Clin Path/Transfusion Medicine/Coagulopathies/hemolymphatic Emergencies Flashcards
Hematopoiesis
Production of all blood cells
Regulated by poietins, colony stimuli, interleukins
Solid cellular portion of blood components
red blood cells (erythrocytes),
platelets (thrombocytes)
five kinds of white blood cells (leukocytes)
– These cells are suspended within plasma, the fluid portion of blood
Erythropoietin
– manufactured in the kidneys
– also facilitates hemoglobin synthesis and stimulates the release of immature RBCs; aka reticulocytes, into the circulation
– increase in production is stimulated by renal hypoxia
– transported to the bone marrow, where it stimulates the proliferation and maturation of erythroid progenitor cell
Erythropoiesis
Cell line up
regulated by?
how long does it take?
Production of RBCs
stem cells → erythroblasts → reticulocytes → erythrocytes
Regulated by EPO which is regulated by blood O2 levels in kidneys
*HYPOXIA stimulates EPO
takes 1 week for RBC maturation
Thrombopoiesis
How long does it take?
what is the precursor cell?
Production of Platelets
Stem cells → megakaryocyte → pieces of cytoplasm become platelets
normal platelet count in small animal patients is 200 000–800 000/mL
Can take 1 week for produciton
Leukopoiesis
types of cells
life span
Production of WBCs
Granulocyte = neutrophils, eosinophils, basophils
Agranulocytes = lymphocytes (T and B), monocytes
– type of cell formed is influenced by cytokines and hormones
– destroyed by lymphatic system
life span ranges from 13-20 days
RBC structure
Life span K9/fel
Contains: H2O, Hb, Bi concave disc shape that creates more membrane for O2/CO2 diffusion
Lacks: Nucleus, mitochondria, ribosomes
K9 Lifespan: 120 days
Fel Lifespan: 68 days
Oxyhemoglobin
when oxygen is bound to hemoglobin
Deoxyhemoglobin
when oxygen is not bound to hemoglobin
CBC Values
vHb - Hemoblobin
HCT
MCV: Mean corpuscular vol; RBC avg size
MCH: Mean corpuscular Hb; Avg wt of Hb in RBC
MCHC: MCH concentration; Avg density of Hb in RBC
RDW: Red Cell Distribution width; EBC size variation
Retic %: Calc # can be overestimated w/ anemia
Absolute Retic count: # of immature RBCs; used to characterize anemia regen vs nonregen
High MCV and low MCHC means:
Regenerative anemia tends to be macrocytic and hypochromic (high MCV, low MCHC)
Normal MCV and Normal MCHC means:
nonregenerative anemias tend to be normocytic and normochromic (normal MCV, normal MCHC).
Low MCV and Low MCHC means:
Iron-deficient anemia (e.g., associated with chronic gastrointestinal hemorrhage) is often microcytic and hypochromic (low MCV, low MCHC).
Dyserythropoiesis
– defective development of RBCs
– non-regenerative anemia, primary bone marrow dz (neoplasia)
– extra marrow suppression from systemic dz
– some toxins (sulfonamides)
– bone marrow typically required for diagnosis
Marrow examination with IMHA
Two major variants are encountered:
1. precursor directed immune-mediated anemia (when reticulocytes are targeted but bone marrow aspirates are still consistent with erythroid hyperplasia)
2. pure red cell aplasia, where earlier precursors are targeted with an absence of these cells on marrow examination.
3
Treatment approach to anemia
therapeutics aim to address impaired DO2, support erythropoiesis, or treat underlying primary disease.
Treatment that Support erythropoiesis
x3
- Darbepoietin: support erythropoiesis in situations of endogenous erythropoietin deficiency
- Iron dextran: support erythropoiesis in iron-deficient patients
- Cobalamin: support erythropoiesis in hypocobalaminemic patients
Senescene
What can exacerbate this?
RBC aging process
1% of old RBCs removed daily via intravascularly or extravasclularly
Oxidative Stress (free radicals) contribute to rapid RBCs aging/destruction
-exacerbated by dz/toxins
Intravascular vs Extravascular Hemolysis
where does it occur? what % is destroyed where?
Intravascular: accounts for 10%, Hb released into blood stream → hemglobinemia → excess unconj Hb in plasma → eliminated by kidneys
Extravascular: 90% of RBC destruction via macrophages in spleen and liver into amino acids, iron, and heme (from Hb)
-AA recycled by Liver
-Iron to bone marrow
-Heme broken into free or unconj bilirubin
What clin path finding is present in intravascular hemolysis but not extravascular?
