Ch.

43 Section III
Plasma Proteins Maintain the Integrity of the Circulatory
System

11/12/2019
J. Brown
 III. Plasma Proteins Maintain the Integrity of the
Circulatory System
Damage to endothelial lining of blood vesselssevering blood vessels blood loss
Subendothelial layer exposed:
basement membrane, smooth muscle cells
Platelets respond to damage  fibrin clot = hemostatic plug
Too much clotting is bad: stroke, ischemia, tissue damage
Regulation: limits clot in size and to area of damage.
Also stability
As vessel heals, clot dissolves: fibrinolysis
Plasma proteins are required
A. Formation of the Hemostatic Plug
1. Platelets: cytoplasm buds off from megakaryocytes (at the 8 nuclei stage)
• form mechanical plugs
• secrete activators of clotting & vascular repair
Non-activated platelets
Invaginated plasma membrane forms canicular open membrane system
increases surface area for receptors & phospholipids (PL)
that accelerate clotting
Microfilaments interior: actin/myosin
3 Types of Granules (secreted during activation)
2. Electron dense: Ca2+, ADP, ATP, serotonin
3. a-granule: heparin antagonist, PDGF, b-thromboglobulin,
vWF, fibrinogen, other clotting factors
3. Lysosomal granules: hydrolytic enzymes
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
A. A. Formation of the Hemostatic Plug
2. Platelet Activation: Adhesion, Aggregation (more platelets recruited), Secretion
of granules: ADP activates
A) Adhesion of platelet to subendothelium at site of injury
(exposed collagen, subendothelial matrix-bound vWF, et al matrix components)
1) GPIa binds collagen (a2b1 heterodimer: integrin)
Platelet changes shape: flat to spherical
long pseudopods extruded (due to
Ca2+ dependent changes in contractile elements)
2)GP1b: binds subendothelial vWF
3) Membrane changes expose GPIIb-IIIa
(aIIbb3 integrin) binds to fibrinogen & vWF
A. Formation of the Hemostatic Plug
2. Platelet Activation: Aggregation
Fibrinogen: protein in platelet granules & circulating in blood
A. Fibrinogen = Factor I: Form 2 triple helices: a b g
joined at N terminal ends by S-S bonds in g chain
a b chains contain Asp & Glu: negative charges
Repel and prevent aggregation to each other
Fibrinogen binds to activated platelets, promotes aggregation
B. Thrombin (Factor II) cleaves fibrinogen N terminal ends
Releases fibrin monomers  aggregate,
form a soft clot
Thrombin also activates platelets:
binds to a specific receptor on platelets
 B. The Blood Coagulation Cascade
Thrombus = clot
Thrombin activation enhances clot formation
serine protease
(serine in the active site)
Clotting factors are either
proteases or cofactors
Either accelerate thrombin
Formation
OR localization
Proproteins: zymogens
Activated by protease cleavage
B. The Blood Coagulation Cascade
Regulation is the key
Proenzymes (serine proteases when activated):
Factors VII, XI, IX and X
Cleave next proenzyme in the cascade
Sequential activation allows acceleration &
Amplification (similar to a kinase cascade)
“a” the activated form
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
B. The Blood Coagulation Cascade
Cofactors:
• Tissue Factor, V and VIII
Serve as binding sites for other factors
• Tissue Factor (Factor III): integral membrane protein
Doesn’t require cleavage to activate
• V and VIII are procofactors: require cleavage
In response to collagen &thrombin,
platelets release vasoconstrictors
• Serotonin
• Thromboxane A2
Reduces blood flow to injured tissue
PDGF is released: stimulates vascular cell proliferation
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
C. The Process of Blood Coagulation
Cascade is triggered when:
Platelets adhere to subendothelial layer.
And at the same time,
Plasma proteins react with subendothelial layer.
Intrinsic (damage induced release of tissue factor)
Thrombin  XI IX  X
and Extrinsic pathways overlap (VIIXI)
Tissue factor (III) VII X
-----positive feedback of Thrombin on VII & XI
PL-phospholipids on platelets & endothelium
Coordinate with calcium to form a complex
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
C. The Process of Blood Coagulation
Extrinsic pathway is first to activate
Intrinsic is activated by Factor VIIa of extrinsic pathway

