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bioanalytical pathology II


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platelet production is stimualted by ________
__________ acts as a bridge bt platelets and subendothelial collagen resultsing in ______ of platelets to collagen
von Willebrand factor
3 steps of the platelet plug
2.granule release reaction
notable granular contnets of platelets
1.calcium:which helps stimulate platelet actions and plays a role in the coagulation system
2.von Willebrand factor: which promotes platelet adhesion to subendothelial collagen
3.fibrinogen: which causes platelets to adhere to each other
4.ADP:which induces expression of platelet surface receptors that bind to fibrinogen
When due platelets release there granules?
when they adhere to subendothelial collagen
platelet aggregation
under the influence of released platelet granule contents, platelets begin to adhere to each other.
Ultimately this results in the formation of an aggregate of platelets that plugs the defect in the vessel wall, and, therefore hemorrhage stops
When can primary hemostasis alone provide permaent hemostasis in an injured vessel wall
in the defect is small and the blood flow in the vessel is slow
intrinsic factors
extrinsic factors
common factors
factor III
Factor III is tissue thromboplastin. It does not circulate, but ratheris attached to fibroblasts and smooth muscle cells where it helps promote the activity of the extrinsic system by activating factor VII
factor IV
factor VI
there is no factor VI
where are coagulation factors synthesized
most are made in the liver.
some are made in macrophages.
factor III is made in fibroblasts and smooth muscle cells
coagulation factors that are dependent on vitamin K
factor V
accelerator that promotes more rapid activation of factor II
factor VIII
accelerator that promotes more rapid activation of factor X
The _________ system is thought to be the primary system responsible for activation of coagulation
factor XIII
fibrin stabilizing factor
severe __________ can result in loss of large amounts of _____ in the urine with susequent thrombosis
2 most important anticoagulant proteins
1.Antithrombin III(AT-3)70%
2.alpha 2 macroglobulin 20%
anti thrombin III
heparin at the endothelial cell surface complexes with AT-3, and this complex inactivates thrombin. AT-3 also inactivates factors IX,X,XI,XII
Factor II
prothrombin (thrombin when activated)
alpha 2 macroglobulin
inhibits thrombin, plasmin, and kallikrein
fibrinolysis is performed by ________ and produces ____________
small peptides and d-dimers (which are refered to as fibrin/fibrinogen degradation products-FDPs
increased bleeding time can occur with the what abnormalities
1.blood vessel abnormalities (esp. abnormal subendothelial collagen)
3.abnormal platelet function
ACT is a test of the function of the ________ and _________ systems. May also be affected by severe _______
Acticated partial thromboplastic time (APTT) is a test of the ________ and _________systems
prothrombin time (PT) is a test of the function of the ________ and _______ systems
petechia and ecchymosis are seen with what type of abnormalites
vascular and platelet
acquired vascular disorders which can lead to a disorder of hemostasis
2.scurvy: vitamin c is needed for normal callagen formation
3.hyperadrenocorticism- leads to decreased collagen production
typical laboratory findings in animals with hemorrhage resulting form vascular disorders
Bleeding time: prolonged
ACT: Normal
APTT: Normal
PT: Normal
TT: Normal
typical laboratory findings in animals with hemorrhage resulting form thrombocytopenia
bleeding time :prolonged
ACT: prolonged if less than 10,000 platelets
APTT: Normal
PT: Normal
TT: Normal
things that cause acquired platelet dysfunction
NSAIDs, abnormal proteins produced in multiple myeloma, phenothiazines, waste products that accumulate during uremia, FDPs
most common inherited platelet dysfunction problem in animals
von Willebrand disease
results in deficiency of von willenbrands factor
typical laboratory findings in animals with hemostatic defects cuased by acquired platelet function defects or inherited platelet function defects other than von willebrand disease
bleeding time: prolonged
ACT: Normal
APTT: Normal
PT: Normal
TT: Normal
von willebrand factor: Normal
typical labortory findings with von willebrand disease
bleeding time: prolonged
ACT: Normal to prolonged*
APTT: Normal to prolonged*
PT: Normal
von willebrand factor: decreased or absent
*ACT and APTT may be prolonged due to the close association of von willebrand factor and factor VIII. decreased vWF may reduce the activity of factor VIII:C even though it is present in adequate concentration
acquired coagulation disorders
1.vitamin K
2.hepatic failure
decreased active vitamin K can result from
1.low dietary intake
3.prolonged bile duct obstruction
5.coumarin (moldy sweet clover
laboratory findings in acuired coagulation disorders resulting from decreased active vitamin K
bleeding time: normal
ACT: prolonged
APTT: prolonged
PT: Prolonged
