Exam 3: Coagulation and Dissolution of a Blood Clot Flashcards
Overview of coagulation: 3 steps & “3-ish big items that are involved”
(putting into words the initial picture from ppt lecture)
- damaged blood vessel - releases clotting factors
- clotting factors turn prothrombin to thrombin
- thrombin turns fibinogen (soluble) to fibrin (insoluble) - this will go to step 3
- damaged blood vessel - release of clotting factors
- plt plug + vasoconstriction to site (limits blood flow)
- development of clot - fibrin strands adhere to the plt plug to form an insoluble clot
Normal Hemostasis: describe the 3 pathways
- a balance between generation of hemostatic clots and uncontrolled thrombus formation
-
Extrinsic pathway: plasma-mediated, initiation of hemostasis
- AKA primary hemostasis
- key: TF (tissue factor)
-
Intrinsic pathway: amplifies and propagates hemostasis
- AKA secondary hemostasis
- key: thrombin
- ppl with congenital issues can still clot, but they might get into trouble in “dire, stressful situations”
- Common pathway: results in an insoluble fibrin clot
“Some extrinsic force causes you to bleed, then your intrinsic forces must stop the bleeding”
Which one is predominant in hemostasis? In general, we are anticoagulated!
Preop Coag testing
- should be based on pt’s history and planned surgery
- BOARDS QUESTION
- balance between risk of surgical bldg and risk of developing postop thrombembolism
- agents
- antiplatelets: ASA
- anticoagulants: coumadin, Heparin
- thrombolytics: TPA
- procoagulants: FFP
- try to remember the test that goes along with each drug (ex: warfarin will be INR to test levels)
Mechanism of Blood Coagulation
- hemostasis is precisely regulated by interactions between blood vessel walls, circulating plts, and clotting proteins in the plasma
- fibrinolysis is the orderly breakdown of a stable blood clot
- > 50 substances that affect blood coagulation have been identified
- fibrin disc in 1800s
- normally, anticoagulants predominate but when a vessel is ruptured, procoagulants are activated
normal hemostasis steps (4)
- vasoconstriction
- plt plug
- clot formation
- clot dissolution
what is down under the endothelium
TF!
- when there’s a break, tissue factor and thrombin interact
- they signal that something is wrong, so plts start coming
- subendothelial TF initiates plasma-mediated hemostasis via the EXTRINSIC pathway
- the intrinsic pathway further amplifies thrombin and fibrin generation
- plts adhere to exposed collagen to undergo activation, resulting in recruitment and aggregation of additional plts
what is visoelastic testing
TEGS and ROTEMS
- Piston inside of a cup of blood. In one of them the cup moves and in one of them the piston moves.
- But basically spins and tells you how long it takes for a clot to:
1. form
2. become stable
3. dissolve
but the results take 30-45 minutes
what categories do each of these belong to:
plts
coagulation factors
- cells = RBCs, WBCs, plts
- plasma = water, proteins, globulins
- globulins = alpha, beta (coagulation factors), gamma (antibodies)
- plasma = water, proteins, globulins
step 1: vasoconstriction
- normally, vasc endothelium provides a nonthrombogenic surface
- “anti-plt, anticoagulant, pro-fibrinolytic” = all just mean NO CLOT
- Damage to the endothelium exposes the underlying extracellular matric and elicits a contraction (vasoconstriction)
- thrombin, hypoxia and high fluid sheer stress can also induce prothrombotic endothelial changes
- “high fluid sheer stress” = stress over an atherosclerotic plaque, or pulmonary stenosis
“damage → reflex vasoconstriction”
step 2: formation of a plt plug
- when plts are exposed to the extracellular matrix in damaged endothelium, they undergo a series of biochemical and physical alterations (grow arms!)
- 3 major phases
- adhesion - sticky plts!
- activation
- aggregation
tell me about plts
(nml concentration, life span of a plt, etc)
- plt = thrombocyte
- thrombus/clot + cells
- formed in bone marrow
-
nml concentration is 150,000-400,000 per microliter
- spontaneous bldg if <50,000 u/L
- lethal if <10,000 u/L
- life of a plt is 8-12 days
- preop interview! they should have held antiplts for 8-12 days!
name the 5 receptors on plts
- ADP: P2Y12
- TXA2: TP
- collagen: GP1a, GPVI
- vW factor: GP Ib-IX-V
- fibrinogen/ vWF
plt adhesion
- going from lazy river → sticky, with feet
- conformational change which enables plts to stick and not be washed away
- exposure to subendothelial matrix proteins allows plts to undergo a conformational change to adhere to the vasc wall
von Willebrand Factor (vWF)
- vWF is produced in the endothelium and plts. it is released by endothelial cells and activated by plts
- vWF’s primary fn is to bind other protiens
- it’s important as a bridging molecule between the subendothelial matrix and plts, forming cross links
- glycoprotein IIb/IIIa = plt-plt
- reopro, aggrastat, integrilin
- glycoprotein Ib/factor IX/factor V receptor complex = plt-vessel
- (these are the names of the receptors)
- glycoprotein IIb/IIIa = plt-plt
- primary thing that enables binding to wall or other plts
enrichment: GPIb-V-IX complex binds vWF, causing what?
