BLOOD

Dr. Hoe See Ziau Department of Physiology Faculty of Medicine University of Malaya
210709 Biomed/Pharm 08/09 Blood Lecture 2

ERYTHROCYTE DISORDERS

Anaemias or polycythaemias

Anaemia
 Reduction

in the Hb concentration below the lower limit of normal range with respect to age, gender and race of an individual  Always associated with pallor (pale)

CAUSES OF ANAEMIA

Insufficient number of RBCs
 RBC

number < 5 X 106/ µl blood anaemia

 Haemorrhagic

Results from blood loss

 Haemolytic

anaemia

Results from RBC rupture, or lyse, prematurely

 Aplastic

anaemia

↓↓ production of RBCs in the bone marrow  Tumors, drugs & chemicals, x-rays

CAUSES OF ANAEMIA

Low haemoglobin content
concentration < 11 – 12 g/dL blood  Hb molecules are normal, but Hb content in RBC < normal  Megaloblastic anaemia  Microcytic anaemia
 Hb

CAUSES OF ANAEMIA

Low haemoglobin content
 Megaloblastic

anaemia

Results from deficiency of vitamin B12 &/or folic acid  Pernicious anaemia
 impaired

absorption of vitamin B12 due to the absence of intrinsic factor that caused by atrophic gastritis and parietal cell loss

Characterised

by many large immature, fragile and dysfunctional RBCs (megaloblasts)

CAUSES OF ANAEMIA
 Low

haemoglobin content
anaemia

Iron-deficiency
 Results

from inadequate intake of ironcontaining food & impaired iron absorption  Most common cause of microcytic anaemia  The RBCs are smaller and paler (hypochromic) than normal

CAUSES OF ANAEMIA

Normal Blood Film

Megaloblastic Anaemia • large immature and dysfunctional RBCs • hypersegmented or multisegmented neutrophils

Microcytic Anaemia • RBCs are smaller and paler

CAUSES OF ANAEMIA

Abnormal haemoglobin
 Due

to a genetic mutation that alters the Hb chain  Thalassaemia

One of the globin chains is absent or faulty

 Sickle-cell

anaemia

Caused by the abnormal HbS formed – results from a change in one of the amino acids in a βchain of the globin molecule  In low O2 concentration condition, HbS molecules interact with each other to form fiberlike structure & cause RBCs to become sickle shape

ANAEMIA
Sickle cell anaemia
Sickle-shaped RBC

Normal RBC

POLYCYTHAEMIA
 

Hb concentration > 16 – 18 g/dL or RBC number > 8 x 106 /µL blood
Cancer of the bone marrow causing ↑ production of RBCs – polycythaemia vera (primary polycythaemia) Conditions causing hypoxia (through EPO) – secondary polycythaemia:
 Natives  Heart  Lung

living in high altitudes

disease disease

ERYTHROCYTE SENDIMENTATION RATE (ESR)

When blood to which an anticoagulant has been added is allowed to stand in narrow tube, the RBCs form a pile of aggregates (rouleaux) Rouleaux gradually sediment leaving a clear zone of plasma above The length (mm) of the column of clear plasma after one hour is the measure of ESR Normal ESR values:
 

Male: 3 – 5 mm Female: 4 – 7 mm

FACTORS AFFECTING ESR

Surface area of falling RBC

Reduced by rouleaux formation, thus ESR ↑
Increased plasma proteins (fibrinogen, 2- and globulins) → ↑ ESR ↑ RBC count → ↓ ESR ESR is low in sickle cell anaemia and congenital spherocytosis because abnormal RBCs fail to form rouleaux

Composition of plasma proteins

RBC count

Size and shape of RBC

IMPORTANCE OF ESR
  

Non-specific indicator of the presence of organic diseases Useful in the evaluation of treatment of various rheumatic diseases Not used for diagnosis of diseases

LEUCOCYTES / WHITE BLOOD CELLS (WBCs)
Nucleated cells  Bigger than RBCs  4,000 – 11,000 / µL blood  Crucial role in body’s immune defense system  Five types of WBC with varying structure, function and number

TYPES OF LEUCOCYTES

The 5 types fall into 2 categories:

DIFFERENTIAL WBC COUNT
Types of WBC
Neutrophils Lymphocytes Monocytes Eosinophils Basophils Cells / µL 3000 – 6000 1500 – 4000 300 – 600 150 – 300 0 – 100

