Transcript HealthyQuest Presentation

Introduction to
Coagulation Testing
Laura Worfolk, Ph.D.
Scientific Director, Hematology
Quest Diagnostics Nichols Institute, Chantilly, VA
Hemostasis
• Intricate system maintaining blood in fluid state
– Reacts to vascular injury to stop blood loss and
seal vessel wall
• Involves platelets, clotting factors, endothelium,
and inhibitory/control mechanisms
– Highly developed system of checks and balances
Normal Hemostasis
Absence of overt bleeding/thrombosis
Bleeding
2
Thrombosis
Interested Specialties
Anesthesiology
Anticoagulant Management
HIT
Vascular Surgery
Cardiology
Graft Occlusion
PAD
Premature CAD
Primary Care
Practice patterns vary
Bleeding &
Thrombosis
OB/GYN
Hematology
Hemophilia
Thrombophilia
Nephrology
Fetal loss, Infertility,
Menorrhagia
AV Graft Occlusion
Neurology
Stroke
3
Hemostasis Statistics
• #1 cause of death is CVD (includes heart attack &
stroke)*
• ~1-2% of population w/ von Willebrand’s disease†
• ~18,000 Americans w/ hemophilia†
• ~600,000/year w/ venous thromboembolism‡
– ~½ with long-term health consequences;
~60,000 fatalities†
– ~5-8% of population w/ thrombophilia†
4
*WHO.
†CDC
‡www.dvt.org..
Primary Hemostasis
• Platelet role:
– Adhesion (via vWF), post injury to vessel wall
– Activation: shape changed, contents released
– Aggregation, ie, “plug formation”
– Formation of surface for coagulation reactions “fibrin glue”
vWF, von Willebrand factor.
5
Coagulation Cascade
XII
XIIa
XI
Injury
XIa
HMWK/Prekallikrein
IX
VII
IXa
TF
VIII
VIIIa
X
Xa
TF/VIIa
Va
Prothrombin
V
XIII
Thrombin
XIIIa
Fibrinogen
Fibrin
(soluble)
6
Fibrin (insoluble)
Cascade Simplified
Activation/Injury
Intrinsic Pathway
Extrinsic Pathway
(XIIa, XIa, IXa, VIIIa)
(TF, VIIa)
Common Pathway
(Xa, Va, IIa, Fibrinogen)
7
Thrombin Regulation
• Activity and formation tightly controlled
– Antithrombin III
• Inactivation of IIa and other enzymes involved in its
formation
– Protein C and protein S pathway
• Inactivation of cofactors Va and VIIIa
– Tissue factor pathway inhibitor
• Turns off extrinsic pathway (TF, VIIa)
Defects in regulatory mechanisms: thrombosis
8
Fibrinolytic Pathway
Clot lysis vital in prevention of vessel occlusion
uPA, tPA
Plasminogen
PAI-1
Plasmin
Fibrin Clot
Alpha-2 AP
Fibrin(ogen) Degradation Products
Defects: bleeding or thrombosis
9
Hemostasis Balance
Thrombin Generation
(ie, Factors II – XII, cells)
Coagulation
Plasmin Generation
(ie, tPA, uPA, cells)
Healing
Fibrinolysis
Thrombin Regulation
Plasmin Regulation
(ie, PC/PS, AT, TFPI, cells)
(ie, PAI-1, cells)
Cellular contribution: platelets, endothelium, monocytes
10
Alteration of Balance
Laboratory testing indicated if
•
•
•
•
•
•
•
Factor deficiencies
Acquired inhibitors
Anticoagulant therapy
Consumption (DIC)
Dysfibrinogenemia
Platelet defects
von Willebrand’s disease
11
DIC, Disseminated intravascular coagulation
Alteration of Balance
Laboratory testing indicated if
• Inhibitor deficiencies
• Acquired inhibitors
(eg, lupus anticoagulant)
• DIC
• Heparin induced thrombocytopenia
12
Case Study #1
• 21 y/o female with vague family history of
bleeding disorder; evaluated prior to taking
scuba diving lessons
• Has nose bleeds following aspirin ingestion
• Differential diagnosis?
– Role of laboratory testing???