allows plt adhesion (to each other and to the vessel wall) and plt plug formation at sites of vasc injury
absence of the GP1b-5-9 receptor is known as Bernard-Soulier syndrome
vWD - von Willebrand Disease
how it’s activated
incidence
where it shows up
- vWF is mainly activated in conditions of high blood flow and shear stress
- 1/100 ppl, but clinically significant cases 1/10,000
- defic of vWF thereshow show primarily in organs with small vessels, such as skin, GI and uterus
- dx’d by measuring amt of vWF in a vWF antigen assay and the functionaliyt of vWF with binding assays
- factor VIII (factor 8) is also measured (they’re buddies)
- think of woman who has really heavy menstrual bldg, nose bleeds, gums bld easily, frequent bruising*
- could show up in post-partum bldg*
Types of vWD (3 + pseudo)
- Type 1 vWD (60-80%) - failure to secrete vWF into circulation or vWF being cleared more quickly than normal
- mild, often undx’d until bldg following surgery, easy bruising or menorrhagia (abn heavy periods)
- drug that helps them secrete the vWF: DDAVP/desmopressin
- Type 2 vWD (15-30%) - qualitative defect and bldg varies. Has 4 subtypes.
- ↓ ability to bind GP1b
- ↓ ability to bind VIII
- Type 3 vWD - most severe, homozygous defective gene, NO production of vWF
- leads to extremely low levels of VIII, since it does normally protects FVIII from proteolytic degradation
- (NIH - vWF is a carrier protein for FVIII)
- can’t give them DDAVP - they need exogenous vWF
- “Type 3, I have no vWF in me”
- Platelet type - aka pseudo-vWD - this is a defect in the plt’s GP1b receptor
- not technically an issue with vWF
- the plt GP1b is abnormal
We need to know what type of vWD they have
“von Willebrand has EIGHT close mates” - vWF and Factor VIII are best buds
What meds do we use that are GPIIb/IIIa inhibitors? (3)
And what class are they?
What’s their MOA?
- antiplts!
- Reopro = abciximab
- Integrilin = eptifibatide
- Aggrestat = tirofiban
- Blocks the ability of fibrinogen to form around aggregated plts
- as a result, no fibrinogen bridging of plts to other plts can occur
- integrilin and aggrestat T1/2 2-4 hrs, so usually in gtt form
- there’s no antidote, so bldg could occur elsewhere in the body and you’d have to wait it out or give plts
- ex of dr. funk’s when they’d be on the gtts in the ICU but they’d be bldg from IV sites or ETTs
plt activation
- after plts adhere to the damaged endothelial wall, several intracellular signaling pathways are activated when ligands bind to plts receptors and a series of physical and biochemical changes occur
- also pls dev pseudopod-like membrane extensions (FINGERS AND FEET) to ↑ plt surface area
plt recruitment
plts rel granular contents, resulting in recruitment and activation of additional plts
- some of the other contents
- ADP, ATP
- serotonin, histamine
- Ca++
- vWF, FV, VII, XI, XIII
- fibrinogen
- PAI-1, TFPI
- thrombospondin, fibronectin
IIb/IIIa inhibitors:
what makes abciximab (ReoPro) different from eptifibatide (Integrilin) and tirofiban (Aggrastat)
- it binds irreversibly to plts
- time to restore plt fn is 24 hrs, compared to 4-6 hrs for integrilin and aggrastat
thromboxane A2 inhibitors
- Aspirin – inhibits the ability of COX enzyme to synthesize the precursors of thromboxane within platelets
- Naproxen – nonselective COX inhibitor
ADP and plts
- P2Y12 receptors further amplify the response to ADP and draw forth the completion of aggregation
ADP receptor antagonist meds?
