Haematology and immune mediated disease Flashcards
What is the difference between petechiae and ecchymoses?
Petechiae - <3mm, capillary bleeding
Ecchymoses - larger, small arteriole bleeding
Causes of thrombocytopenia
Increased destruction, consumption, sequestration
Reduced production
Coagulation abnormalities in DIC
Initially hypercoagulable, progresses to consumptive coagulopathy
Causes of increased platelet consumption
DIC
Causes of platelet sequestration
Hepatomegaly, splenomegaly, hypotension, hypothermia, endotoxaemia
Name some inherited thrombopathias and the breed association
Glanzmann’s - Great Pyrenees/Otter Hounds
Thrombopathia of Bassett Hounds and Spitz
Delta storage pool deficiency - American Cocker Spaniel
Causes of acquired thrombopathias
Infectious - rickettsial
Hepatic disease/uraemia
Neoplasia
Drugs - aspirin/clopidogrel, carprofen, hydroxylethyl starch solutions, omega-3 FA (in vitro only)
Likely cause of surface bleeding in patient with normal platelet number and function
Vascular disorder
Vascular disorders
Primary immune-mediated
Secondary to medications, infections, neoplasia
Normal BMBT (dogs and cats)
<4 mins (dogs)
<2 mins (cats)
Clinical signs of primary haemostat disorder
Petechiation, ecchymoses, mucosal bleeding, ocular haemorrhage, haematoma formation
Calculation for manual platelet count
Platelets/hpf x 15-20
What haematology findings are suggestive of increased platelet production?
Increased MPV, PDW, reticulated platelets
What is the general pathogenesis of most immune-mediated haematological diseases (Th-1/Th-2)
Th-2 (antibody-mediated)
What are the mechanisms through which immunosuppressive medication act to reduce immune-mediated disease?
1 - reduced production of antibody by lymphocytes
2 - suppressing clearance of opsonised cells by macrophages or complement
Which antibody class is most associated with IMHA? What is it's half-life?
IgG
7 days
How can platelet function be assessed?
Impedance whole blood aggregometry
Plasma-based light transmission aggregometry
Platelet function analysed (PFA-100)
Which PFA cartridge has the highest sensitivity for canine platelets?
Collagen/ADP
What is assessed by platelet aggregation?
Platelet number
Platelet function
vWF
Which pathway and factors are assessed by PT?
Extrinsic
FVII, X, V, II, I
What does an elevated PT with a normal aPTT indicate?
FVII deficiency
What pathways and factors are assessed by aPTT?
Contact pathway, intrinsic pathway, common pathway
Prekallikrein, FXII, FXI, FVIII, FIX, FX, FV, FII, FI
Which factors do not affect aPTT?
FVII and FXIII
Are PT/aPTT affected by heparin?
PT - no
aPTT - yes
What is a common clinically irrelevant incidental finding on coagulation testing in cats
Factor XII deficiency (prolonged aPTT, normal PT)
What conditions are associated with an elevated aPTT but normal PT?
Haemophilia (A, B, C), heparin, FXII def
What conditions are associated with elevated PT and aPTT?
Rodenticide toxicity, liver disease, DIC, hypo-/dysfibrinogenaemia
What is tested by the ACT?
All factors but VII and XIII
What are FDPs?
What does an elevation suggest?
Created when plasmin lyses fibrinogen, fibrin monomers, insoluble fibrin, cross-linked fibrin
Plasmin activity
What are D-dimers?
What does their elevation indicate?
What is their half life?
Form of FDP generated from cross-linked fibrin
Thrombin generation, fibrin formation, cross linking by FXIIIs and plasmin activity
Short half life (5h) - indicate recent fibrinolysis
How does altered PCV affect TEG analysis?
Higher hct - hypo coagulable tracing
Lower hct - hyper coagulable tracing
What is assessed by a calibrated automated thrombogram (CAT)?
The ability to generate thrombin in real time in-vitro in response to a specific stimulus
Used with platelet-poor or platelet-rich plasma
Virchow’s triad
Vascular stasis
Increased coagulability
Disruption or activation of vascular endothelium
By how much does a clotting factor need to be reduced to elevate PT/aPTT
30-50%
List acquired hyper coagulable states
IMHA Cardiac disease (HCM, endocarditis, dirofilaria) PLE PLN Neoplasia Endocrine dz (HAC, hypoT, DM) Prednisolone Inflammation/Sepsis
List acquired hypo coagulable states
Vit K deficiency - rodenticide
Hepatic failure
DIC
Factor VIII and IX inhibitory antibodies as consequence of multiple transfusions in haemophilia
Vit K dependent coagulation factors?