Hemoglobinemia and hemoglobinuria are not seen in extravascular hemolysis since the cells are destroyed outside the blood vessels via normal mechanisms
Pathophysiology of Icterus
3 types
-Excessive RBC breakdown → excessive unconj bilirubin in plasma
-Unconj bilirubin exceeds Livers ability to conjugate → desposits to tissues (PRE-hepatic)
-Liver dz/dysfunction cannot handle processing unconj. bilirubin → buildup
- Bilirubin obstruction → conj bilirubin backs up into blood stream then tissues (POST- hepatic)
Regenerative Anemia
What do the RBCs typically look like?
– bone marrow responds appropriately by increasing red blood cell production and increased reticulocytes release
– occurs either due to hemorrhage or hemolysis → result of internal loss, external loss, destruction by way of hemolysis, or dilution
– polychromasia → more immature RBCs than mature
– can be macrocytic → larger than normal OR anisocytosis → various different sizes
– peripheral metarubricytosis (nucleated red cells)
– hypochromic = low hemoglobin concentration and hence appear a paler color due to the abundance of reticulocytes in circulation
Two forms of reticulocytes observed in cats:
- aggregate → shorter lived and should be used to gauge regeneration in cats
- punctate
Metarubricytes in the face of nonregenerative anemia is:
an inappropriate marrow response, and reasons for bone marrow derangement should be considered (e.g., bone marrow neoplasia, heat related illness, feline leukemia).
How long does it take for bone marrow to respond to acute blood loss?
– bone marrow requires time to respond to acute blood loss, and this typically takes 2–3 days in cats and 4–5 days in dogs
– anemia can be “preregenerative”
5
Macrocytic, Normocytic Anemia
- FeLV infections with no reticulocytosis (common)
- Poodle macrocytosis (not anemic)
- Hyperthyroid cats (slight macrocytosis without anemia)
- Spurious with erythrocyte agglutination
- Spurious in cats and dogs with persistent hypernatremia
Regenerative Anemia Absolute retic count #
Absolute reticulocyte/PCM count:
> = 100,000/ul (dog)
> = 60,000/ul (cat)
non regen < than these #s
Non-regen, Normocytic, Normochromic anemia
the type of anemia in which circulating RBCs are the same size (normocytic) and have a normal red color (normochromic)
– Pre-Regenerative anemia → not enough time has passed for reticulocyte generation to respond
Non‐Regenerative Anemia
2 categories
Describe RBC appearance
– occurs secondary to impaired erythropoiesis associated with primary bone marrow disease or systemic disease leading to secondary extra-marrow suppression.
– Most cases of non‐regenerative anemia have normal red blood indices as they are normocytic and normochromatic
– two categories:
1. hypoproliferative anemia
2. abnormal RBCs are sequestered in the bone marrow
– red blood cells appear microcytic after Hb, iron, and RBC reserves are exhausted
Hypoproliferative anemia
– bone marrow RBC production is static or reduced
What is the best way to evaluate regeneration response?
the presence or absence of reticulocytes in the peripheral blood
7
Potential causes for NNN anemia
- Chronic inflammation and neoplasia (sometimes slightly microcytic)
- Chronic renal disease
- Endocrine deficiencies
- Selective erythroid aplasia
- Aplastic and hypoplastic bone marrow
- Lead toxicity (may not be anemic)
- Cobalamin deficiency
Non Regen Anemia: Primary Bone
Marrow Disorder
- Pure Red Cell Aplasia (PRCA)
- Aplastic Anemia
- Myelodysplastic
- Syndrome
- Myeloproliferative
- Diseases
- Myelophthisis
- Myelonecrosis
- Viral Infection
* Parvovirus
* (Dog/Cat)
* FeLV (Feline)
* FIV (Feline) - Drug-Induced
- Hematologic Dyscrasia
4
Non Regen Anemia: Secondary Suppression
of Bone Marrow From Systemic Disease
1 Chronic Inflammatory Disease
*Infection
*Inflammation
*Neoplasia
2. Chronic Renal Failure
3. Endocrine Disorder
*Hypothyroidism
*Hypoadrenocorticism
*Estrogen toxicity
primary failure of erythropoiesis
severe erythroid hypoplasia of the bone marrow occurs and precursor red cells are not produced
– uncommon, but primary failure is typically immune mediated, acquired, and may be associated with many disease conditions
Aplastic anemia: Non regen or Regenerative
Non-regenerative: characterized by bone marrow failure
Examples:
Estrogen induced
Drug use : antibiotics
Chemotherapy
Radiation
Infectious disease
Can be primary or secondary to another dz process: thymoma, leukemia, infection, toxicities, or renal disease
Marrow infiltration
another form of bone marrow failure associated with
1. neoplasia
1. myelofibrosis: the replacement of bone marrow with fibrous connective tissue
1. osteopetrosis
Hemolysis: Non-regen or Regenerative?
