Path - Blood and Clots - Exam 3 Flashcards
What is haemostasis?
Haemostasis is the human body’s response to blood vessel injury and bleeding. It involves a coordinated effort between platelets and numerous blood clotting proteins (or factors), resulting in the formation of a blood clot and subsequent stopping of the bleed.
Balance between clot formation (thrombin) and clot lysis (plasmin)
What occurs in a bleeding phenotype and clotting phenotype?
Bleeding:
- increased clot lysis
- decreased clot formation
Clotting:
- decreased clot lysis
- increases clot formation
What is it about vascular endothelium injury that affects haemostasis? I.e. what triggers it?
Following injury there is:
- smooth muscle vasospasm – constriction of blood flow and reduced production of prostacyclin and NO which usually prevent platelet adherence.
- exposure of collagen – initiator of platelet activation (initiates intrinsic coagulation pathway)
- exposure of tissue factor – thrombin generation (initiates extrinsic coagulation pathway)
What triggers constriction of blood flow on vascular endothelium injury?
- direct injury to vascular smooth muscle
- chemical release by endothelial cells/platelets
- reflexes triggered by nociceptors
What is primary hemostasis and what are the steps involved?
Platelet plug is formed to rapidly stop the initial bleeding after injury. Short term control of bleeding.
- platelet adhesion
- platelet shape change
- granule release
- recruitment
- aggregation (hemostatic plug)
- SECONDARY – permanent plug
What is secondary hemostasis and what are the steps involved?
Secondary hemostasis is the process where the platelet plug initially created in primary hemostasis is reinforced by the conversion of fibrinogen to fibrin. Long-term control.
- factor cascade
- fibrin-cross linking at site of injury
What is fibrinolysis and what are the steps involved?
The breakdown of clots
What causes platelet activation and what do they do once activated?
Circulating platelets bind sub-endothelial matric proteins through surface receptors, i.e. collagen and tissue factor.
Once activated they:
- change shape – to form platelet plug and formation of pseudopods to mediate adherence
- release granules (ADP, fibrinogen vWF etc). Some recruit more platelets, fibrinogen starts clot formation.
- express fibrinogen receptor
What mediates ‘bridge’ formation between platelets and what mediated platelet adhesion?
Bridge
- Fibrinogen receptor
- GP IIb/IIIa
- Fibrinogen binds platelets together via receptor
Adhesion:
- von Willebrand factor (vWF)
- binds to collagen and GP1b/IX/V receptors
What are the key roles of platelets?
- adhesion/aggregation to form the primary hemostatic plug
- release okatelt-activating and pro-coagulant molecules from granules
- provide a pro-coagulant surface for subsequent activity of the coagulation activity
What are the steps in coagulation?
- initiation – vessel injury exposes TF – fibroblasts to circulating FVIIa in plasma. These form an active complex
- Vascular spasms
- Platelet plug formation:
- The TF-FVIIa complex activates other clotting factors – this generates small amount of thrombin localized to vascular defect
- Propagation:
a) Thrombin IIa activates:
- plateles
- coagulation proteins required for its own production
b) large scale production of thrombin occurs on platelet surface - Coagulation cascade leads to fibrin formation
- Fibrinogen converted to fibrin, which reinforces and stabilizes platelet plug
- Fibrinolysis activated to localize clot site and then to remove it when injury is healing
What are the steps in fibrinolysis?
- plasminogen converted to plasmin
2. plasmin enzymatically degrades fibrin so clot can be removed
What are the common features of a platelet defect vs clotting factor deficieny:
- Platelet Defect:
a) Excessive bleeding after minor cuts: YES
b) Petechiae: COMMON
c) Ecchymoses: SMALL/SUPERFICIAL
d) Haemarthroses, muscle haematoma: UNCOMMON
e) Bleeding with proceedures: OFTEN IMMEDIATE, VARIES IN SEVERITY
f) Bleeding phenotype: MUCOCUTANEOUS
- Clotting factor deficiency:
g) Excessive bleeding after minor cuts: NOT USUAL
h) Petechiae: UNCOMMON
i) Ecchymoses: MAY BE LARGE/EXTENSIVE
j) Haemarthroses, muscle haematoma: YES
k) Bleeding with proceedures: PROCEDURAL OR DELAYED
l) Bleeding phenotype: DEEP TISSUE BLEEDING
What is important to ask about in clinical history for haemostasis disorder?
a) bleeding history
- timing
- triggers
- severity
b) drug use
c) family history
What sort of things are screened for in the lab in coagulation disorders?
- platelet count
- prothrombin time (PT)
- activated partial thromboplastin time (APTT)
- +/- thrombin time (TT) and fibrinogen
- APTT – instrinsic pathway
- PT – extrinsic pathway
- TT – fibrinogen to fibrin
How are coagulation assays performed?
Performed on plasma
What does prothrombin time tell you?
- Evaluates extrinsic/common pathways:
- II, V, VII, X, fibrinogen
- Reference range established depending on test
- Relatively insensitive to heparin
- . High INR = a calculated value allowing for comparison of PT results between labs. Use as if PT.
What does Activated partial thromboplastin time (APTT) time tell you?
- induces clot
- Evaluated FVII, IX, XI, XII
- Generally remains normal until a single factor is
What does thrombin time (TT) tell you?
- measures conversion of fibrinogen to fibrin
- sensitive to inhibitors such as heparin
What types of fibrinogen assays exist?
