Pathology 3 Flashcards
What is the predominant inflammatory cell type in acute inflammation?
Neutrophils
What are the predominant inflammatory cell types in chronic inflammation/
Macrophages, lymphocytes and plasma cells
What are the gross characteristics of acute inflammation?
Often abundant exudation of fluid, plasma proteins and leukocytes
What are the gross characteristics of chronic inflammation?
Fibrosis, tissue destruction and repair
What are potential causes of chronic inflammation?
- Persistent infection
- Prolonged exposure to toxic agents
- Some foreign materials
- Immune mediated inflammatory disease
- Unidentified mechanisms
Give examples of microorganisms that may cause persistent infection
- Ones that are difficult to eradicate
- e.g. mycobacteria, Histoplasma capsulatum
Give examples of toxic agents that may lead to chronic inflammation
Barbiturates or aflatoxins (causing chronic hepatitis)
Give examples of foreign materials that may lead to chronic inflammation
- Occurs where these are indestructible
- e.g. plant material, asbestos fibres, some suture material
Give examples of immune mediated inflammatory diseases that may lead to chronic inflammation
- Autoimmune disease e.g. masticatory myositis
- Immunodeficiencies e.g. hereditary defects in leukocyte function
Give an example of an unidentified mechanism that may lead to chronic inflammation
Granulomatous meningoencephalitis (GME)
Describe the morphological features of chronic inflammation
- Tissue destruction
- Attempts of healing: replacement of damaged tissue by connective tissue (fibrosis and angiogenesis) and tissue proliferation
- Infiltration with mononuclear cells (macrophages, lymphocytes, plasma cells)
- Thickening due to proliferation of epithelial cells
- Nodular lesions
What nodular lesions may occur in chronic inflammation?
- Abscesses
- Granulomas
Describe the appearance and cause of abscesses
- Collection of pis circumscribed by fibrous capsule that is visible grossly
- Pus composed mostly of leukocytes, exuded plasma and proteins
- Most commonly caused by bacteria
Describe the development of a granuloma
- Nodular aggregation of macrophages, surrounded by collar of mononuclear leukocytes mainly lymphocytes)
- Macrophages may fuse to form multinucleated giant cells
- Chronic inflammation walled off by fibrotic tissue
What agents may lead to granuloma formation?
- Parasites
- Fungi and algae
- few bacteria e.g. Mycobacteria)
- Few viral infections e.g. Porcine circovirus type 2)
- Foreign body material
Where are monocytes found?
Blood
Where are Kupffer cells found?
Liver
Where are sinus histiocytes found?
Lymph nodes and spleen
Where are alveolar macrophages found?
Lungs
Where are microglia found?
Central nervous system
Where are osteoclasts found?
Bone
What do macrophages, Kupffer cells, sinus histiocytes, alveolar macrophages, microglia and osteoclasts have in common?
Are all derived from monocytes produced in the bone marrow. Differentiate when infiltrate tissue
Describe macrophage migration
- Extravasation of monocytes controlled by adhesion molecules and chemokines
- Reach tissues in similar way to neutrophils in acute inflammation (adhere to wall, emigrate)
What factors are required to activate macrophages?
- Exogenous factors (microbial prodcts, foreign bodies)
- or Endogenous factors (cytokines e.g. IFN-y)
What are the actions of activated macrophages?
- Eliminate injurious agents
- Initiate process of repair
- Responsible for much of tissue injury in chronic inflammation
- Some products are toxic to microbes and also host cells (ROS, NOS), potentiate inflammation (e.g. cytokines) or healing by inducing fibrosis and angiogenesis
What is the connection between lymphocytes ad macrophages?
- Act in bidirectional way
- Recruit and activate each other
- May lead to very chronic and severe reactions
- T cells secrete IFN-y
- Macrophages secrete T cell cytokine IL-12
What is the role of plasma cells in chronic inflammation?
- Activate B lymphocytes
- Produce antibodies direct against persistent foreign or self antigens
What is the role of eosinophils in chronic inflammation
- Present in parasitic and some fungal infections
- Some hypersensitivity reactions
What is the role of neutrophils in chronic inflammation
- Characteristic of acute inflammation, but still present in some forms of chronic inflammation
- e.g. abscesses
Outline regeneration in wound healing
- Proliferation of cells and tissues to replace lost structures
- Complete resolution of lost structures
Outline repair in wound healing
Combination of regeneration and scar formation by deposition of collagen
What is the proportion of regeneration vs scarring dependent on?