Regenerative
Immune mediated
Neonatal isoerythrolysis
Transfusion reaction
Infectious
Heinz body anemia
Zinc toxicity
Copper toxicity
Pyruvate kinase deficiency
Phosphofructokinase deficiency
Impaired red cell production: Non-regen or Regenerative
Non-Regenerative
Immune mediated → can be both
Neoplasia → can be both
Drug induced
Inflammatory disease→ Always Non Regen
Chronic renal disease
Endocrine diseases
Bone marrow infiltration: Non-regen vs Regenerative
Non‐regenerative
Neoplasia
Myelofibros
anemia of chronic inflammatory disease
Type of anemia
suspected mediator
– usually mild to moderate, normocytic, normochromic, and always non‐regenerative
– decreased RBC lifespan
– suspected mediator is hepcidin, a protein produced by the liver in response to the release of IL‐6 as part of host defense against inflammatory stimuli
– Hepcidin is also produced in hypoxic or iron‐deficient states and inhibits iron export from duodenal enterocytes and macrophages, resulting in decreased iron absorption
Polycythemia
Definition and consequences
abnormally increased packed cell volume, red blood cell count, and hemoglobin concentration
– induces an increase in blood viscosity, particularly when the packed cell volume rises above 50–60% in dogs
– polycythemia increases cardiac workload and hinders microcirculation.
– potentiates thrombosis, causes tissue hypoxia, and makes neurological complications more likely
Absolute vs Relative
Absolute polycythemia
primary vs secondary
caused by an increased total RBC mass, divided into primary or secondary polycythemia
– primary when the increase in red cell mass results from an abnormality of the myeloid stem cells
– secondary when the increase in red cells is in response to increased levels of erythropoietin inappropriately
* ex: PDA, congenital heart defects, renal neoplasia, → local hypoxemia triggers erythropoietin
Relative polycythemia
hemoconcentration due to decreased plasma volume
because of the shifting of body fluids from the vascular space into the interstitial space due to dehydration, excessive external loss of body fluids, or overuse of diuretics
– ex: hemorrhagic gastroenteritis, who lose a large volume of fluids rapidly
– Splenic contraction can lead to an increase in circulating red blood cells but not in the total number of red cells in dogs; this normal physiological response should not be interpreted as polycythemia
Acanthocytes
spiculated red cells with a few projections
RBCs with blunt finger like projections
Injury via RBC damage traveling thru vasculature or fragile from iron deficiency
Liver dz, DIC, vasculitis
3 examples
what do these suggest?
Spherocyte; more disc shape than biconcave, smaller, more dense lacking central pallor
IMHA; regenerative
Oxidative injury
Coral snake
– suggest immune mediate destruction
Causes x4
Schitocytes; RBC fragments
mechanical injury to erythrocytes, turbulent blood flow
-Vascular abnormalities; DIC, vasculitis, PSS, hemangioma
Causes x4
Echinocytes “Spur cells” (tiny sun)
Drugs; chemo agents, furosemide
Renal dz; glomerulonephritis
Snake envenomation
Reticulocytes; immature RBCs. Lg amounts of RNA form dark blue clumps/strands
Bone marrow respone to anemia
Ghost Cells; pale RBCs due to loss of hemoglobin thru intravascular hemolysis
-IMHA
-Oxidant injury
-Snake envenomation
Basophilic stippling; numerous small blue punctuate stippling in RBCs
-Regenerative Anemia
-Lead poisoning
caused by;
examples x6
Heinz bodies; dense granuel on the edge of RBC caused by oxidative injury = hemoglobin to precipitate
Propofol; acetaminophen in cats
toxins; Onion, zinc in dogs
anticoagulant rodenticide
Skunk musk
Howell- Jolly bodies; small round dark purple dots (nucleus remnants) within RBCs
Regenerative anemia
impaired splenic function (corticosteroids/splenectomy)
Nucleated RBC - most mature nucleated RBC
present in strongly regenerative anemias
Band Neutrophil; immature neutrophils
↑ bands present = left shift with acute or severe inflammation
Toxic neutophilic change
Neutrophils
how much of count do they make up?
how do they migrate to tissues?