II, VII, IX, X
Protein C+S
How to anticoagulant rodenticides cause bleeding?
Inhibit vitK epoxide reductase - reduced vita essential for carboxylation of glutamic acid residues on coagulation factors
How soon after rodenticide ingestion is bleeding typically observed?
2-5 days
What can be measured to indicate rodenticide ingestion?
PIVKA - elevates within 12 hours of ingestion
What is the primary mediator of DIC?
Tissue factor
What are the risk factors for DIC development?
Prolonged hypotension
SIRS
Disturbed blood flow to major organ
Major tissue trauma
What has been associated with Increased mortality in dogs with DIC?
High D-dimer, low antithrombin, hypo coagulable TEG trace
Are the following indicated in DIC;
- Heparin
- Plasma
Heparin - no
Plasma - if active bleeding
Labile clotting factors?
V and VIII
Scott syndrome Breed Inheritance Signs Cause Test
GSD
Autosomal recessive
Epistaxis, soft tissue haemorrhage, surgical haemorrhage
Defect in procoagulant activity on platelet surface
Prothrombin consumption assay
Haemophilia - factor deficiency? Signs Test Grading Prognosis
A - VIII B - IX Prolonged bleeding, haematoma formation, mucosal bleeding, haemarthrosis Prolonged aPTT/ACT, normal PT Measurement of factor activity Mild - 6-20% Moderate - 2-5% Severe - <2% Variable
Where does erythropoiesis occur in:
a) The foetus
b) Normal Adult
c) Anaemic adult
a) BM and liver
b) BM
c) BM, liver and spleen (EMH)
How long does it take for red blood cells to mature and be released from the bone marrow?
5-7 days
Describe cat’s reticulocytes
Aggregate reticulocytes develop into punctate reticulocytes, then remain in circulation for 2-3 weeks
How is the corrected reticulocyte count calculated? When is it useful?
CRC = (reticulocyte count x hct)/normal hct
RC normally expressed as % of circulating RBCs, may falsely elevate in anaemia
How long does it take for reticulocytosis to develop after haemorrhage/haemolysis?
2-5 days
Ddx for erythrocyte destruction
Non-immune haemolytic anaemia (intoxication, infection, red cell fragmentation syndrome, hereditary, hypophosphataemia (C))
Haemophagocytic anaemia
IMHA (associative/non-associative)
What is the mechanism of hypophosphataemia causing haemolysis?
Inhibition of ATP production
Leads to decreased membrane stability, increased osmotic fragility, susceptibility to oxidative stress and haemolysis
Name 3 infections associated with haemolysis
Babesia
Haemoplasmosis
Feline cytauxzoonosis
Name 3 red cell fragmentation syndromes
Microangiopathic haemolytic anaemia - HSA, other neoplasia, CHF, glomerulonephritis, myelofibrosis
Haemolytic uraemia syndrome - renal arterial/arteriolar endothelial lesions induce local thrombi
DIC
What are the hereditary haemolytic anaemias? What are their mechanisms?
Pyruvate kinase deficiency and phosphofructokinase deficiency - impaired erythrocyte energy metabolism
Stomatocytosis - genetic membrane disorder
Pyruvate kinase deficiency Species Inheritance Clinical presentation Age at diagnosis
Dogs and cats
Autosomal recessive
Varying degree of regenerative anaemia. Progressive iron overload => haemosiderosis and liver fibrosis.