- Ability of the tissue to regenerate (i.e. labile vs permanent tissue)
- Extent of the damage
What are the 3 stages of cutaneous wound healing?
- Blood clot and inflammation
- Proliferation and granulation tissue
- Remodelling and maturation
Describe the blood clot and inflammation phase of cutaneous wound healing
- Occurs within 24 hours
- Activation of coagulation pathways
- Blood clot formation
- Neutrophils appear
Describe the proliferation and granulation tissue phase of cutaneous wound healing
- Occurs between 2-7 days
- Macrophages replace neutrophils, clean and promote proliferation
- Proliferation of epithelial cells
- Fibroblasts and endothelial cells proliferate to form granulation tissue
- Macrophages produce FGF to stimulate fibroblasts to produce collagen to carry out fibrosis
- Fibroblasts and connective tissue grow parallel to wound surface and perpendicular to proliferating capillaries
Outline the process of remodelling and maturation in cutaneous wound healing
- Weeks
- Leukocytes and increased vascularity disappear during second week
- Granulation tissue converted into pale, avascular scar composed mainly of fibroblasts and dense collagen)
- Wound contraction occurs in large wounds
Define clinical pathology
The practice of pathology as it pertains to the care of patients
What is the importance of the development, application and interpretation of clinical pathology laboratory procedures?
Can be used for:
- Establishing diagnosis and/or prognosis
- Monitoring treatment of sick animals
- Monitoring animal health
How can clinical pathology be used in healthy animals?
- Monitoring production/performance e.g. metabolic profile of dairy herd
- Monitoring during critical periods e.g. anaemia in piglets, subclinical fatty liver in recently calved cows
- Special purposes e.g. transport/export, selling, animal exhibition, prior to slaughter
What is involved in clinical pathology?
- Haematology, clinical biochemistry, cytology
- Blood, urine, FNAs, effusions, cerebrospinal fluid, lavages, synovial fluid, faeces, ruminal fluid
What is meant by anatomic pathology?
- Biopsy
- Necropsy
What factors may induce variability in clinical pathology results?
- Laboratory quality (pre-analytical, analytical, post-analytical)
- Biological variables
- inter-individual factors
- Intra-individual factors
- Pre-instrumental factors
Give examples of pre-analytical factors that may affect laboratory quality
- Something biological in the patient unrelated to the disease
- Patient preparation
- Sample preparation (e.g. choice of collection tube)
- Shipping (temperature of sample)
Give examples of analytical factors that may affect laboratory quality
- Appropriate equipment/reagents used?
- Analytical sensitivity, specificity, precision
Give examples of post-analytical factors that may affect laboratory quality
- Results given for the correct patient
- Appropriate interpretation
- Diagnostic sensitivity and specificity
Give examples of biological variables that may induce variability in clinical pathology results
- Species, breed, age and sex
- Intra-individual variation
What is meant by inter-individual factors in relation to clinical pathology?
- Inherent differences between groups of animals due to
- Species e.g. cat PCV < dog PCV
- Breed (akitas have lower mean cell volume)
- Age (growing dogs have lower PCV and total protein concentration than adult dogs)
- Sex (males usually have higher PCV values)
What is meant by intra-individual factors in relation to clinical pathology?
- Transient differences in the ame animal due to usually environmental or external factors
- Can induce outlier results when compaed to usual reference ranges for disease-free animals
- Can be minimised by standardising procedures e.g. fasting before sampling
Give examples of factors that may lead to intra-individual variation
- Fasting/post-prandial samples very different
- Diet: low protein diet = low blood urea
- Excitement/anxiety: increase glucose and lymphocytes in cats, adrenergic response increases PCV in horses
- Reproductive status: lactation leads to reduced serum Ca
- Drugs/therapy e.g. glucocorticoids increase ALP and ALT in dogs
- Method of blood sampling
- Sampling site e.g. mammary vein has less glucose vs jugular in lactating cows
Give examples of pre-instrumental factors that may affect the clinical pathology of a blood sample
- Poor sampling method e.g. pumping of syringe punger
- Haemolysed, lipaemic or icteric plasma
- Incorrect tube choice
- Wrong anticoagulant :blood ratio
- Transportation of specimen
- Storage of specimen
What is the difference between serum and plasma?