Percentage Distribution
60 – 70 % 20 – 40 % 2–6% 1–4% 0 – 0.5 %

LEUKOPOIESIS

CHARACTERISTICS OF LEUKOCYTES

Chemotaxis

Attracted to chemicals from pathogens or damaged tissues

Amoeboid movement

Allows leukocytes to move through tissues

Diapedesis

Leave capillaries by squeezing themselves between endothelia cells and enter tissues

Phagocytosis

Remove pathogens and cell debris by engulfing them

CHARACTERISTICS OF LEUKOCYTES

GRANULOCYTES
  

Nuclei segmented into several lobes of varying shapes Cytoplasm contains an abundance of membraneenclosed granules 3 types:

Neutrophil  Eosinophil  Basophil

Life span:

<12 hours in transit in the blood before entering tissues  3 – 4 days in tissues

GRANULOCYTES
Neutrophil

 

Lobed nucleus (3 – 6) Purplish granules
Number ↑ (neutrophilia) in acute bacterial infections First WBCs that react to inflammatory responses triggered by bacterial infection

Destroy bacteria by phagocytosis

GRANULOCYTES
Eosinophil


Bilobed, purplish nucleus Dark pink granules (contain histamine, serotonin) Number ↑ (eusinophilia) during allergic diseases (asthma, hay fever, food allergy), skin diseases & parasitic infestations (worms) Inactivate inflammation-inducing mediators released from mast cells during allergic reactions

GRANULOCYTES
Basophil

  

Bilobed nucleus Dark blue granules (contain histamine, serotonin, heparin) Release histamine with IgE antibody signal Number ↑ (basophilia) in inflammatory & allergic reactions

AGRANULOCYTES


Single, large, non-segmented nucleus Few granules 2 types:

Monocytes  Lymphocytes

AGRANULOCYTES
Monocytes

  

Life span: months - years Dark blue-purple , kidney shape nucleus; gray-blue cytoplasm Digests bacteria, antigen & old or damaged cells by phagocytosis Leave the blood after 1 – 2 days & enter tissues to become macrophages Number ↑ (monocytosis) in chronic infections (tuberculosis, syphilis, malaria)

AGRANULOCYTES
Lymphocytes


   

Life span:100 – 300 days Spherical dark purple-blue nucleus; pale blue cytoplasm Body immunity Protection against bacterial & viral infections B Lymphocytes

Produce antibodies against different antigens

T Lymphocytes

Respond against virus infected cells & tumor cells Release chemicals that destroy the victim cells

LEUCOCYTE DESTRUCTION

Aged leucocytes are destroyed in the liver, spleen, bone marrow and lymph nodes by macrophages Large numbers of leucocytes dies in helping to defend the body against infection, forming together with necrotic tissues → pus

LEUKOCYTES DISORDERS

Leukopaenia
 Abnormally low WBC count  HIV infection, radiation therapy,

chemotherapy, glucocorticoids

LEUKOCYTES DISORDERS

Leukocytosis
 An

increase in the number of leukocytes over the upper limits  A modest leukocytosis is normal during an infection  Extreme leukocytosis generally indicates leukaemia

LEUKOCYTES DISORDERS

Leukaemia
 Cancerous

conditions involving WBCs  Two general types
myelogenous leukaemia

Cancerous production of young myelogenous cells in the bone marrow

lymphocytic leukaemia

Cancerous production of lymphoid cells, usually beginning in a lymph node or other lymphocytic tissue

 Acute

and chronic leukaemia

THROMBOCYTES / PLATELETS
    

Small non-nucleated cells (2 – 4 m in diameter) Irregularly shaped; stained dark purple Granules contain serotonin, ADP, von Willebrand factor, fibrinogen Produced in bone marrow from special cells called megakaryocytes 150- 400 x 103 /µL blood


Life span: 7 – 14 days
Destroyed in the spleen and liver

FORMATION OF PLATELETS
Thrombopoiesis

FUNCTIONS OF PLATELETS
 

Prevents bleeding
Coagulation of blood

HAEMOSTASIS

Prevention of blood loss following an injury
4 mechanisms involved:
 Vasoconstriction

(vascular spasms)  Formation of platelet plug  Formation of blood clot  Permanent repair and fibrinolysis

VASCULAR SPASMS
 

First response to vascular injury Vasoconstriction of injured vessel due to contraction of smooth muscle in the wall of vessel Triggered by
  

Local myogenic reflexes (direct injury to vascular smooth muscle) Reflexes initiated by local pain receptors Vasoconstrictors (serotonin, thromboxane A2) released by endothelial cells & platelets