– phagocytes aka “clean up crew” that engulf micor-organisms and tissue debris
–Circulates for few hours then migrates into tissues
– most numerous circulating leukocyte in the dog and cat
– become attracted to site of potential infection or inflammation through a chemical signaling process known as chemotaxis that employs endogenous inflammatory mediators.
Eosinophils
what % of count do they make up?
how long do they stay active?
Respond to histamines → allergies, anaphylaxis, parasitic infx, sometimes cancer
– ≤5% of the total WBC count
– do not stay in the circulating pool for very long and migrate into tissues within hours
– granules contain anti-inflammatory subtances
– effective against protozoas an parasitic infections
Basophils
what do they initiate? what do they release and in reponse to what?
Initiate immune/allergic responses
Release histamines in response to chemicals from mast cells
– rarest WBCs in circulation
– granules contain histamine and heparin, making these cells effective at initiating inflammation by working in conjunction with eosinophils
– Eosinophils are attracted to the site of allergic reaction by chemotactic factors released from basophilic granule
– Basophilia is most commonly associated with an allergic or hypersensitivity reaction
Monocytes
how do they enter tissue site?
make up % of circulating WBCs
- Mono-nuclear
- Phagocytosis, process antigens
- become macrophages → most of their phagocytosis occurs in the tissues
- monocytes enter tissues by the process of chemotaxis.
– after entering tissues = tissue macrophages
– make up 5–6% of the circulating WBCs
– mature much faster than neutrophils,
– stay in peripheral blood longer
– largest WBC in circulation
Tissue macrophages
Where are they mostly found? #4
most prevalent in filter organs such as the liver, spleen, lungs, and lymph nodes
Mononuclear phagocyte system (MPS)
Monocytes and tissue macrophages
responsible for the clean‐up of variety of microscopic debris in the body:
* cellular debris left over from inflammation and infection,
* specific antigens that have been destroyed by lymphocytes,
* other foreign substances.
Lymphocytes
2 types
only WBC type w/o phagocytic capability
Memory cells of immune system
– B cells = divide into plasma cells to create antibodies, Humoral immunity (lymphoid tissues)
– T cells= Cytokine production, cell-mediated immunity (killing/activation/regulation) lymphoid and other tissues
T cells
what % do they make up?
how long do they survive?
Role
– predominant circulating lymphocyte, accounting for up to 80% of peripheral blood lymphocytes
– very long‐lived, surviving on average from 6 months to 10 years.
– there are two subtypes:
* killer T cells and helper T cells
B cells
- After B lymphocytes recognizes an antigen, → transforms into a plasma cell that can release antibodies in a process called humoral immunity.
- Plasma cells derived from B lymphocytes produce, store, and release antibodies that are known as immunoglobulins
- Plasma cells are found in all tissue in the body, but are most common in the lymph nodes and spleen.
Leukocytosis / Neutrophilia mechanisms
#4
- ↑ production due to inflammation (appropriate)
- ↑ production due to neoplasia (inappropriate)
- Demargination
- ↓ egress from circulation
Clin path findings with Increase in production of leukocytes/neutrophils due to inflammation
–Degenerative Left shift: absolute #s of band/immature neutrophils are > than mature ones
–Leukemoid response: strong inflammatory stimulus that can mimic neoplastic response (can be seen with infx)
Demargination
Corticosteriod response = stress leukogram from endongenous or exogenous steriods
–15 - 20K cells/uL
–Steriods usually clear w/i 24hrs but may take longer with prolonged use
– Epinephrine response: excitement (puppies/kittens)
Decreased neutrophil transit time into tissues
caused by
Corticosteriods ↓ neutrophil transit time
Left Shift
↑ immature neutrophils (bands)
– sign of inflammation or infection as the body increases neutrophil release in response to injury
Immune system responsible for causing WBCs to be released early
↑ # of monocytes
– degree of left shift corresponds with the severity of the inflammation or infection
Degenerative vs Regenerative
Regenerative Left Shift
mix of adult and immature cells
– neutrophilic leukocytosis with an increase in immature neutrophils but a larger population of mature neutrophils
Degenerative left shifts
Condition where a neutrophilic leukocytosis is present but there are more immature neutrophils than segmented neutrophils in the blood
– associated with severe disease and typically indicate a poor prognosis
Toxic neutrophils
what is it in response to?