- Serum: forms 3 layers, blood at bottom, gel in middle, serum at top. No proteins
- Plasma: need to centrifuge blood to get plasma. Contains proteins
What are the advantages of serum?
- Plasma not recommended for some analyses e.g. direct bilirubin, bile acids, haptoglobin, protein electrophoresis
- Autoanalysers may work better with serum
- Most “normal values” are for serum
What are some disadvantages of serum?
- Separation times time for clots for form (unless serum accelerator used, can interact with some assays)
- Separation may be more likely to result in haemolysis, lower yield
- Cannot be used to measure fibrinogen
Why must plasma and serum be separated from blood as soon as possible?
- Some compounds may not be stable in whole blood (e.g. glucose)
- Red cells can haemolyse
- Haemoglobin coloured, will affect test results where colour is assessed
Describe appropiate storage and transportation of serum/plasma samples
- Separated from blood asap
- Closed tubes, cool and dark
- Get sample to lab without delay
- Storage: freeze serum (-20) and plasma (-70)
What is the anticoagulant used in clinical pathology haematology?
EDTA
What samples are used in clinical chemistry for clinical pathology?
- Li-heparin plasma
- EDTA plasma
- Plain serum
What sample type is used for glucose tests in clinical pathology?
Fluoride oxalate
What sample type is used to assess haemostasis/coagulation for clinical pathology?
Citrate
When should EDTA not be used?
When measuring potassium and ionised calcium values, as K-EDTA increases K and decreases Ca (binds out calcium to block clotting cascade)
Describe the appearance of haemolysed, icteric, and lipaemic blood samples
- Haemolysed: red
- Icteric: yellow
- Lipaemic: opalescent/cloudy
What is the effect of haemolysis in a sample on laboratory tests?
- Increases palsma/serum values for some compounds/enzymes due to higher concentration within RBC
- Decreases plasma/serum values of compounds due to lower concentration in RBCs and being diluted
- Interferes with determination by colour interferences or chemical interactions
Give examples of compounds that are found in higher concentrations within RBCs vs plasma
- K+ in horses, inorganic phosphate
- Enzymes e.g. lactate dehydrogenase, AST
Give examples of compounds that are found in lower concentrations in RBCs vs plasma
- Ca
- Glucose
- Mg in cattle
Give examples of the interfering effect of haemolysis on colour determination and chemical interactions in a sample
- Bilirubin falsely increased due to similar absorbance range to haemoglobin
- Chemical interaction: creatinine (Jaffe colorimetric method) falsely decreased in ketotic plasma
How can lipaemia in a sample be avoided?
Fast patients appropriately prior to sampling
What are the main changes in a sample, associated with lipaemia?
- Increase in total lipid, TAG and cholesterol
- many determinations either cannot be carried out or results significantly affected
How does lipaemia increase or decrease values of some compounds in plasma/serum?
- Presence of extra lipid fractions
- Turbidity caused by the lipids
What is the importance of vaildation methods and techniques in clinical pathology?
Aim is to remove/account for instrumental variation
How are instrumental factors causing variation in results controlled?
- Validation of techniques used (assay parameters, machine set up, analytical range) - must be known before analysis starts
- Quality control must be performed regularly
- Can batch samples or have continuous analysis, but QC important for both
What factors are included in the validation of a clinical pathology technique?
- Precision (repeatability, reproducibility)
- Accuracy (measuring right thing correctly)
- Specificity (ability of a technique to measure one single analyte in a complex solution)
- Sensitivity (range and linearity, interval between lowest and highest concentrations method can measure)
- Other
In what 2 situations are imprecise and accurate results possible in clinical pathology?
- Where there are wide differences between physiologic and pathologic state
- Where the use of mean of repeated measures used to increased precision
Explain how post-instrumental or analytical factors may affect clinical pathology interpretation?
- Validation of the result (analytical and biological), is the clinical interpretation of the result
- Transfer of the result to the final user: verbal vs written, easy to make mistakes
- Archiving the result and storing the specimen provides opportunity for reexamination
What is meant by test precision?
- Repeatability/reproducibility
- The ability of a technique to give the same result when the same specimen is analysed several times
How is repeatability of a test assessed?
- Is the within assay precision
- Consecutive measurements within a series of tests
- Usually assessed by replicate QCs in an assay
- Gives intra-assay coefficient of variation
How is reproducibility of a test assessed?