Instantly reduces the flow of blood from the ruptured vessel

FORMATION OF PLATELET PLUG

Injury to a vessel disrupts the endothelium & exposes the tissue collagen molecules Platelets adhere to collagen via von Willebrand factor (vWF) Binding of platelets to collagen triggers the platelets to release ADP & serotonin – platelet activation

Platelet activation causes new platelets to adhere to the old ones – platelet aggregation
Platelet aggregation rapidly creates a platelet plug inside the vessel

Adhesion of platelet induces synthesis of thromboxane A2 – which further stimulate platelet activation & aggregation, and vasoconstriction

FORMATION OF PLATELET PLUG

FORMATION OF PLATELET PLUG

Adjacent undamaged endothelial cells synthesis & release  prostacyclin (PGI2) – profound inhibitor of platelet aggregation  Nitric oxide (NO2) – vasodilator, inhibitor of platelet adhesion, activation & aggregation to prevent the platelet plug from spreading away from the damaged endothelium along intact endothelium

FORMATION OF BLOOD CLOT: BLOOD COAGULATION
    

Transformation of blood into a solid gel – clot or thrombus Consisting mainly protein polymer – fibrin Occurs locally around the original platelet plug The dominant haemostatic defense Three major steps:

 

Formation of prothrombin activator Conversion of prothrombin to thrombin Conversion of fibrinogen to fibrin

BLOOD COAGULATION

(Active factor XII)
Factor XII

Formation of prothrombin activator

Coagulation

(Fibrin stabilising factor)
Ca2+

BLOOD COAGULATION

Brought about via intrinsic (blood) and/or extrinsic (tissue) system
12 clotting factors (proteins) in the plasma (Factor I – Factor XIII; no Factor VI) Most of the factors are synthesised in the liver Under normal condition, all clotting factors are present in the inactive forms in plasma

 

CLOTTING FACTORS

CLOT PATHWAYS

•Occurs when blood is exposed to collagen fibres •Slower •Can be brought about in vitro

Platelet Factor 3 (PF3) & Ca2+ are involved in both pathways

Prothrombin activator

•Triggered by release of tissue factor (tissue thromboplastin) •“shortcut” mechanism

Ca2+

Haemostasis

PERMANENT REPAIR & FIBRINOLYSIS

Contraction of platelet trapped within the clot shrinks the fibrin mesh


Draws the edges of the damaged vessel together
Serum is squeezed from the clot (serum: plasma without clotting factors) Platelet-derived growth factor (PDGF) – stimulates fibroblast migration & fibrous tissue growth → Scar

The clot is gradually dissolved away (fibrinolysis)

FIBRINOLYSIS
 

Dissolution of blood clots Brought about by a proteolytic enzyme – plasmin

Plasmin digests fibrin into soluble fibrin fragments, thereby dissolve the clot
Plasminogen activators

Plasminogen

Plasmin

Fibrin

Soluble fibrin fragments

BLOOD CLOTTING DISORDERS

Haemophilia
 Genetically

inherited clotting insufficiency  Haemophilia A
Classic haemophilia (85 %)  An abnormality or deficiency of Factor VIII
 Haemophilia

B (Christmas disease)

15 %  A deficiency of Factor IX
 Haemophilia

C

Very rare  A deficiency of Factor XI

BLOOD CLOTTING DISORDERS

Haemorrhagic disorders that due to
 liver

disease

deficiency of coagulation factors that are synthesised in the liver (Factors I,II, V, VII, IX and X)

 Vitamin

K deficiency

Synthesis of Factors II (prothrombin),VII, IX and X are reduced  Vitamin K is required as coenzyme for synthesis of these factors

BLOOD CLOTTING DISORDERS
 Thrombosis
 Formation

of clots (thrombus) inside the blood

vessel
 Causes

of thrombosis  Roughed endothelial surface of a vessel caused by arteriosclerosis, infection, or trauma  Blood flow is sluggish and allows activated coagulation factors to accumulate

 Emboli

- freely flowing clots in the blood stream to other organs and cause damage

ANTICOAGULANT AGENTS

Agents that prevent clotting of blood


In vitro and in vivo
Heparin
 Naturally

occurring anticoagulant  Facilitates the action of antithrombin → inactivates thrombin and other proteases involved in blood clotting  Found in the granules of basophils and mast cells  Used in vitro and in vivo

ANTICOAGULANT AGENTS

Chelating agents
 Citrate,

oxalate, flouride, EDTA  Combine with plasma Ca2+ & form insoluble complexes  Used in vitro only

Agents that inhibit the action of vitamin K
 Coumarin

derivatives: warfarin, dicumarol  Clinically used in vivo only as drug for treatment of thrombosis