produces in response to bacterial infx tissue necrosis
Lg # = poorprognostic indicator
– very granular appearance
Affect of Addisons on WBCs
Will not see stress leukogram
–body unable to respond phisologically to stress
Leukopenia
– abnormally low white blood cell count, usually due to a decreased neutrophil count known as neutropenia (most abundant WBC)
Leukopenia causes
x7
Decrease production in bone marrow
–Viral: Parvo/Panleuk/FIV/FeLV
–Neoplasia, radiation
–Necrosis
–Drug Toxin; chemo/chloramphenicol/TMZ
–Increased utilization; fulminant inflammation, bacterial sepsis, endotoxemia
-Destruction; immune mediated
–2nd to lymphopenia
Leukocytosis
elevated white blood cell count and is common in a variety of inflammatory states
– historical or clinical evidence of inflammatory disease such as pyrexia, weight loss, loss of appetite, infected wounds, and specific organ system involvement
Infectious Agents/Parasites that cause Leukopenia
#7
Histoplama
Hepatozoon
Ehrlichia
Anaplasma
Babesia
Mycoplasma
Distemper
Thrombocytopenia
4 mechanisms
N = > 200,000 (8-15 phpf)
4 mechanisms of thrombocytopenia:
1. Sequestration; rare in animals
2. Consumption; DIC, Parvo, heatstroke, MODS
3. Destruction; ITP
4. Hypoproliferation; Bone marrow dysfunction/Liver dz = decrease thrombopoeitin production
*CKCS have abnormally low platelets due to them being oversized - idiopathetic asymptomatic thrombocytopenia
Immune‐Mediated Thrombocytopenia results from:
x4
Can be due to:
1. decreased production, → myelosuppression (chemo/drugs)
1. increased consumption (e.g. a large blood clot), DIC
1. increased sequestration, → splenomegaly
1. increased destruction = ITP/DIC
ITP definition
increased platelet destruction results due to an inappropriate immune response
– either primary or secondary
– Regardless of cause ITP is a type II hypersensitivity reaction that results in destruction of circulating platelets when antiplatelet antibodies attach to the surface of the cells and are removed by the monocyte macrophage phagocytic system in the spleen
< 30,000/mL
Secondary ITP
or acquired ITP can occur due to infectious, inflammatory, neoplastic, or toxic processes
Primary ITP
diagnosed based on exclusion of other causes; in cases of acquired ITP, the underlying cause must be determined in order for treatment to be effective.
Blood smear platelet count
Each 100× field should contain 8–15 platelets, with each platelet visualized representing approximately 15 000/μL
Why is vincristine used for ITP treatment?
– vinca alkaloid and immunomodulating antitubular agent
– works by inducing release of platelets from megakaryocytes through premature fragmentation and by impairing the consumption of platelets by macrophages
Veterinary blood products and components that contain platelets
fresh whole blood,
fresh platelet concentrate,
cryopreserved platelets,
lyophilized platelets, and
platelet‐rich plasma
Effusions
Pure Transudate: < 2.5g/dl
–PLE, PLN, portal hypertension, PSS
Modified transudate: 2.5-5.0 g/dl
–blood, neoplasia, CHF, Liver dz
Exudate: > 5 g/dl
–purulent, pyothorax
Lactate forms found in the body
where is it metabolized?
2 types
D-Lactate = non-physiological (short bowel syndrome)
L-Lactate = main form of lactate in body
–produced in skeletal muscle, brain tissue, adipose tissue and circulating blood cells
–Metabolized in Liver (60%) and Kidney (30%)
– <3 uses lactate for ATP production
Normal energy Metabolism
Glucose → glycolysis → pyruvate + 2 moles of ATP per mole of glucose
→ pyruvate enters mitochondria → 36 moles of ATP/glucose mole
Energy metabolism under Anaerobic conditions
Abnormal anaerobic conditions → inadequate O2 supple
–glycolysis rate too high for mitochondria metabolism to keep up → accumulation of pyruvate and NADH = becomes main source of energy production for ATP
Glycolysis role in producing lactate
Glycolysis → glucose = pyruvate → lactate → NADH
–Lactate dehydrogenase activity ↑ = ↑ lactate to pyruvate ratio
Measured Lactate
– Indirect measure of tissue O2 balance
Liver cannot eliminate amount of lactate produced by hypoxic tissue = level of lactate we measure
Cori Cycle
Reverses lactate metabolism when aerobic metabolism restored
Type A Lactate examples
Oxygen Related
↑ O2 demand:
–exercise, shivering, tremors, sz
↓ O2 availability:
– shock, LV failure, CPA, Pulmonary dz
↓ Arterial O2 content:
Acute Anemia, hypoxemia, CO poisoning
Type B Lactate
Not directly O2 related → metabolic
B1= Systemic dz → inflammatory process affecting cells ability to proccess O2
B2 = Drugs or Toxins → medication related to metabolic hyperlactemia, acetaminophen, corticosteriods, Catecholamines
B3 = Hereditary dz → congenital hyperlacemia (GSP), enzyme deficiencies, pyruvate dehydrogenase
FFP
– plasma that has been separated from red blood cells and frozen within 8 hours
– contains all of the coagulation factors, anticoagulants, alpha macroglobulins, albumin, fibrinogen, and fibronectin
Frozen Plasma
fresh plasma is frozen for more than a year (frozen plasma) or kept at room temperature for more than 8 hours, it loses its labile clotting factors (factors V and VIII).