Dogs (not cats) develop progressive myelofibrosis and sclerosis
Dogs young, cats old
Phosphofructokinase deficiency
Species
Inheritance
Clinical presentation
Dogs only
Autosomal recessive
Some have no clinical signs, most have persistent haemolytic anaemia exacerbated by sporadic haemolytic crises secondary to exercise-induced hyperventilation alkalaemia
Haemophagocytic anaemia
Breed predispositions
Bernese Mountain Dog, Rottweiler, Golden Retriever, Flat Coat
Haemophagocytic anaemia
Pathogenesis
Sub-group of histiocytic sarcoma derived from macrophages. Demonstrate infiltrative growth in liver, spleen, lung and BM
Haemophagocytic anaemia
Clinical signs
Vague
Most have hepatosplenomegaly
Moderate-severe anaemia with abundant reticulocytosis
Haemophagocytic anaemia
Diagnosis
Demonstration of histiocytes containing RBCs or haemosiderin deposits
Immunophenotyping CD11/CD18
Haemophagocytic anaemia
Prognosis
MST 4 weeks
When does feline alloimmune haemolysis occur?
When type A/AB cat receives type B colostrum
What genetic mutation has been associated with polycythaemia vera in humans and dogs?
JAK2
What other haematological abnormality is described in cats with primary polycythaemia?
Thrombocytosis
What are the pathways for erythrocytosis?
Renal hypoxia => hypoxia inducible factor => erythropoietin (cortical fibroblasts)
Thyroid hormone => increased proliferation of erythroid progenitor cells
=> increased hypoxia induced EPO production
Glucocorticoids - synergise with HIF - form burst forming units - erythroid cells capable of self renewal
Growth factor - directly stimulates erythropoiesis
Which 2 endocrine conditions are associated with erythrocytosis?
Hyperthyroidism
Hyperadrenocorticism
What are the ddx for erythrocytosis
Physiological
Primary
Secondary - physiologically appropriate
(R=>L shunt, PDA, TOF, pulmonary disease)
- physiologically inappropriate (renal hypoxia/aberrant EPO production)
What are the clinical signs of erythrocytosis?
Hypervolaemia - ocular - engorged retinal vessels, uveitis, retinal detachment, glaucoma
- engorged mucous membranes
Hyperviscosity - CNS - weakness, ataxia, seizures
Haemorrhage
PUPD
V/D
What ABG findings are consistent with a physiologically appropriate secondary erythrocytosis?
Arterial O2 sat <92%
What medical treatments have been described for primary erythrocytosis?
Hydroxyurea
Chlorambucil (humans only)
Radiophosphorus
What are the possible complications with phlebotomy?
Iron deficiency
Hypoproteinaemia => oedema
What are the three components of primary haemostats?
Platelets
VWF
Vessel wall
Describe platelet development
Megakaryocyte precursor cells form platelet-specific organelles and surface proteins
Develop large pseudopodia, stretch to form pro-platelet processes
Platelet organelles move to end of pro platelet
Branch and constrict - beaded appearance
Proplatelets break away from megakaryocyte and fragment into platelets
What is the lifespan of a platelet in the circulation?
6-10 days
Describe platelet activation
Triggered by injury to vessel
Platelets transform into adhesive spiny spheres - recognise and bind sub endothelial matrix
What are coated platelets?
Subset of collagen and thrombin-activated platelets
Scramble phosphatidylserine from inner membrane to outer - release PS micro-particles - act as scaffold for tens assembly
What substances antagonise platelet receptors and dampen platelet reactivity?
Prostacyclin, PGE2, PGD2
What is the most abundant platelet integrin receptor? What is it’s function
Alpha-IIb-beta-3 complex (GPIIb/IIIa)
Activation-dependent receptor for fibrinogen, fibronetin, vWF
What autoantibody target has been identified in fogs with ITP?
GPIIb/IIIa
Which drugs have been implicated in development of ITP?
Sulphonamides, cephalosporins
What infectious agents have been implicated in 2ndry ITP?
Anaplasma phagocytophilum, Babesia sp, Ehrlichia Canis, Leptospira sp, Leishmania infantum
What conditions have been associated with platelet-bound antibodies in dogs?
Lymphoma, HSA, histolytic sarcoma
Chronic hepatitis, pancreatitis, SIRS
What are the mechanisms by which neoplasia causes thrombocytopenia?
Consumption (bleeding/DIC), sequestration, myelopthisis, platelet bound antibodies
How can a diagnosis of ITP be further supported? What are the limitations of the test?
Flow cytometric assay for platelet-bound antibody
Sensitive but not specific
Increase in stored samples
For which agent is there evidence as a sole agent in ITP?
MMF
What are the recognised types of vWD?
What are their characteristics?