- functional – evaluates both quantity and quality
- diluted plasma clotted with very high concentration of thrombin
- reduces impact of inhibitors
- high dose thrombin – time independent of thrombin therefor just functional fibrinogen - immunological – quantity only
- use when functional abnormalities are suspected to get quantity
What do you do if you get an abnormal assay result? (clotting)
- repeat, especially if unexpected or weird
- mixing studies:
- assay performed mixing patient plasma with 1:1 normal plasma
- correction to normal:
factor deficiency - failure to correct:
- suggests coagulation inhibitor
What are some of the potential errors with coagulation assays?
- pre-analytical errors: can result in incorrect diagnosis or inappropriate management
- analytical
- post-analytical
What are some of the pre-analytical errors that can occur with coagulation assays?
- sample volume
- sample handling
- storage – length and temp
- Patient factors:
- age
- gender
- blood group
- pregnancy
- stress/exercise
- anticoagulation
- other pathology
- diurnal
What if lab results are normal but they have a history of bleeding?
Abnormalities in secondary haemostasis more or less excluded:
- moderate/severe factor deficiency unlikely if normal APTT/PT
- significant fibrinogen abnormality unlikely if normal Clauss fibrinogen
Abnormalities in primary haemostasis not excluded
- functional platelet abnormality
- other, i.e. von Willebrand disease
Test fibrinogen to compare Clauss (functional) and antigenic fibrinogen. If normal, do von Willebrand studies (FVIII and vWF)
What would you suspect if everything but APTT was normal (APTT high), but on mixing the APTT becomes normal? What if treatment is effective, but then patient comes back and APTT is still high, and this time remained high on mixing?
Clotting factor deficiency. Common = haemophilia
Treatment: factor replacement
Loss of response to previously effective factor replacement and non-correction on mixing suggests:
Acquired factor inhibitor
Reason: typically alloimmune phenomenon against recombinant FVIII.
With APTT, what will mixing tell you?
- If mixing study full corrects….
- suspect factor deficiency
- VII, IX, XI, XII - If non-correction…
Evaluate bleeding phenotype
a) if bleeding – factor studies (especially VIII) +/- inhibitor assay
b) if clotting – test antiphospholipid antibodies
What would you suspect if someone has a high PT, high APTT and low fibrinogen? How would you check?
Fibrinogen issue. Tests Clauss (functional) and Antigenic fibrinogen independently. If they have low Clauss fibrinogen, this suggests dysfibrinogenaemia (functionally abnormal fibrinogen)
What would you suspect if someone has high haemaglobin and haematocrit, and high PT and APTT?
Test error – citrate:plasma ratio is high. Repeat collection with lower citrate dose is likely to show a normal result.
What are the components of blood
- 55% plasma
- ## 45% erythrocytes
What are two automated test methods for doing a full blood count?
a) Aperture Impedance
b) Light Scatter (laser)
How is haemoglobin measured?
NOT measured by cell sorter
- RBCs lysed and light absorbance is measures
How is the Red Cell Count measured?
number of cells through light source
How is the Hematocrit measured?
– measures % of blood taken up by red cells
- summation of number of pulses and average pulse height
What is the RDW (Red Cell Distribution Width)?
Red Cell Distribution Width (RDW)- measure of the range of variation of red blood cell (RBC) volume
i) RDW-SD – when reported as standard deviation
- calculates the wide at the 20% height levels of the histogram to find MCV
ii) RDW-CV – when reported as coefficient of variation
- reference range = 11.5-14.5%
How are reticulocytes singled out for automated measures?
Why are they measured?
Can target RNA with dye as reticulocytes have more RNA than mature RBC.
Measurement used to check for:
- anaemia
- increased in haemolysis/acute bleeding
- BM failure/CRF/B12/fol/Fe deficiency
How are white cells distinguished from red cells to measure white cell count?
What are some potential errors that could occur despite this distinguishment?
RBCs are discriminated from platelets by size and then lysed.
Potential errors: - large platelets - white cell agglutination - NRBCs (nucleated RBCs) What are some factors that could cause bad results for RCC or WCC?
What are the 5 different white blood cells that can be measured in a WCC and how are they distinguished from each other?
Can get 5 part differential:
- neutrophils
- lymphocytes
- monocytes
- eosinophils
- basophils
Differentiated by:
- volume
- complexity/light scatter
- fluorescence after labeling
What can the mean platelet volume tell you?
Younger platelets are smaller so can tell you about age
What are some factors that could cause bad results for RCC or WCC?
- cold agglutinants - increases MCV and MCHC
- lipemia - high lipid levels causes turbidity and increases Hb and MCH
- Nucleated RBCs - will be counted as white cells
- High WCC - can result in high Hb, RCC and RBC indices
- platelet clumps - decreases platelets but increases WCC
What 2 factors cause cytopaenias (low cell counts)?
a) lower production
b) higher destruction
What are the 3 broad categories of anemia?
a) Microcytic - presence of small, often hypochromic, red blood cells in a peripheral blood smear and is usually characterized by a low MCV (small and pale).
- Iron deficiency
- thalassemia
- chronic disease
- rare causes
b) Macrocytic – large RBCs
- megaloblastic (B12 or folate deficiency) and non-megaloblastic
c) Normocytic – normal MCV, but decreased Hb and hematocrit.
- BM Failure
- Haemolysis
What are some of the clues in blood of iron deficiency?
- thrombocytosis (too many platelets)
- low RCC
- high RDW (RBC volume)
- Blood film: pencils, elliptocytes, target cells.