– Frozen plasma contains clotting factors II, VII, IX, X and plasma proteins
Cryoprecip
created by slowly thawing and centrifuging FFP
– centrifuged supernatant is discarded, and the remaining solution (CRYO) contains vWF, factor VIII, fibrinogen, and fibronectin
Cryopoor Plasma (CPP)
CPP is the supernatant removed after FFP has been centrifuged
– contains albumin, globulin, antithrombin, protein C, protein S, and factors II, VII, IX, and X
– albumin concentration and COP were highest in CPP compared with CRYO or FFP
– CPP is a reasonable alternative to FFP for albumin replacement and oncotic support
Albumin
6 important functions
Albumin makes up 70%–80% of COP within the body
– plays an important role in wound healing; as an antioxidant, free-radical scavenger, and transport agent; and in preserving normal platelet function
Causes of hypoAlb
protein-losing nephropathy, protein-losing enteropathy, end-stage liver failure, malnutrition, and systemic inflammatory states
Alb deficit equation
albumin deficit (g) = 10 × 0.3 × BW (kg) × (desired albumin – patient albumin
HSA administration dose
2ml/kg 25% HSA is administered IV over 2 hours followed by 0.1 to 0.2 ml/kg/hr for 10 hours, for a total dose of up to 2 g/kg
Marked leukocytosis and polycythemia with hypoglycemia
because of increased cell utilization of glucose
Pathologic hemolysis
– occurs when the rate of destruction is increased, and the life span of RBCs is shortened
Hemolytic anemia (HA)
results when regeneration of RBCs from precursor cells is inadequate to replenish the destroyed cells
– caused by several immunologically and nonimmunologically mediated mechanisms
9
Causes of Hemolysis
- Fragmentation hemolysis
- Toxicant-induced hemolysis
- Foodstuffs and additives
- Drugs
- Chemicals
- Immune-mediated hemolysis
- Heritable hemolysis
- Infection-related hemolysis
- Miscellaneous
Fragmentation hemolysis
How does it occur?
What type of cell is typically seen?
– mechanical process, most commonly the result of shearing of the RBC membrane in the small vessels (microangiopathic hemolysis) or from altered rheologic forces
– Because shearing occurs inside the vascular space, hemoglobinemia and hemoglobinuria commonly result
– observation of schistocytosis on a peripheral blood smear provides supportive evidence of fragmentation; keratocytes and acanthocytes also are seen
– Assays of coagulation to check for DIC
Toxicant-induced hemolysis
– often related to oxidative injury
– Oxidation of hemoglobin iron results in the formation of methemoglobin, which, although unable to bind oxygen, does not shorten RBC life span
– Cats are more susceptible to chemical oxidant injury than dogs
What causes Heinz-bodies?
Oxidation of hemoglobin causes Heinz body formation
– ultimately resulting in RBC removal
Propofol use in cats
Substances that can cause toxicant-induced hemolysis
5 examples
- Onion/Garlic
- Propylene glycol → propofol
- Zinc/Copper
- Skunk Musk
- Acetaminophen
- Benzocaine
Heritable hemolysis
– relatively uncommon
– all hereditary erythrocyte defects lead to HA
– Osmotic fragility, Phosphofructokinase deficiency, Pyruvate kinase deficiency
Infection-related hemolysis
result of direct cell damage, through initiation of an immunologically mediated response to infection, or both
– Systemic infections also can lead to microangiopathic hemolysis
Infection of red blood cells causes in US:
- hemotropic Mycoplasma spp.
- protozoan parasites → Cytauxzoon felis and Babesia spp
Mycoplasma haemofelis
causative agent of feline infectious anemia
– Presumably transmitted by fleas, the resulting hemolysis is variable in severity, cyclic in nature, and often Coombs positive
Babesia most likely to cause HA in the United States
*B. gibsoni *
any breed can be infected, but infection in North America is most common in pit bull terriers.