(P2Y12)
prodrugs and direct-acting ones
- prodrugs (have to go thru the liver to start working)
- ticlopidine = Ticlid
- clopidogrel = Plavix
- prasugrel = Efient
- direct-acting (more effective)
- ticagrelor = Brilinta
- cangrelor = Kengrexal
- before ADP inhbitiros, the only antiplt on the market was ASA
- researchers began in the 1970’s to look for drugs to target other important signaling pathways
- remember that 15% of pts who metabolize plts differently
plt aggregation
which receptor links what
- plt aggregation completes the formation of a plt plug
- activators released during the activation phase recruit and amplify the response of additional plts to the site of injury
- newly activated glycoprotein IIb/IIIa receptors on the plt surface bind fibrinogen to provide cross-linking with adjacent plts
- covalent bond = strong, tight
image of 2 plts stuck together:
- GP IIb-IIIa = vWF + fibrinogen “net” (plt to fibrinogen)
- GP Ib-IX-V (1b-9-5) = vWF + other vWF (plt to each other)
step 3: formation of blood clot
- Following platelet adhesion, activation and aggregation…the blood clot begins to form
- The plasma protein fibrinogen breaks down to produce fibrin, which subsequently becomes cross-linked into a stable mesh
- Coagulation factors are then activated and initiate the coagulation cascade
- Soluble fibrinogen is converted to insoluble fibrin
- The key step in blood clotting is the conversion of fibrinogen (I) to fibrin (Ia) by thrombin (IIa)
- the body is already incorporating plasminogen into the clot
- thrombin = makes clot strong
- “strong-bin”
- alpha angle on TEG/ROTEM
- MCF - maximum clot firmness
- alpha - leading up to how strong of a clot it’s going to get
- MCF - how strong the clot it
coag cascade summary
- The classic description of coagulation includes initiation of this chain reaction via either the intrinsic or extrinsic pathway
- Intrinsic pathway is triggered when blood contacts a negatively charged surface (i.e. exposed subendothelial collagen)
-
Extrinsic pathway is activated when blood contacts cells outside the vascular endothelium.
- Nonvascular cells express a membrane protein called tissue factor (III) which initiates this pathway
- Common Final Pathway is where the intrinsic and extrinsic pathways converge with the activation of factor X
coagulation factors:
how are they named?
where are they synthesized?
which ones are Vit-K dependent for utilization?
- Were identified using Roman numerals assigned in order of discovery
- Most coagulation factors are enzymes with some exceptions
- vWF and Tissue Factor (III) are glycoproteins
- Most are synthesized in the liver with some exceptions
- Calcium (IV) comes from diet
- Von Willebrand is synthesized in endothelial cells and platelets
- Factors II, VII, IX and X are Vitamin K dependent for utilization (coumadin) **Test question for sure!
- Circulate in an inactive state
- A lower-case letter “a” is the active enzyme
- E.g.: Inactive prothrombin is referred to as factor II and active thrombin is identified as factor IIa
intrinsic pathway of coagulation
- “Contact Activation System”
- Begins with damage to the blood vessels themselves
- Formation of the primary complex on collagen and thrombin generation by way of factor XII and ultimately merges to the common pathway an activates factor X
- The formation of this primary contact in initial clot formation may prove to be a minor role as patients with these severe factor deficiencies do not have bleeding disorders
- May be more involved with inflammation, amplification, and propagation of the hemostatic response to maximize thrombin generation
***TEST***
Which factors are Vitamin-K dependent for utilization?
2, 7, 9, 10
Interchangeable names for :
Factor I
Factor II
Factor III
Factor IV
Factor I = Fibrinogen
Factor II = Prothrombin
Factor III = Tissue Factor
Factor IV = Calcium
Calcium is always 4! Ca-Ch blockers is class IV, and now this!
Extrinsic pathway of coagulation
- “Tissue Factor Pathway”
- Initial step in plasma-mediated hemostasis
- Following damage to the blood vessel, Factor VII comes into contact with tissue factor (which is prevalent in the sub-endothelial tissues surrounding vasculature) and forms an activated TF-VIIa complex
- The TF-VIIa circulating the plasma activates factor X to promote the conversion of X to Xa.
7-(W)TF-Extrinsic - “7 ways to extrinsic-ally cause bleeding, wtf”
For the test, we don’t have to memorize all of this, but we DO need to know where a drug works (ex: heparin works in the coagulation cascade, has nothing to do with plts)
Common pathway of Coagulation
- Common to both extrinsic and intrinsic
- It depicts thrombin generation and subsequent fibrin formation - prothrombin (II) is cleaved by activated factor X to produce thrombin (IIa)
- “10 cleaves 2 (prothrombin) into 2a (thrombin)”
- Signal amplification
- Thrombin activity
- Fibrinogen (I) to fibrin
- activates platelets and factor XIII (fibrin-stabilizing factor)
- converts inactive cofactors V (proaccelerin) and VIII (antihemophilic factor A) to active forms
- activates factor XI (plasma thromboplastin antecedent) and up-regulates tissue factor
- stimulates vascular endothelium to express downregulation of fibrinolytic activity
Ppl with Factor 13 deficiency
can make clots but they won’t stay
Blood clot
(somewhat repetitive but it had its own slide for reinforcement)
- Prothrombin gets activated to thrombin
- Thrombin activates fibrinogen to form fibrin
- Fibrin creates covalent bonds and cross-linking of fibers create a meshwork in all directions of blood cells, platelets and plasma which adhere to the surface of damaged blood vessel
- After a clot is formed, the actin and myosin of the platelets trapped in the fibrin mesh interact in a manner like that in muscle contraction
- Ex: stroke tx window is BEFORE it’s stronger, before it contracts down
cobwebs and clotting
• In traditional European medicine, cobwebs were used on wounds and cuts and seem to help healing and reduce bleeding. Spider webs are rich in vitamin K, which can be effective in clotting blood. Webs were used several hundred years ago as gauze pads to stop an injured person’s bleeding.