Type I - low VWF concentration, full array of multimers, mild-moderate bleeding
Type II - variable reduction in VWF concentration, absence of high molecular weight multimers, moderate-severe bleeding
Type III - complete absence of VWF - severe bleeding
Which breeds are associated with type I VWD?
BMD, Corgi ,Coton de Tulear, Doberman, Dutch Partridge dog, German Pinscher, Goldendoodle, Irish Setter, Kerry Blue Terrier, Manchester Terrier, Papillon, Poodle, Stabyhound, WHWT
Which breeds are associated with type II VWD?
GSHP, GWHP
Which breeds are associated with type III VWD?
Dutch Kooiker, Scottish Terrier, Shetland Sheepdog
What laboratory assessments of VWF are available?
VWF:Ag (ELISA)
DNA testing
Qualitative assay - WWF:collagen-blinding assay
How is VWF:Ag interpreted?
What factors can alter it?
Normal - 70-180% Boderline - 50-69% Abnormal - 0-49% Decrease - haemolysis Increase - ill - inflammatory/septic, pregnancy
What are the modes of inheritance for VWD
Type I - autosomal dominant or recessive
Type II - autosomal recessive
Type III - autosomal recessive
How are qualitative VWF assays (CBA) interpreted?
Used to diagnose type II VWD Assesses binding to collagen type I/III - dependent on high molecular weight multimers Reported as ratio of VWF:CBA to VWF:Ag 1 = normal 2 = Type II VWD
When should VWD be treated?
Type II and III - bleeding inevitable
Type I - phenotype variable
Aim to control spontaneous/trauma-related bleeding
What are the treatment options for VWD?
Desmopressin - 30 mins before surgery or SID
Blood component - FWB, FFP, CRYO
Cryo ideal - 1 unit/10kg, ever 8-12 hours
What is the half life of plasma VWF?
12 hours
What is the evidence for DDAVP use in dogs with Type I VWD? Effect on VWF:Ag?
Raises plasma VWF concentration from 10 => 17%
Improved haemostat function (PFA-100)
Shortened BMBT
What are the risks of DDAVP treatment of VWF, particularly at high doses?
Water retention and hyponatraemia
Tachyphylaxis
Failure to stimulate VWF release from Weibel-palade bodies with repeat administration
What has been associated with acquired VWD in dogs?
MMVD and sub aortic stenosis (CKCS)
Following tetrastarch bolus
Angiostrongylus (case report)
What type of VWD has been reported in cats?
Type III
List some hereditary platelet dysfunctions
Glanzmann thrombasthenia
Calcium-diacylglycerol guanine exchange factor thombopathia (Basset thrombopathia)
Platelet procoagulant deficiency (Scott syndrome)
What tests are available for the diagnosis of Scott syndrome?
Flow cytometry
DNA testing
What acquired platelet dysfunctions have been recognised in dogs?
Uraemia, hepatobiliary disease, paraproteinaemia
What anti-platelet medications are available and what are their mechanisms?
Aspirin - irreversible cyclooxygenase inhibitor
Clopidogrel - P2Y12 ADP receptor antagonist
What congenital WBC disorders are described in dogs/cats?
Chediak-Higashi Syndrome Pelger-Huet Anomaly LAD Trapped neutrophil syndrome Pyruvate kinase deficiency X-linked SCI Autosomal recessive SCI Cyclic haematopoiesis of grey collies Common variable immunodeficiency Lysosomal storage disease
What secondary WBC disorders are described?
Immune-mediated neutropaenia Primary MDS Secondary MDS Secondary dysmyelopoiesis Myeloproliferative disease Myelopthisis Myelotoxicosis Viral - parvo/distemper/FeLV/FIV/FIP Rickettsial/fungal
What are the causes of immune-mediated neutropaenia?
Primary
Secondary - tick-borne
- drugs (phb, cephalosporins)
How is primary immune-mediated neutropenia diagnosed?
Exclude other causes - overwhelming demand, reduced production (haemic neoplasia), drug-induced, sequestration (splenomegaly)
Flow cytometry (anti-neutrophil ABs) described
Rapid response to immunosuppression
What are the features of MDS?
Non-regen anaemia or multiple cytopaenias
Normal/hypercellular BM
BM - increased immature precursors, increased dysplastic cells, megaloblasts and erythroid precursors
Blast cells <30% nucleated cells
Lack of increased myeloid cells
