BLOOD AND IMMUNOLOGY

REBECCA A. VILLANUEVA, M.D.

Department of Biochemistry & Nutrition
Associate Professor, Diplomate, Fellow, PAAB

RULE OF 2/3
TOTAL BODY WEIGHT

2/3 WATER 2/3/ICF ICF 1/3 ECF

1/3 SOLIDS

2/3 INTERSTITIAL

1/3 INTRAVASCULAR (BLOOD) ~7% BW

BLOOD
viscous fluid with a cellular portion and fluid portion I. Cellular Portion WBC platelets RBC II. Fluid Portion

Plasma
Serum without clotting factors

Properties of Blood
7 10 % of total body weight To = 36.5 C to 37.5 C pH = 7.35 7.45 Osmotic pressure = 7 8 atm Freezing point = -0.537C Sp. Gravity = 1.035 1.075 Viscosity = 4 5 x greater than water

Blood: Functions
I. Transport

A. Respiration
B. Nutrition C. Excretion

D. Fluids, electrolytes, minerals, vitamins

Blood: Functions
II. Regulatory

A. Blood pH
B. Body Temperature C. Body Hormones

D. Body Fluids & Electrolytes

Blood: Functions
III. Protective A. Infections (WBC) Bacterial Parasitic Viral Fungal

Blood: Functions
III. Protective

B. Immune Diseases Allergies Immunity Self / Non-self C. Bleeding (Hemorrhage)

D. Excessive clotting

PLASMA PROTEINS

Major Fractions of Plasma Proteins

1. Fibrinogen

2. Gammaglobulin
3. Prothrombin 4. Blood grouping globulins

5. Pure Albumin a. 100 mL of 25% solution = 1 unit of

blood

Major Fractions of Plasma Proteins

small amount of albumin and globulin large amount of salts

many non-protein substances of low MW

GENERALIZATIONS
Most plasma proteins are synthesized in the liver

Plasma proteins are generally synthesized on membrane bound polyribosomes

Almost all plasma proteins are glycoproteins

GENERALIZATIONS
Many plasma proteins exhibit polymorphism Each plasma protein has a characteristic half-life Levels increase during acute inflammatory states or secondary to certain types of tissue damage

PLASMA PROTEINS
comprise the major parts of solids of plasma separation by Salting Out Method Na+ OR Ammonium Sulfate

PLASMA PROTEINS
Plasma CHON = 7.75 g/dl (6-8)
Albumin = 4.7 5.7 gm%

Globulin = 1.3 2.5 gm%

Fibrinogen = 200 400 mg%

ALBUMIN

ALBUMIN
most abundant, most homogenous in electrophoresis MW - 69,000

Ellipsoidal in shape
150 Amstrom x 38 Amstrom small & compact

Single polypeptide chain (585 a.a.)

with 17 disulfide bonds

ALBUMIN
compact, small in size fast to travel in electrophoresis high solubility reactive groups in the molecule cause albumin to combine reversibly with a great diversity of anions and cations

responsible for most of oncotic pressure of blood

ALBUMIN
N-terminal a.a. Asp

C-terminal a.a.
Leu human,dog & rabbit

Ala other species

IpH = 4.7 More phototrophic group (Proton binding groups)

ALBUMIN
Titration studies show molecule of albumin at ph 7.4

100 COOH groups

58 NH2 groups 16 Imidazole

19 phenolic
22 guanidine groups

ALBUMIN
Major protein of human plasma 4.5g/dl Produced in the Liver 12 g/day 60% of Total Plasma Protein 75 80% of Plasma Oncotic Pressure Initially synthesized as PRE-PRO PROTEIN with a signal peptide

ALBUMIN
Decreased A:G Ratio Liver and Kidney Diseases Malnutrition

Binds various LIGANDS FFA, Calcium, Steroid Hormones Bilirubin, Tryptophan, Copper Drugs: Sulfonamides, Penicillin G Dicumarol, Aspirin