dissolution of blood clot
- Clot lysis occurs when plasminogen (normally found in the blood) is activated to plasmin
- Activation occurs by tissue plasminogen activator (t-PA) released from the tissue, vascular endothelium, plasma, and urine (urokinase)
- Plasmin is an enzyme (resembles trypsin in pancreatic secretions) which digests fibrin fibers, fibrinogen, Factor V, Factor VIII, and Factor XII
schematic pictures of fibrinolysis, describe it
plasminogen
↓
plasmin
(tPA ↑s this rxn, antifibrinolytics like TXA or amicar ↓s this rxn)
↓
fibrin(ogen) → fibrin(ogen) degradation products**
**this is why ppl with DIC have ↑ fibrin degrad. products! everything else is low, but FDP is high bc they’re breaking down clots over and over again!
tissue plasminogen activator
exogenous plasminogen activators such as streptokinase are used to dissolve intravascular clots
-use in thrombotic stroke
thrombolytics
what do they do?
when do they work best?
- Possess inherent fibrinolytic effects or enhances the body’s fibrinolytic system by converting endogenous pro-enzyme plasminogen to the fibrinolytic enzyme plasmin
- More capable of dissolving newly formed clots (platelet rich and weaker fibrinogen bonds)
- Older clots have more cross-linking and are more compacted = more difficult to dissolve
POC coag measures:
PT
PTT
- Prothrombin time (PT) - evaluates the extrinsic pathway. A sample of blood plasma is incubated with tissue factor in the presence of excess Ca2+ . It is particularly sensitive to three of the four Vit-K dependent factors (II, VII and X). Commercial prothrombin reagents vary markedly in their responsiveness to warfarin-induced decreases in clotting factors and are not interchangeable between laboratories, therefore the INR is used for standardization
- Partial thromboplastin time (PTT) – indicates the performance of the intrinsic pathway. A sample of blood is triggered by adding an activator surface ( e.g., silica) plus phospholipid and CA2+
POC coag measures:
ACT
TEG/ROTEM (viscoelastic testing)
- ACT - Performed by mixing whole blood with an activated substance to initiate activation of the clotting cascade. Widely used and is reliable for high heparin concentrations. Influenced by hypothermia, thrombocytopenia, coagulation deficiencies
- Viscoelastic Testing – Thromboelastometry (TEG) & Rotational Thromboelastometry (ROTEM) – a global assay for whole blood clotting including coagulation factors, inhibitors, anticoagulant drugs, platelets, and fibrinolysis
POC coag measures:
bldg time
heparin [] measurements
plt fn tests
- Bleeding time – sensitive test of platelet function. A small standardized incision is made in the underside of the forearm and the amount of time it takes for bleeding to stop is recorded
- old-timey
- Heparin Concentration Measurements – increasing concentrations of protamine are added to samples of heparin-containing blood. Time to clot is measured the sample in which heparin and protamine are most closely matched will clot first. (1mg Protamine will inhibit 1mg (100 units) heparin)
- issue with standardization
- Platelet Function Tests - The classic method involves centrifugation of patient blood to obtain platelet- rich plasma, which is then analyzed in a cuvette at 37°C placed between a light source and photocell.
- bc CBC tells you the number but not if they’re working
Coagulation Test Normals:
bldg time
plt count
PT
bldg time: 3-10 min
plt count: 150-350,000 mm3
PT 12-14 sec
(prothrombin time)
Coagulation Test Normals:
INR
aPTT (activ partial thromboplastin time)
TT (thrombin time)
INR: 0.9-1.2
apTT 25-35 sec
thrombin time 25-35 sec
Coagulation Test Normals:
ACT (activ coagulation time)
Fibrinogen
ACT 80-150 sec
Fibrinogen >150 mg/dL
Remember all of them, but DEF the ACT. But all of them LOL.
List some of the granular contents that plts release, which cause recruitment and activation of additional plts.
(dense granule and alpha granule)
total of 12 granules
- dense granules
- ADP
- ATP
- serotonin
- histamine
- Ca++
- alpha granules
- vWF
- fibrinogen
- F5, F7, F11, F13
- PAI-1
- TFPI
- thrombospondin
- fibronectin