Cytauxzoon felis
transmitted by
CS seen
causes a tick-transmitted infection endemic to the southeastern, midwestern, and mid-Atlantic regions of the United States
– Cats typically become sick in the warm weather months with an acute febrile illness often accompanied by icterus and pancytopenia.
Systemic infections that may cause HA
– hemolysis may be due to hemotropic mycoplasmosis
– Feline leukemia
– feline immunodeficiency virus
Immune-mediated hemolysis
secondary vs primary
– destruction of RBCs is mediated by antibody or complement-triggered events
– i.e., secondary IMHA
→ infection, drugs, cancer, procedure
– occurs more commonly in dogs than cats (rare) → cocker spaniels overrepresented
– primary IMHA is idiopathetic
7
Drugs that might trigger Secondary IMHA
- acetaminophen,
- cephalosporins,
- NSAIDs,
- penicillins,
- phenylbutazone,
- tetracyclines,
- trimethoprim‐sulfa
What is the most common form of IMHA?
immunoglobulin-mediated type II hypersensitivity reaction causing extravascular hemolysis
3
IMHA autoantibodies invovled
IgG, IgM, and IgA
– produced against the animal’s own red blood cell membrane antigens regardless of the age or health of the cells
– in normal scenarios, these autoantibodies regulated by suppresor T cells
– animals with IMHA have poor T cell
Microangiopathic hemolytic anemia
what do they result in?
condition in which RBCs are physically damaged during circulation due to vessel occlusion, abnormal vascular morphology, or fibrin shearing.
– Ex: splenic torsion, HWD, DIC, IVCs, hemangiosarcoma
result in Schitocytes
5
Complete blood count findings in IMHA
- regenerative anemia,
- leukocytosis with left shift,
- lymphocytosis,
- thrombocytopenia,
- elevated reticulocyte percentage
Agglutination
– can be seen grossly and microscopically but must be differentiated from rouleaux formation
How to differentiate between Agglutination and Rouleaux?
Rouleaux disperses with saline washing
True agglutination does not
Treatment for IMHA: Immune suppression
Glucocorticoids mainstay of treatment for IMHA
– positive response may still take 3-5 days
– adjunctive immunosuppressive drugs include azathioprine and the less myelotoxic/hepatotoxic but similarly acting agent mycophenolate mofetil
– cyclosporine
Treatment for IMHA: supportive care
provision of adequate oxygen-carrying capacity via transfusion of RBCs
– need for transfusion based on clinical signs
– however reaching a certain level of anemia despite CS can still warrant transfusion due to likely hypoxia (< 12%)
Anti-thrombotics for IMHA
– dogs with IMHA commonly euthanized due to complications from IMHA such as PTE or DIC
– anticoagulants = rivaroxiban, UF heparin
– antiplatelet therapy = clopidogrel, aspirin
Other possible complications with IMHA
x3
– Gastric ulceration and erosion
– both as a result of poor oxygenation of GI tissues and due to the use of high-dose glucocorticoids
– use of proton pump inhibitors as gastric protectants is only recommended routinely when ulcers are suspected
Other causes of immune-mediated hemolytic anemia
transfusion reactions and
neonatal isoerythrolysis result from immunologically mediated but normal attack against RBCs.
– in dogs subsequent transfusions may result in hemolysis
Feline neonatal isoerythrolysis
Type A or type AB kittens born to type B queens can develop hemolytic anemia after absorption of alloantibodies via colostrum.
Hemophagocytic histiocytic sarcoma syndromes
Clin path findings
– severe regen anemia with thrombocytopenia
– result in a rapidly progressive severe extravascular HA that can be confused with IMHA
– Despite HA, the spherocytosis or autoagglutination typical of IMHA is absent and affected dogs are Coombs negative
– hyperbilirubinemia with hypoalbuminemia and hypocholesterolemia are commonly identified.
Platelet aggregation
what does it need to occur?
In the presence of thrombin, platelets change shape and develop pseudopods that allow them to intertwine with each other.
– platelets squeeze together to form a primary hemostatic plug
Thrombin Definition
an enzyme in blood plasma which causes the clotting of blood by converting fibrinogen to fibrin
– essential product of secondary hemostasis
– end‐result is a stable fibrin clot
Thrombin actions
– stimulates platelets and endothelial cells to enhance clot formation
– affects the natural inhibitors of coagulation, causing consumption of antithrombin (AT) and protein C
Fibrinogen
what does it become and how?
a soluble plasma protein present in blood
– from which fibrin is produced by the action of the enzyme thrombin.
Fibrin
an insoluble protein formed from fibrinogen during the clotting of blood.