GLOBULIN

GLOBULIN
Fractions: A. Pseudoglobulin B. Euglobulin
glob IpH=5.06 glob IpH=5.12 transport (LP) transport (LP)

globulin IpH=6.0 Immune system-Ab

FIBRINOGEN

FIBRINOGEN: Factor 1
soluble plasma glycoprotein
46 nm in length

MW: 340,000
6 polypeptide chain

(A)2: (B)2;2
A2B2;2

FIBRINOGEN: Factor 1
end highly negatively charged (Asp & Glu residues A, B) B - (-) tyrosine O SO4 residue

Hence:
Contribute to its H2O solubility

Repulsion of termini of other fibrinogen molecule preventing aggregation

Diagram Of Fibrinogen

Fibrin Polymerization

Major Protein Component Of Human Plasma

Concentration Approximate mg/100 ml M.W. Albumin 3 4.5 x 103 68,000 Other Function Osm reg transport of FA, bilirubin CHO 40 70% lipid Not known Lipid transport

Globulin glo lipo

100 350 450 400 900

40 35,000 200,000 450,000 200,000 200,000

2 Glo 2 glyco

CHO

Unknown

Major Protein Component Of Human Plasma

Concentration Approximate mg/100 ml M.W. Other Function

Cerulo plasmin
Prothrombin Glo lipo protein

30

150,000
63,000

CHO
CHO

Copper transport
Blood clotting

600 1,200 350 450 3 20 M 80 90% lipid Lipid transport

Major Protein Component Of Human Plasma

Concentration mg/100 ml lipo protein Transferrin Plasminogen 350 450 40 Approx. M.W. 200,000 450,000 85,000 90,000 Other 40 70% lipid CHO Function Lipid transport Iron transport Precursor of Fibrinolysin Antibody Blood clotting factors

Globulins Fibrinogen

700 1,500 300

150 th 340 th

CHO CHO

HAPTOGLOBIN

HAPTOGLOBIN
Plasma glycoprotein
MW = 100,000 daltons

Alpha-phoretic mobility
Irreversibly binds extracorpuscular Hgb in a tight non-covalent complex (Hb-Hp)

HAPTOGLOBIN
Hb Kidney Excreted in urine or precipitates in tubules (iron is lost) Kidney

Hb+Hp

Hb-Hp Complex

Catabolized By Liver Cells (Iron is Conserved)

HAPTOGLOBIN

CERULOPLASMIN

CERULOPLASMIN
2-Globulin Serum Glycoprotein MW = 160,000 Carries 90% of Cu present in plasma (60% in RBC Superoxide Dismutase)

Deep blue color

Single subunit contains 6 atoms of copper Displays ferroxidase activity

CERULOPLASMIN
Physiological functions of copper: 1. Oxidation of iron 2. Erythropoiesis and Leucopoiesis 3. Bone Mineralization

4. Elastin & Collagen cross-linking

5. Oxidative phosporylation

CERULOPLASMIN
Physiological functions of copper:
6. Catecholamine metabolism

7. Melanin formation
8. Myelin Formation

9. Glucose homeostasis
10.Antioxidant protection of cells

-1 ANTITRYPSIN (-1 Antiproteinase)

-1 ANTITRYPSIN (-1 Antiproteinase)

polymorphic single chain plasma protein (394 AA); MW 52,000

90% of -1 fraction of human plasma synthesized by hepatocytes and mactophages

-1 ANTITRYPSIN (-1 Antiproteinase)

principal serine protease inhibition (serpin or Pl) of human plasma inhibits trypsin, elastase and other proteases genetic defect which leads to deficiency causes emphysema and liver diseases

-1 ANTITRYPSIN (-1 Antiproteinase)

smoking
oxidizes methionine residue

inactivates -1 antitrypsin
increase proteolytic destruction of lung tissue emphysema

HEMOSTASIS

HEMOSTASIS VASOCONSTRICTION

HEMOSTASIS PLATELET PLUG

HEMOSTASIS FIBRIN CLOT

HEMOSTASIS PLASMIN - FIBRINOLYSIS

Modifying Signal

Inactive Enzyme A

10x
Active Enzyme A

Amplification 10x

Inactive Enzyme B

10x
Active Enzyme B

Amplification 100x

Inactive Enzyme C

10x
Active Enzyme C

Amplification 1000x

PLATELET ADHESION & ACTIVATION

ACTION OF PLATELETS
COLLAGEN vWF

PLATELETS SEROTONIN GROWTH FACTOR FACTOR IV

LIPOPROTEINS

FACTOR V 20%
ADP

FACTOR XIII (ALPHA S.U.)

