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CLINICAL BLOOD HAEMOSTASIS - COAGULATION

COAGULATION: FIBRINOGEN, PT, BT, TT, APTT.

FIBRINOGEN

Fibrinogen/Coagulation Factor I is a plasma glycoprotein produced by the liver. Fibrinogen is the action substrate for both thrombin (final coagulation enzyme) and plasmin (fibrinolytic system enzyme). Fibrinogen belongs to the acute phase proteins (appearing at 24-48 hours post-event). 

Defective fibrinogen levels show liver disfunction. Fibrinogen levels help assessing the body's ability to form a blood clot.  

While coagulation approaches its end, soluble fibrinogen is transformed into insoluble fibrinogen fibres. Fibrinogen fibres protect the wound by intercalating; along with the platelets, they form a clotted blood barrier that blocks bleeding until healing. 

Fibrinogen analysis measures the soluble amount of Factor I (fibrinogen dissolved in the blood), before its conversion into insoluble fibrin.

Fibrinogen normal levels: 200-400 mg/dL (according to the laboratory reference values - reactive used etc.).
 
 
PROTHROMBINE TIME (PT; INR): 

Prothrombin time (PT) evaluates the activity of factors involved in the extrinsic and common coagulation pathway. Evaluates both the activity of vitamin K-dependent coagulation factors (except F IX), factor V and fibrinogen, as well as the function of liver protein synthesis, with diagnostic and therapeutic implications. 

In vitro, main route of blood coagulation initiation: the extrinsic system. This includes blood components and vascular elements, the initiation of coagulation occurring when the tissue factor (FT) is bound to F VIIa. 
The F VIIa - FT enzyme complex activates both F IX and F X. 
F Xa interacts with its cofactor F Va, forming the prothrombinase complex; the complex: enough to generate few thrombin amounts next to FT-expressing cells. 

In vitro, PT detects clot formation. This represents fibrin polymerization, resulting from thrombin action.
Being absent in normal plasma, the tissue factor (FT) must be provided from an external source. This is why the cascade of enzymatic reactions triggered in PT is known as the "extrinsic pathway".  

Prolonged PT indicates deficiency of coagulation factors (I, II, V, VII, X) or an inhibitor's presence. 

PT is the most commonly used test monitoring oral anticoagulant therapy. Results may be expressed as follows:
- as clotting time - in seconds; PT - time (s); normal: 11-13.5 s;
- as a percentage (%) of normal prothrombin activity; measurable range: 12.5-120%; AP - volume fraction (%)
- as prothrombin ratio (PR = PT patient in seconds / normal plasma PT in seconds); PR - prothrombin time ratio (1)
- as INR - International Normalized Ratio - (1); normal: 0.8-1.1.
INR = (PT patient / PT normal plasma) x ISI; ISI: international sensitivity index of used thromboplastin, calculated in report to reference thromboplastin for which ISI = 1. 

INR = 2.0-3.0 target of oral anticoagulant treatment for most clinical cases;
INR = 2.5-3.5 target in anticoagulant treatment for: recurrent deep vein thrombosis, recurrent systemic embolism, cardiac stent, mechanical heart valves.


BLEEDING TIME (BT/CT)

A test that investigates primary haemostasis, thus being an indicator of the vascular and platelet phases efficiencies. 

BT depends on the:
- platelets (function and number)
- adhesion plasma proteins
- vascular wall matrix integrity
Critical values:  >15 minutes. 

Bleeding Time is increased when platelet levels are low or when platelets are qualitatively abnormal. 


THROMBIN TIME (TT)

TT measures the time of fibrin formation under the action of thrombin, and its aggregation to form insoluble clot - the final coagulation step. Assesses the activity of fibrinogen.

Under thrombin action on fibrinogen, fibrinopeptides A and B are released. Upon their cleavage, fibrin monomers form soluble aggregates. 
Due to the action of Factor XIII (activated by thrombin), along with that of Ca ions, a transverse polymerization of fibrin monomers takes place, with the formation of insoluble fibrin. 


Thrombin Time mainly reflects the function and interaction between the exogenous thrombin and the endogenous fibrinogen. 

TT measurable range: 13-240 seconds. Thrombin clotting time is generally <22s, between 14-16s (every lab sets its own reference values, according to the reagent kits used; other labs may have: 7,0-12,0s). Critical values: >60s. 


ACTIVATED PARTIAL THROMBOPLASTINE TIME (APTT)

Also called Partial Thromboplastin Time (PTT), is a functional test evaluating both "intrinsic" and "common" coagulation pathways. 
In vitro, the contact system is involved when blood interacts with a foreign surface, such as the cardiopulmonary bypass. 

Kallikrein cleaves HK, releasing bradykinin and kinin-free kininogen (activated HK). 
In vitro, activation of HK is achieved by adding:
- "partial thromboplastin"; composed only of phospholipids; no protein, no TF;
- silica - a surface activator; provides the negatively charged surface.
Than, calcium chloride is added and time expressed in seconds is measured until the clot's formation. 
The so-called "partial thromboplastin" does not contain the tissue factor (protein) needed to initiate coagulation in PT, as in "complete thromboplastin" stage. The silica provides the negatively-charged surface needed for coagulation contact pathway activation. 

The cascade of reactions triggered in PTT were called as "intrinsic pathway" based on the misconception that coagulation is initiated without the addition of any external factors. The external factor involved in initiating coagulation in PTT: the negatively-charged glass surface of the reaction tube; this can be potentiated by adding silica, kaolin, ellagic acid, in what we now refer to as the "activated" partial thromboplastin test.  

Deficiency/inhibition of HK, prekalikrein or Factors XII, XI, IX and VIII causes prolongation of aPTT with normal PT, while deficiencies of "common pathway" coagulation factors (X, V, II, Fibrinogen) can prolong both aPTT and PT. aPTT is not influenced by FVII or FXIII deficiencies. 

Normal aPTT: 21-35s (therapeutic: 2,0-2,5x normal)



COAGULATION: 


"INTRINSIC PATHWAY": prekalikrein, high-molecular weight kininogen-HK, Factors XII, XI, IX and VIII;
"COMMON PATHWAY": Factors X, V, II and I. 


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