TT:Normal for vit K alone, prolonged in hepatic failure
von willebrand factor: normal
DIC laboratory findings
palelet count:decreased
bleeding time: prolonged
AT-3: decreased
*ACT,APTT, and PT may be shortened early in DIC due to the marked activation of the coagaulation cascade, but in most cases these are prolonged by the time the affected animal is presented to the veterinarian. TT can be expected to be prolonged if plasma fibrinogen concentration is decreased
reasons for doing a bone marrow
1.unexplained cytopenia
2.searching for the presence of neoplastic cells
3.staging of lymphomas
4.fever of unknown origin
5.principal means of evaluating the response to anemia in the horse-rarely done
bone marrow aspiration preferred sites and dog:
2.cow and hores:
1.iliac crest, femur, humerus
2.sternum, rib
3.proximal tibiotarsal joint just below the femoral-tibiotarsal joint, sternum
bone marrow aspiration cytology versus core biopsy
1.quick turn around time
2.more cellular detail
3.No tissue archiecture
4.cellular density only estimated
5.the most useful and common evaluation
Core biopsy:
1.slower turn around time cellular detail
3.tissue architecture is intact
4.cellular density more accurately assessed
5.less commonly performed
-hypoplastic marrow
-no cells on aspirate
_________ is the most consistent hematologic abnormality in both acute and chronic ehrlichiosis
hematologic abnormalities in acute ehrlichiosis
thrombocytopenia-due to sequwstration, destructionor consumption
bone marrow:megakaryocytic hyperplasia, bone marrow hyperplasia
anemia: suppression of erythropoiesis, destruction of erythrocytes
leukocyte count is variable in the acute phase
laboratory abnormalities in chronic ehrlichosis
pancytopenia <25% of dogs
severe bone marrow hypoplasia
bone marrow plasmacytosis
hyperglobulinemia 50-75%
protein eletrophoresis may be polyclonal or monoclonal
FeLV bone marrow:
may produce hypoplastic bone marrow. hypoplasia may be restricted to one cell line or may be multiple
pure transudate:
modified transudate:
pure transudate:form due to hypoalbuminemia
modified transudate: form due to impaired blood or lymph flow
exudates:form due to increased capillary permeability
transudate or exudate
1.appearance-clear protein<3g/dl
4.clot form-no
1.appearnce-cloudy protein>3g/dl
4.clot form-yes
joint fluid analysis
A. suppurative:
B. monnuclear:
A. usually immune-mediated disease. If septic difficult to see bacteria
B. degenerative disease or trauma
causes of lymphadenopathy
1.hyperplasia (antigenic stimulation)
2.reactive (antigenic stimualtion)
3.inflamed (lymphadenitis)
4.primary neoplasia (lymphoma)
5.metastaic neoplasia
Indications for Transfusion
SEVERE ANEMIA- Numerical guidelines
Acute anemia: Dog PCV < 20, cat < 12
Chronic anemia: Dog < 15, cat < 12
Must consider clinical signs!!!
Indications for transfusion
Acute blood loss (surgery or trauma)
Can use whole blood for RBC and volume
Packed cells and crystalloids preferable
Indications for transfusion
Defect of hemostasis (eg Vit K antagonists)
Replace coagulation factors
Must use fresh whole blood, fresh plasma, fresh
frozen plasma, or cryoprecipitates
Indications for transfusion
Albumin < 1.5 g/dL
Only for acute replacement
Difficult and expensive
Fresh plasma or frozen plasma
Indications for transfusion
Severe thrombocytopenia
Platelet-rich plasma
Used for acute replacement of platelets if
animal needs surgery
VERY difficult to increase platelet numbers
with fresh whole blood
General principles of blood
Antibodies to KNOWN RBC antigens are
used to type
Antibodies are added to RBCs to be typed
Secondary antibodies (against antiRBC
antibodies), +/- complement
Agglutination or hemolysis of positive RBC
Important blood groups of
domestic animals
Dog erythrocyte antigens-
DEA 1.1, 1.2 (A), 7 most important
Cats- 3 blood groups
A, B, AB
Large animals
Zillions of blood groups
Functional significance of
blood groups in animals
Naturally occurring antibodies
antibodies against RBC antigens which occur
due to exposure to a different environmental
antigen (eg microorganism)
Can cause immediate transfusion reactions
Previous sensitization
Through transfusions, pregnancy or vaccines
with blood products
Used to detect incompatibilities between
donor RBC and recipient serum
Detects the presence of antibodies in the
recipient serum which may react with donor
RBC to cause an immediate transfusion rxn
Naturally occurring antibodies
MAJOR crossmatch most important
Dogs may be transfused one time without a
crossmatch because they have few naturally
occuring antibodies
Minor crossmatch
Detects antibodies in donor plasma to recipient
Much less important
Used if large amounts of plasma are to be given
to a patient (eg failure of passive transfer)
Acute transfusion reactions
Acute hemolysis
Naturally occuring antibodies
Antibodies from previous sensitization
Intra- or extravascular hemolysis
Hemoglobinuria, hemoglobinemia
bilirubinemia, bilirubinuria, shock, DIC
Platelet, leukocyte or plasma sensitivity rxns
Allergic reactions
IgE mediated
1-45 min post transfusion
urticaria, pruritis, erythema
Stop the transfusion!!