THROMBOXANE A2

Arachidonic Acid
Cyclooxygenase

Endoperoxidase
Thromboxane synthetase Prostacyclin synthetase

Thromboxane A2

Prostacyclin (PGI2)

Thromboxane B2 (platelets)

6 Keto PGF1 (endothelial cells)

PROPERTY STRUCTURE ROUTE ONSET OF ACTION M.O.A. ANTIDOTE USE

HEPARIN Large Acidic polymer Parenteral Rapid (Seconds)

WARFARIN Small lipidsoluble mol. Oral Slow

Activates Antithrombin III Protamine Acute (Days)

Inhibits Vitamin K Vitamin K / Plasma Chronic (Wks. Mos.)

ANTIPLATELET DRUGS
ASPIRIN / NSAIDS
Inhibits thromboxane synthesis by blocking cycloxygenase Used in prophylaxis and treatment of MI, TIA and other thrombotic events

ANTIPLATELET DRUGS
TICLOPIDINE (Ticlide 500mg)
Inhibits ADP Formation
Prevents TIA, for patients who cant tolerate aspirin

ANTIPLATELET DRUGS
CLOPIDOGREL (PLAVIX 75 mg)
A. Inhibits the ADP pathway B. Both are used for people with stent

ANTIPLATELET DRUGS
CILOSTAZOL (PLETAAL)
A. Inhibits Phosphodiesterase use especially with people with intermittent claudication

SERINE PROTEASE INHIBITORS

Aprotinin * Risk of kidney failure

ANTIPLATELET DRUGS
DIPYRIDAMOLE
Inhibits phophodiesterase increasing cAMP Used in prevention of thrombosis in artificial heart valves

ANTIPLATELET DRUGS
SOLUDEXINE (VESSEL DUE-F)

ANTIPLATELET DRUGS
SULFINPYRAZONE
Mechanism unknown

Rarely Used

COAGULATION PATHWAY

Numerical System For Nomenclature of Blood Clotting Factors

Factor I
Factor II

Fibrinogen
Prothrombin

Factor III
Factor IV

Tissue Factor
Ca++

Numerical System For Nomenclature of Blood Clotting Factors

Factor V Proaccelerin, labile factor, accelerator globulin Proconvertin, serum prothrombin conversion accelerator (SPCA), cothromboplastin

Factor VII

Numerical System For Nomenclature of Blood Clotting Factors

Factor VIII Antihemophilic Factor A, Antihemophilic globulin Factor IX Antihemophilic Factor B, Christmas Factor, plasma thromboplastin component (PTC)

Numerical System For Nomenclature of Blood Clotting Factors

Factor X Factor XI Factor XII Stuart-Power Factor Plasma thromboplastin antecedent (PTA) Hageman Factor

Factor XIII

Fibrin Stabilizing factor (FSF), Fibrinoligase

ZYMOGENS OF SERINE PROTEASES

Factor XII Binds to exposed collagen at site of vessel wall injury activated by HMWK & kallikrein Factor XI Activated by Factor XIIa

ZYMOGENS OF SERINE PROTEASES Factor IX Activated by Factor XIa in the presence of Ca++ Factor VII Activated by thrombin in the presence of calcium

ZYMOGENS OF SERINE PROTEASES

Factor X Activated on surface of activated platelets by tenase complex (Ca++, Factors VIIa and IXa) and by Factor VIIa in the presence of Factor III & IV

ZYMOGENS OF SERINE PROTEASES Factor II Activated on surface of activated platelets by prothrombinase complex (Ca++, Factors Va and Xa)

COFACTORS
Factor VIII
- Activated by thrombin

- Cofactor in the activation of Factor X by Factor IXa

COFACTORS
Factor V
- Activated by thrombin

- Cofactor in the activation of Prothrombin by Factor Xa

COFACTORS
Factor III (Tissue Factor)
- Lipoprotein found on the surfaces of extravascular cells that acts as a cofactor for factor VII

