also prevents the activation of new thrombin.
• Heparin sulfate: polysaccharide that enhances antithrombin activity
• Protein C catalyzes reactions that degrade clotting factors Va and VIIIa
  • Protein C is activated by protein S

Disorders of Clotting

• A disorder is a condition in which clotting is not regulated correctly and has drastic consequences for maintaining homeostasis
  • Bleeding disorders
  • Hypercoagulable conditions
• Bleeding disorders can increase blood loss from even minor cuts.
• A clotting protein deficiency prevents the blood from clotting (hemophilia).

Hypercoagulable Conditions

• Results in inappropriate clot formation (thrombosis).
• The clot/thrombus is dangerous, obstructing blood flow through a vessel.
• Thromboembolism: When a thrombus breaks off and occludes smaller vessels downstream.
• Thrombi commonly form in deep veins of the legs causing deep vein thrombosis (DVT).
• Pulmonary embolism: dangerous complication of DVTs in which, emboli break off thrombi in legs and lodge in small blood vessels in lungs

Blood transfusions

• Blood taken from a donor is given to recipient
• Blood transfusions were not always the common treatment modality they are now
• Discovery of surface markers/antigens biological molecules and cells (including erythrocytes) lead to safer blood transfusions
• The Antigen is recognized as foreign and the immune system trys to remove them

Blood groups

• Antigens on erythrocytes give rise to this
• Blood groups are genetically determined carbohydrate chains
• Two groups of the 30 different antigens found on erythrocytes are particularly useful for clinical use: (1) ABO blood group (2) Rh blood group

Blood Typing (ABO)

• Features two antigens: A and B antigens
• Giving rise to four ABO types: a. In Type A, only the A antigen is present on erythrocytes. b. In Type B, only the B antigen is present on erythrocytes. c. In Type AB, both the A and B antigens are present on erythrocytes. d. In Type O, neither A nor B antigens are present on erythrocytes. There is no O antigen.

Rh Blood Group

• Rh antigen was first discovered in rhesus monkeys.
• Individuals with Rh on their erythrocytes are Rh-positive (Rh+).
• Those without Rh antigen are Rh-negative (Rh-).
• The ABO and Rh blood groups combined to create eight common blood types.
• Type O+ is the most common blood type in the U.S. populations while AB- is the least.

Blood Typing in Lab

• Blood typing uses antibodies to bind to individual antigens on erythrocytes
• When antibodies (agglutinins) bind to surface antigens
• The surface antigens clump together or agglutinate.
• Agglutination promotes erythrocytes destruction (hemolysis)
• A blood sample is treated with three separate antibodies in order to determine blood type
• If agglutination occurs in response to an antibody, antigens are present
• That specific antigen is absent when no agglutination occurs
• The three antibodies used are: a. Anti-A antibodies bind and agglutinate A antigens b. Anti-B antibodies bind and agglutinate B antigens c. Anti-Rh antibodies bind and agglutinate Rh antigens

Blood Transfusions:

• Antigens and antibodies are the basis for blood matching
• Blood taken from a donor screened for compatibility prior to administration to a recipient
• A match occurs if donor blood type is compatible with recipient blood type

Transfusion Reaction

• During reaction, recipient antibodies bind to donor antigens
• Antibodies cause agglutination, subsequently destroying erythrocytes
• It can possibly lead to kidney failure and death.

Universal Donor

• Universal Donor is has blood type O- because their erythrocytes do not have A, B, or Rh surface antigens
• This blood type can be given if no blood matching is available
• Universal Recipient has blood type AB+
• This individual with blood type does not make antibodies against the A, B, or Rh antigens
• Matching is still the safest practice