Circulatory overload- heart failure patients
Citrate toxicity- hypocalcemia
Ammonia toxicity- patients with liver
Transmission of infectious agents
Delayed transfusion reactions
3-21 days after transfusion
due to transfusion of RBC with an
imcompatible blood group
Results in shortened life of RBC
This is not prevented by crossmatch!
Jaundice, fever, + Coombs
Blood donor selection- Dogs
Weight > 25kg
DEA 1 and 7 negative
Easily accessible veins (Greyhound)
Screened for infectious diseases
Initial CBC, chemistry panel, fecal exam
and urinalysis
Blood donor selection- cats
Weight 5-7 kg ideal
No universal donors in cats (types A, B,
PCV > 30%
Initial CBC, chemistry panel, fecal exam,
Blood donor selection- large
No equine or bovine “universal donors”
High degree of blood type polymorphisms
Crossmatch necessary
3 categories of disorders of the immune system
lymphoproliferative disorder
clincal course of animals with complement deficiency
1.recurrent extracelluar bacterial infections
2.not usually fatal
3.Particular infections shown in people- Neisseria menigititis
4.Membranoproliferative glomerulonephritis later in life
5.Neutrophil migration and function appear normal
6.normal antibody response
7.similar incidence of toxocara infections in dogs
8. no adverse concequnces to vaccine with live virus
assessment of complement function
1.amount of complement in canine serum can be assessed by immunodiffusion assay
2.Human hemolytic complement assay can be used on veterinary species, must be serum and requires control animal sample
3. if one or both pathways are deficient consult lab for further tests
neutrophil failure to migrate clinical presentation
bacterial infection with very high blood neutorphil counts
chronic granulatmous diease can be caused by what type of neutrophil problem
failure of intracellur killing
assessment of neutrophil function
2.measure expression of cell surface proteins known to be involved in function-flow cytometry
3.measure functional response to activation:NBT assay, assay to measure oxidative burst
4.measure the ability of neutrophils to phagocytize-rare
rate of adaptive immune system deficiency by cell type
1.b-cell most common
2.b and t cell next most common
3.t cell least common
general characteristics of b cell immunodeficiency
bacterial infections, after 3-6 months when maternal antibody wanes
may be low AB with normal number of b cells or low numbers of b cells
general characteristics of t cell immunodeficiency
infection with intracellular pathogens, fungi, bacteria, protozoa, viral infections- those organisms requiring t cell mediated killing or activation of macrophages for immunity
general characteristics of deficiency of b and t cells
1.severe lymphopenia, hypoglobulinemia
2.usually survive the first few months of life but thats it
3.susceptible to all types of infections
enumerate lymphocyte subsets: flow cytometry
1.lymphocyts can be distinguished by their surface proteins:
t cells:cd8 or cd4
b cells:cd21
2.measures size and scatter of cells
and anitbody binding by fluroescence
lymphocytes are small and not complex
lympocyte function assays
1.lymphocyte blastogenesis:ability of t cells to divide
2.DTH:ability of t cells to divide and migrate
3.vaccine titers:both b and t cell function
to methods of phenotyping lymphocytes
1.immunohistochemistry in formalin fixed tissue-ab react to b or t cells
2.lymphocytes in suspension can be idifed by surface markers
Whats your diagnosis:
lymphocyte pleural effusion in a cat
1.low serum to effusion triglyercide and cholestrol ratios
2.flow cytometry showing mixture of cd4 and cd8 t cells
Whats your diagnosis:
lymphocyte pleural effusion in a cat
1.high serum to effusion triglyceride and cholesterol ratios
2.flow cytometry shows mixture of cd4 and cd8 t cells and b cells
Whats your diagnosis:
lymphocyte pleural effusion in a cat
1.flow cytometry shows single phenotype of lymphocytes (all cd4 t cells,all cd8 t cels, or all b cells)
Whats your diagnosis:
lymphocyte pleural effusion in a cat
1.small lymphocytes
2.flow cytometry show unique phenotype (express both cd4 and cd8 at same time)
PARR assay
PCR for antigen receptor rearrangments:
used to determine if lymphocytes are derived from a single clone.
the presence of a predominat single clone is very specific for the presence of lymphoid neoplasia

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