Arvin (extract from venom of Malayan viper)
 Depletes

fibrinogen by forming imperfect fibrin which is removed by macrophages (defribrination)

BLEEDING TIME
  

A medical test done on someone to assess their platelet function The time it takes for bleeding to stop (as thus the time it takes for a platelet plug to form) is measured Normal value: 2 – 9 min Bleeding time ↑ when:
 Thrombocytopenia

(platelet difficiency)  Disseminated intravascular coagulation (DIC)

CLOTTING TIME
The time taken for blood to clot in vitro  Normal clotting time: 6 – 12 minutes  Clotting time ↑ when:

 Clotting

factors are ↓ than normal  ↓ vitamin K  Chronic liver disease

BLOOD GROUPS

Two major blood group systems:
 ABO

system  Rhesus system (Rh)

Based on the presence of antigens on the RBC membrane

BLOOD GROUP ABO SYSTEM

BLOOD GROUP ABO SYSTEM

Knowledge of blood group is important for transfusion purpose
DONOR Antigen O (anti-A, anti-B) RECIPIENT A (anti-B) B (anti-A) AB (-)

A (A)
B (B) AB (A,B) O (-)

X
X X √


X X √

X
√ X √


√ √ √

BLOOD GROUP ABO SYSTEM
Universal donor

Universal recipient

RHESUS SYSTEM

Based on the presence of antigen D on the RBC membrane Presence of antigen D: Rh + (85 – 90 %)
Absence of antigen D: Rh – (10 -15 %)

RHESUS SYSTEM

Rh negative individuals:
 Plasma

does not contain antibodies to antigen D produce antibodies naturally or spontaneously

 Cannot

RHESUS SYSTEM

Anti-D antibodies can be produced when:
 Rh+  Rh

blood is transfused into an Rh- individual

mother becomes pregnant with an Rh+ foetus
Haemolytic disease of the newborn / erythroblastosis foetalis

RHESUS SYSTEM
2. Rh- individual (no ag. D) 1. Rh+ blood transfusion (1st time)

3. Body produces anti-D antibodies in 2 – 3 weeks
4. Rh+ transfusion (2nd time after 4 – 6 months) 5. Agglutination & haemolysis

RHESUS SYSTEM

1st pregnancy
 

2nd pregnancy

Rh- mother becomes pregnant with Rh+ foetus

Foetal RBC may cross the placental barriers into maternal circulation – mainly occurs during parturition
Induce mother to synthesise anti-Rh antibodies In future pregnancies, these antibodies may cross the placenta to attack and haemolyse the RBC of an Rh+ foetus – cause haemolytic disease of newborn

 

BLOOD TRANSFUSION

The process of transferring blood or blood-based products from one person into the circulatory system of another Main reasons for blood transfusion:
  

To replenish the volume of blood lost during trauma To treat a severe anaemia or thrombocytopenia caused by a blood disease People suffering from hemophilia or sickle-cell disease

BLOOD GROUPING
  

Performed to identify a person’s blood group Antisera containing Anti-A, anti-B or aniti-D antibodies are used A drop of the blood to be tested is placed on a microscopic slide and mixed with the antiserum containing the specific antibody After several minutes, the mixtures are observed under microscope If the blood has become clumped (agglutinated) → antibody-antigen reaction has resulted

BLOOD GROUPING

CROSS-MATCHING
 

Performed to determine the compatibility of a donated unit of blood for its intended recipient Involves testing for agglutination of donor RBCs (antigen) by the recipient’s plasma (antibody), & of the recipient’s RBCs by the donor plasma

Donor O+

Recipient A+

(RBC)

No antigen

Antigen A
Anti-B

(Plasma) Anti-A & Anti-B

During an emergency, O Rh- blood is used for transfusion

COMPLICATIONS OF BLOOD TRANSFUSION
Allergic reactions: fever, chills  Hyperkalemia  Hypocalcemia  Transmission of infectious diseases: AIDS, malaria, syphilis, hepatitis B  Volume overload  Agglutination, haemolysis, jaundice

TRANSFUSION REACTIONS

Resulting from mismatched blood types
 Agglutination
 

and haemolysis of RBCs (particularly those from the donor)
Agglutinated clumps can plug small vessels Haemolysed RBCs release large amount of Hb which are converted to bilirubin. High bilirubin concentration in the body fluid can cause jaundice Excess free Hb in the plasma leaks through the glomerular membranes into the kidney tubules → precipitation of Hb in the tubules and block the tubules → leading to acute kidney shutdown

TRANSFUSION REACTIONS

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