– It forms a fibrous mesh that impedes the flow of blood by keeping platelets in place
Primary hemostasis definition
activated platelets form a platelet plug in minutes
Secondary hemostasis definition
what initiates it?
reinforcement of the frail platelet plug with fibrin strands occurs within hours
– Factor VII and tissue factor (factor III) are considered the main initiator of secondary hemostasis.
Fibrinolysis definition
the enzymatic breakdown of the fibrin in blood clots
Primary hemostasis pathway
where does it take place?
how is a clot formed?
what actions do platelets have?
Takes place between the injured vessel wall and platelets.
→ results in the formation of a platelet plug at the site of a vascular injury; activated platelets, along with endothelial cells = form an unstable fibrin clot.
→ This platelet plug fulfills multiple purposes:
* provides a physical barrier to inhibit loss of blood
* provides membrane surfaces as binding sites for procoagulant enzymes that aid the formation of thrombin in secondary hemostasis.
→ In primary hemostasis, following platelet adherence, platelets undergo conformational changes and release substances that stimulate platelet aggregation.
→ Aggregated platelets constitute the primary hemostatic plug.
3 phases: 1 Initiation, 2 Extension, Stabilization
Main mechanism for primary hemostasis
platelets
Primary Hemostasis
1 Initiation
binding of platelets to collagen and vWF triggers platelet activation
Primary Hemostasis
2 Extension
Extension of the adhered platelet monolayer on injured vessels undergoes further activation resulting in secretion, shape change, formation of the procoagulant membrane, and integrin activation.
Primary Hemostasis
Stabilization
forces generated by contraction of actin/myosin filaments in platelet further strengthen platelet-to-platelet interactions by narrowing the gap between platelets and preventing the diffusion of activators away from the platelets, hence fostering a local procoagulant microenvironment
Secondary hemostasis pathway
→ formation of fibrin in and around the primary plug, stabilization of the platelet clot through the generation of thrombin (factor II), and activation of the fibrinolytic system.
→ known as the coagulation cascade.
two pathways for the activation of the coagulation cascade: intrinsic and extrinsic.
main initiator of secondary hemostasis
Factor VII and tissue factor (TF; Factor III)
– Factor VII synthesized in liver
Intrinsic pathway
What does it work on?
What factors does it involve?
surface activated, operating strictly with components present in the blood
Factors XII, XI, IX, VIII
12-11 = 1, 9-8 = 1
Extrinsic pathway
What does it work on?
What factors does it involve?
requires tissue factor for activation
Factors: III (TF), VII
Where are clotting factors produced?
All coagulation factors are produced in the liver, with the exception of factor VIII
What is factor IV?
Calcium
– required for most reactions and is the reason why calcium chelators such as citrate and ethylenediaminetetra‐acetic acid (EDTA) are used for blood collection
– Binding agent for Vit K dependent factors
Which factors require Vit K?
Factors: II, VII, IX, X
2+7 = 9 then 10
What initiates the coag cascade?
exposure of tissue factor (TF) which is also known as factor III
Coag Cascade pathway
– TF is exposed from vascular endothelial cell damaged but also contributed by activated monocytes and inflammatory mediators.
– After being exposed, TF binds to factor VII → forms a thrombin complex that initiates the extrinsic cascade.
→ small amount of thrombin produced activates clotting factor XI to factor XIa,
→ which activates factors V, VIII, and XIII.
– Through processes, thrombin formation is maintained.
What does each value mean?
- Reaction time (R): The time to onset of fibrin formation (or when the curve deviates by more than 2 mm from midline).
- K: The time it takes for the curve to deviate 20 mm from the centerline (rate of fibrin formation).
- α: The angle between the time point K and the centerline (rate of fibrin formation).
- Maximum amplitude (MA): Strength of the clot.
- hypocoagulable (A), prolonged CT/R and CFT/K and decreased α angle and MCF/MA.
- normal (B), four standard parameters are within the reference intervals defined at our center.
-
hypercoagulable (C) shortened CT/R and CFT/K, and increased α angle and MCF/MA.
profiles obtained by the viscoelastic tests.
hyperfibrinolytic profiles
1. (A) Note that the beginning of the tracing is also in favor of hypocoagulability. Fibrinolysis occurs 18 minutes after the initiation of the test.
2. (B) Fibrinolysis occurs 27 minutes after the initiation of the test
3. (C) cat with aortic thromboembolism 10 minutes after the administration of intravenous alteplase. Fibrinolysis occurs 5 minutes after the initiation of the test with lysis indices equal to 0% and the maximum lysis equal to 100%