- Is the between assay precision
- Different series of measurements, usually assessed by running same QCs across different assays
- Inter-assay coefficient of variation produced
Define test accuracy
- The ability of the test to measure the right thing correctly
- I.e. the agreement between the true value and the observed value
How is test accuracy assessed?
- Measurements are repeated in order to minimise the influence of imprecision
- Given as percentage: (true value-mean of measurements)/true value
Define assay specificity
- The ability of a technique to measure one single analyte in a complex solution
What may affect the specificity of an assay?
Presence of similar molecules e.g. testing for cortisol, but also picks up other similar shaped steroids = low specificity
Give examples of specificity interfering substances
- Substances that interact with analytical techniques e.g. drugs, detergents, anticoagulants, pollutants
- Ascorbate (vit C, reducing agent) interferes with glucose oxidase reaction of urine test strips
- EDTA (anticoagulant) interferes with calcium, magnesium, iron and potassium
Define assay sensitivity
- Interval between lowest and highest concentrations the method can measure
- The analytical range of a technique is the range of values between which a measurement of the concentration of the analyte is possible with a known imprecision
What is the formula for coefficient of variation?
Standard deviation/mean
Compare the Gaussian curves of a precise and an imprecise technique
- Precise: narrower curve, more results around the mean
- Imprecise: wider curve, wider distribution of results around the mean
Outline the distribution of fluid in the body
- 60% water
- 2/3rds intracellular, 1/3rd extracellular
- Extracellular: 80% interstitium, 20% in plasma
Give examples of how abnormal fluid distribution may occur
- Imbalance between intracellular and interstitial compartments may lead to swelling/shrinkage, potential for serious cell injury
- Imbalance between intravascular and interstitial components may lead to oedema
What are the main mechanisms of oedema?
- Increased microvascular permeability
- Increased intravascular hydrostatic pressure (high BP)
- Decreased intravascular osmotic pressure
- Decreased lymphatic drainage
What might cause increased microvascular permeability in oedema?
- Inflammation
- Toxins
- Anaphylaxis
- Clotting abnormalities
What may cause increased intravascular hydrostatic pressure?
- Portal hypertension
- Pulmonary hypertension
- Localised venous obstruction
- Fluid overload
- Hyperaemia
What may cause decreased intravascular osmotic pressure?
- Decreased albumin production (liver disease)
- Increased albumin loss
- Water intoxication
What may cause decreased lymphatic drainage in oedema?
- Lymphatic obstruction/compression
- Lymphangitis
- Lymphangiectasia
- Lymphatic aplasia/hypoplasia
What are some effects of oedema?
- Decreased wound healing/clearance of infection
- Pulmonary oedema leads to decreased oxygen diffusion
- Brain oedema is acutely life threatening
What is haemostasis?
Arrest of bleeding
Outline the process of haemostasis
- Formation of platelet plug (primary haemostasis)
- Formation of fibrin meshwork (secondary haemostasis)
- Removal of platelet/fibrin plug (fibrinolysis, thrombus retraction)
Outline the formation of the platelet plug (primary haemostasis)
- Transient arteriolar vasoconstriction
- Platelet adherence and activation
- In minimal vascular injury, platelet plugs alone may resolve the damage
Outline the formation of the fibrin meshwork
- Local activation of coagulation results in fibrin polymerisation
- Fibrin “cements” platelets into definitive secondary haemostatic plug
Outline the removal of the platelet/fibrin plug
- Release of tissue factors: thrombomodulin (stops the clotting cascade) and tPA (tissue plasminogen activator, stimulates fibrinolysis)
- Fibrin-platelet thrombus dissolved after healing of the vessel (thrombolysis)
- Fibrin dissolution (fibrinolysis) initiated immediately on vessel injury by plasmin
What are the 2 pathways of the coagulation cascade and what is the result?
- Extrinsic
- Intrinsic
- Both activate factor X, both produce proteolytic enzymes that initiate fibrin formation
Outline the extrinsic coagulation pathway
- Occurs outside the blood vessel wall when shed blood contacts tissue debris
- Thromboplastin plays major role
- Produces tissue factor VIIa
Outline the intrinsic coagulation pathway
- Triggered by effects of abnormal surfaces on components normally present in blood
- All coagulation factors are present in normal plasma
- Leads to cascade of complex enzymatic reactions until factor X activated