13
Case #1: Lab Testing
• Screening assays
– aPTT: assesses intrinsic & common pathways
– PT: assesses extrinsic & common pathways
– Fibrinogen: hypo- or dysfibrinogenemia?
– CBC: platelet count
• von Willebrand’s disease (vWD) evaluation
– Multiple tests required to classify vWD type
• Antigenic and functional assays
14
Case #1: Test Results
Test
aPTT
Result
(Ref. Range)
33.7 sec
(25.3 – 35.8)
Platelet count
231 K/L
Comment
Detects intrinsic/common
pathway factor deficiency
Rule out thrombocytopenia
(130 – 400)
Factor VIII
activity*
vWF antigen*
85%
Rule out FVIII deficiency
(50 – 150)
40%
Consistent with vWD
(50 – 150)
*Acute phase proteins.
15
Case #1: Test Results
Test
ABO blood type
Result
Comment
Type O:  levels of vWF Ag
AB+
vWF Functional Assays
Test
Ristocetin cofactor
activity
Platelet
aggregation
Result
(Ref Range)
22%
(50 – 150)
Comment
If abnormal activity:antigen
ratio, suspect qualitative (ie,
Type II) defect
No aggregation Indicates abnormal vWF
function
16
Case #1: Multimer Analysis
Shown is representative gel
of normal and type 1 and 2A
vWF deficiencies
Patient results demonstrated
absence of high and
intermediate molecular
weight multimers consistent
with type 2A vWD
17
Case #1: Summary
Probable diagnosis
von Willebrand’s disease type 2A (bleeding
disorder)
18
Case Study #2
• 38 y/o Caucasian man admitted for evaluation
of portal hypertension; history of recurrent
thrombosis (>10 years)
PT, aPTT, fibrinogen: normal
• Positive family history; father and sister with
venous thrombotic episodes, but no laboratory
investigation
• Differential diagnosis??
19
Inherited Thrombophilia
Risk Factors
% in
Healthy
% in
VTE
RR (%) of
Thrombosis
5
21
3–7
0.02–0.17
1
15 – 40
Protein C deficiency
0.3
3
5 – 12
Protein S deficiency
0.7
2
4 - 10
2
6
2–3
5–10
10 – 25
3-4
Condition
APC resistance/FV Leiden
mutation
AT deficiency
Prothrombin (FII) 20210GA
mutation
Hyperhomocysteinemia
VTE, venous thromboembolism; RR, relative risk; APC, activated protein C; AT,
antithrombin.
20
Case #2: Lab Testing
Test
Result (Ref. Range) Comment
Protein C activity 80% (70-180%)
Protein S activity 95% (70-150%)
AT III activity
110% (80-120%)
APC Resistance 1.1 (< 2.0)
Prothrombin
gene mutation
Activity assays detect
qualitative or quantitative
deficiencies
Positive; suggestive of FV
Leiden mutation; genetic
testing for confirmation
Not detected
21
Case #2: Summary
Probable diagnosis
Thrombosis caused by APC
resistance/factor V Leiden mutation
22
Value of Thrombophilia Testing
• Testing does not affect management of acute
events
• Test results may influence decisions
– How long & how intensively to treat
• Prevention of recurrence
– Prophylaxis during high-risk procedures
– Need to evaluate family members
– Estimate future risk (ie, risk associated with HRT)
HRT, hormone replacement therapy.
23
Case Study #3
• 40 y/o woman with iron deficiency anemia due
to menorrhagia; hysterectomy delayed due to
prolonged screening test
aPTT:
PT:
Fibrinogen:
47.8 sec (elevated)
13.0 sec (normal)
300 mg/dL (normal)
• No history of bleeding or bruising; no family
history
• Differential diagnosis??