Diagrammatic Representation of Prothrombin

NH2 (Gla-XI)
P R O

Arg Thr Arg IIe

T
H R O M B I

COO-

THROMBIN ACTIVATES FACTOR VII FACTOR V FACTOR XIII FACTOR VIII

PROTEIN C

REGULATORY AND OTHER PROTEINS

Protein C Activated to protein Ca by thrombinbound to thrombomodulin; then degrades Factors VIIIa and Va

REGULATORY AND OTHER PROTEINS

Protein S Act as a cofactor of Protein C; both has Gla residues

THROMBOMODULIN Protein on the surface of endothelial which then activates Protein C cells binds thrombin

REGULATORY AND OTHER PROTEINS

PS PC PCa

THROMBI N THROMBOMODULIN
Va

VIIIa

FIBRIN CLOT

FORMATION OF FIBRIN CLOT

Thrombin
Arg Gly

NH3+

FIBRINOPEPTIDE (A OR B)

STABILIZATION OF FIBRIN CLOT

FIBRIN-CH2-CH2-CH2CH2-NH3+ (LYSYL)

NH2-CO-CH2-CH2FIBRIN (GLUTAMINYL)

NH+

FACTOR XIIIa (TRANSGLUTAMINASE)

FIBRIN-CH2-CH2-CH2-CH2-NH-C-CH2-CH2-FIBRIN

FIBRINOLYSIS

DISSOLUTION OF BLOOD CLOT

FIBRIN CLOT
PLASMINOGEN

TPA

FIBRIN CLOT

PLASMIN
Arg Val FIBRIN

SOLUBLE FRAGMENTS

t.PA
t.PA PLASMIN

PAI-1 PAI-2

INHIBITORS

RESPECTIVE INACTIVE COMPLEXES

THROMBOLYTIC AGENTS

ALTEPLASE (tPA)
Large human protein, MW50,000 Produced in bacteria through recombinant DNA

Directly converts Ferin-Bound Plasminogen to Plasmin

THROMBOLYTIC AGENTS

ANISTREPLASE
APSAC (Anisoytated Plasminogen Streptokinase Activator Component)

Prodrug (ANISOYL, Group HYDROLYZED in VITRO)

Slow Release (Long Half Life) Human TPA from recombinant DNA

THROMBOLYTIC AGENTS

STREPTOKINASE
Obtained from bacterial cultures Forms a complex with plasminogen prior to conversion

THROMBOLYTIC AGENTS

UROKINASE
Extracted from cultured human kidney cells

Directly converts plasminogen to plasmin

PLASMINOGEN ACTIVATION VARIOUS STIMULI (+) BLOOD PROACTIVATOR tPA, UROKINASE BLOOD ACTIVATOR (+) (+) (-) ANTIACTIVATORS INHIBITION

(-)
AMINOCAPROIC ACID TRANEXAMIC ACID

STREPTOKINASE

(+)
ACTIVATOR PROACTIVATOR ANISTREPLASE Arg Val

(+) DEGRADATION PRODUCTS

PLASMIN

(+)

THROMBIN FIBRINOGEN FIBRIN

FIBRIN SPLIT PRODUCTS

ACTIVITY OF HEPARIN
HEPARIN
+

ANTITHROMBIN III PROTAMINE (-) SULFATE

HEPARIN ANTITHROMBIN COMPLEX

THROMBIN

XII a

XI a

Xa

IX a

VITAMIN K CYCLE
WARFARIN COUMARIN

DIHYDROQUINONE / VITAMIN K(H2) + O2 + CO2

DITHIOLS (THIOREDOXIN) / X-(SH)2

PROTEIN CARBOXYLASE
VITAMIN K EPOXIDE / VITAMIN K(O) + H3O+ FACTORS II, VII, IX, X PROTEIN C / S

PLURIPOTENT STEM CELL

MYELOID STEM CELL

LYMPHOID STEM CELL

PROERYTHROBLAST
MEGAKARYOCYTES N. MYELOCYTES

ERYTHROCYTE
THROMBOCYTES

PRE B CELL

B LYMPHOBLAST
PMN

B CELL

PROTHYMOCYTE PROMONOCYTES E. MYELOCYTES

B. MYELOCYTES

MONOCYTE EOSINOPHIL
BASOPHIL

T LYMPHOBLAST

T CELL

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