Common Pathway:
X Thrombin Fibrinogen hydrolysis Fibrin, which
aggregates (soft clot)
Factor VIII  cross linked (hard clot)

Thrombin activates Factor VIII, V, XI, VII, platelet degranulation

amplifies cascades

V & VIII: cofactors, limiting amounts

Act as sites for assembling enzyme-cofactor
complexes on platelet surfaces
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
C. The Process of Blood Coagulation

1. Cross linking of Fibrin

Factor VIII only enzyme in coagulation cascade that is NOT a serine protease
catalyzes transamidation between Gln & lys side chains
on adjacent fibrin monomers
Creates covalent amide bonds

Fibrin network traps platelets & other cells

Forms a plug in vascular wall
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
C. The Process of Blood Coagulation
2. Factor Complexes

VII, IX, X, prothrombin

• all have a domain with carboxylated Glutamate residues
• Requires Vitamin K & occurs in the liver
• Prothrombin & Factor X have 10 or more poly carboxyGlu
• Bind Ca2+ which binds phospholipids on platelets (neg. charge)
• Localizes complexes & thrombin to platelet surfaces
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
2. Factor Complexes
Factor V contains a binding site for Factor Xa & prothrombin
Prothrombin is the substrate for Factor Xa
Complex allows 10,000-15,000 fold increase in rate of prothrombin
conversion
Factor VIII: forms a complex on surface of activated platelets
With IXa and X: increases activation of X to Xa
Tissue Factor III: complex with factor VIIa which activates X
1. Complexes Enhance clotting several 100,000 fold!
Clot forms fast enough to maintain hemostasis
2. Complexes Localize clot formation to site of vascular injury
Some Phospholipids in cell membrane face only the cytoplasm so
cell injury must take place to activate clotting cascade.
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
D. Regulation through Feedback Amplification and Inhibition
1. Role of Thrombin in Regulation
Activation: Activates V, VIII & XI
Activates platelet aggregation
Stimulates factor VIII release from vWF
Cleaves factor VIII to VIIIa
Inhibition:
Thrombin binds to endothelial cell receptor thrombomodulin
abolishes clotting function of thrombin & allows thrombin to activate Protein C, an
Anticoagulant
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
D. Regulation through Feedback Amplification and Inhibition
2. Proteins S and C
Protein C: regulated by cleavage
Active form (APC) binds Protein S
Protein S: Cofactor of protein C: anchors protein S to the clot
through Ca2+/ g-carboxyglutamate binding to platelet PL
APC is a protease which destroys VIIIa and Va by proteolysis
APC stimulates endothelial cells to increase PGI2 reduces platelet aggregation
 III. Plasma Proteins Maintain the Integrity of the Circulatory
System
D. Regulation through Feedback Amplification and Inhibition
3. Serpins: Serine Protease Inhibitors
Present in plasma at high concentrations (10% of plasma proteins)
Limits protease activity in circulation
Limit coagulation and clot dissolution (fibrinolysis)
Each serpin has a binding site for a specific protease: acts as a trap:
enzyme-inhibitor complex
Antithrombin III is serpin for Thrombin. Arg in ATIII binds active site serine in Thrombin (1:1)
Critical because thrombin activates both coagulation and fibrinolysis
Heparin (a glycosaminoglycan) binds to Lys in ATIII: enhances complex formation with Thrombin
Heparin molecule released so can be reused (so it’s catalytic)
ATIII-Heparin also inactivates XIIIa, XIa, Ixa and Xa, but no effect on VIIa or APC
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
F. Fibrinolysis
Clot formation must be limited to plug
Plasminogen: zymogen circulating, binds to fibrin in clot
when activated Plasmin: serine protease: cleaves fibrin
Plasminogen activator (Urokinase) circulating in blood
Plasminogen Plasmin most efficient on clot surface
Protein C (APC) stimulates release of tPA from tissue
How do we prevent plasminogen activators from creating plasmin in circulation????
a2-antiplasmin circulating protease inhibitor: inhibits circulating plasmin
but not clot-bound plasmin
III. Plasma Proteins Maintain the Integrity of the Circulatory
System
F. Fibrinolysis
How do we prevent plasminogen activators from creating plasmin in
circulation????
a2-antiplasmin circulating protease inhibitor: inhibits circulating plasmin
but not clot-bound plasmin