24
Lupus Anticoagulants
• Antiphospholipid antibodies (APA) are directed
against proteins bound to phospholipid
membrane surfaces
• Lupus anticoagulants (LA) are a type of APA
– Associated with thrombosis & recurrent fetal
demise
– Characterized by prolongation of phospholipid
dependent clotting assays (ie, aPTT)
25
ISTH Criteria for
Lupus Anticoagulants
1. Prolongation of a phospholipid dependent
clotting assay (ie, aPTT)
2. Evidence of inhibition in mixing studies
3. Evidence that inhibition is phospholipid
dependent
4. Lack of specific inhibition by any one
coagulation factor or other circulating inhibitor
(ie, FVIII inhibitors, heparin)
26
Case #3: Lab Testing
Test
Result
Comment
aPTT mixing
studies
No correction
Differentiate factor
deficiency from inhibitor
Lupus Anticoagulant Testing
Test
Result
Comment
dRVVT screen &
confirm
Positive
Consistent with
presence of lupus
anticoagulant
Hexagonal phase Positive
confirm
27
Case #3: Summary
Probable diagnosis:
Lupus anticoagulant
LA may be asymptomatic or associated with thrombotic
events or recurrent abortion. A bleeding history
requires other coagulopathies be excluded. Since LA
may be transient, international consensus guidelines
suggest waiting at least 12 weeks before retesting to
confirm antibody persistence.
J Thromb Haemost. 2006;4:295.
28
Role of Laboratory Testing
• Assist in diagnosis of bleeding and thrombotic
disorders; for example:
– Screen for von Willebrand’s disease in patients
with menorrhagia (ACOG recommendation)
– Test for thrombophilia risk factors in patients with
recurrent spontaneous abortion or thrombotic
events
• Monitor anticoagulant therapy
– Oral anticoagulants, heparin, thrombin inhibitors
29
Role of Laboratory Testing
• Monitor replacement therapy
– Factor levels (ie, FVIII, vWF)
• Pre-op screening
• Risk assessment
30
Pre-analytical Considerations
• Proper specimen handling, processing, and
storage is critical for accurate and precise results
• General specimen requirements available
– www.questdiagnostics.com (click on Test Menu)
– www.nicholsinstitute.com (click on lab information
specimen requirements)
– Quest Diagnostics Nichols Institute Directory of
Services (contact your local representative)
31
Resources
• Laboratories performing routine, specialty, and
esoteric hemostasis testing
• Consultative services available @ Quest
Diagnostics Nichols Institute
Mervyn Sahud, MD
San Juan Capistrano, CA
949-728-4794
Jeffrey Dlott, MD
Chantilly, VA
703-802-6900, x7259
• Quest Diagnostics Interpretive Guide:
http://www.questdiagnostics.com/hcp/intguide/hc
p_ig_main.html
32
Case-Oriented Symposium on
Bleeding & Thrombosis
• October 11-12, 2007, Renaissance Hotel,
Washington DC; topics:
–
–
–
–
–
–
–
Pediatric hemostasis issues
Thrombophilia
Platelet disorders
Thrombotic thrombocytopenia purpura
FVIII Inhibitors
Point of Care testing
New technologies & more
For more information on this CME approved symposium, go to:
http://www.nicholsinstitute.com/Coagulation/Default.htm.
33
References
• ACOG committee opinion. von Willebrand’s disease
in gynecologic practice. Int J Gynaecol Obstet.
2002;76:336.
• Brandt JT, et al. Laboratory identification of lupus
anticoagulants: Results of the Second International
Workshop for Identification of Lupus Anticoagulants.
On behalf of the Subcommittee on Lupus
Anticoagulants/ Antiphospholipid Antibodies of the
ISTH. Thromb Haemost. 1995;74:1597.
• Miyakis et al. International consensus statement on
an update of the classification criteria for definite
APS. J Thrombo Haemost. 2006;4:295.
34
References
• Press et al. Clinical utility of FV Leiden testing for the
diagnosis and management of thromboembolic
disorders. Arch Pathol Lab Med. 2002;126:1304.
• Sadler et al. Update on the pathophysiology &
classification of von Willebrand disease: a report of
the Subcommittee on von Willebrand Factor. J
Thrombo Haemost. 2006;10:2103.
• Thrombophilia: Laboratory support of risk
assessment and diagnosis. Available at:
http://www.questdiagnostics.com/hcp/intguide/jsp/sh
owintguidepage.jsp?fn=CF_Thrombophilia/CF_Thro
mbophilia.htm. Accessed March 21, 2007.
35
Thank you.