Lab 6 - WBC Flashcards
Infection could be
Parasitic,
bacterial (septic),
rickettsial,
fungal,
viral (infectious agent!!)
Inflammation could be like:
- Infection:
- endotoxin mediated
- sterile necrosis (i.e.
- pancreatitis,
- tumour, trauma),
- chemical (injections, toxins, acids, alkalines),
- immune mediated (SLE)
To evaluate the severity, locality and ethiology of inflammatory processes what are needed?
Laboratory examinations are needed.
Laboratory examinations:
- Haematology (qualitative and quantitative=complete blood count, CBC)
- Erythrocyte sedimentation rate (ESR) cytology
- serum biochemistry
- routine biochemical parameters (substrates, enzymes)
- acute phase proteins
- microbiology
- parasitology
- immunology (ELISA, RIA etc. methods)
- molecular biology
- morphological pathology,
- histology
What is nessesary for White Blood Cell (WBC) counting
For WBC counting it is necessary to use anticoagulated blood
For WBC counting - what is used as a anticoagulant?
Na2EDTA, K2EDTA, or Na- citrate can be used as anticoagulant.
In case of bird, reptile, fish samples Li-, or Ca heparin is used.
Methods used for
White Blood Cell (WBC) counting
- WBC counting by using Bürker-chamber
- WBC counting by hematology analyser
- Automatic cell counters can count WBC (electrical impedance of a particle (cell))
- Laser cell counters
Qualitative blood count
Smear - Staining
After preparing smears several STAINING methods can be used, i.e.
- May-Grünwald,
- Romanowsky,
- Giemsa,
- Diff-Quick etc.
The smears should be analysed at the edge of the slide and in the middle, too.
How to perform Qualitative Blood cell count
- Prepare smare
- Stain
Low magnification - High magnification. First low power should be used, then high.
1000x magnification can be provided by using an immersion lens (and immersion oil).
- One should count at least 50-200 cells and differentiate according to the morphological pattern of the cell types and make the ratio in % among the different cell types:
Qualitative blood count
Neutrophil granulocyte cell line:
- Myeloblast
- Promyelocyte myelocyte
- Metamyelocyte (Jugend)
- Band forms (stab)
- Segmented forms
Qualitative blood count
Lymphoid cell line:
- Lymphoblast
- Small lymphocyte
- Middle sized lymphocyte
- Reactive T lymphocyte
Qualitative blood count
Eosinophil cell line:
- Young form (band nucleated) eosinophil granulocyte
- Segmented form eosinophil granulocyte
Qualitative blood count
Basophil cell line:
- Young form (band nucleated) basophil granulocyte
- Segmented form basophil granulocyte
Qualitative blood count
Monocyte cell line:
- Monocytic (young) form
- Reactive macrophage form
Function of different white blood cells
The function of different white blood cells is mentioned during the lectures.
Absolute WBC numbers
Relative % is defined routinely, but it is useful to know the absolute number of the different cell types.
Neutrophil granulocyte ratio is 70 % in a dog. This is physiological.
Example:
WBC count: 20 x109/l (high!)
Neutrophil granulocyte: 70 %
Lymhocyte: 29 %
Eosinophil granulocyte: 1 %
This value is pathological: 14 x10^9/l (normal: 3-11,8 x10^9/l).
Diagnosis: neutrophilia.
2.
Lymphocyte is 60 % in a cat, that seems to be increased.
WBC count is 3,5 x109/l.
WBC count: 3,5 x109/l (low!)
Neutrophil granulocyte: 40 %
Lymhocyte: 60 %
Absolute neutrophil granulocyte No is 1,4 x 10^9/l (normal: 3-11,8 x109/l).
Diagnosis: neutropenia.
WBC (x 109/l) in Swine, Sheep, Cow and Rabbit
Swine 7 – 20 (x 109/l)
Sheep 4 – 12 (x 109/l)
Cow 4 – 12 (x 109/l)
Rabbit 4.5 – 11 (x 109/l)
Lymphocytes (%) in Swine, Sheep, Cow and Rabit
Swine 40 – 60 (%)
Sheep 40 – 70 (%)
Cow 40 – 70 (%)
Rabbit 40 – 80 (%)
General normal values for dog, cat, horse, humans:
Neutrophil granulocyte
metamyelocyte (Jugend)
segmented form
% and Absolute No x10^9/l
1. Absolute No x10^9/l
Neutrophil granulocyte = 3-11,8 x10^9/l
- metamyelocyte (jugend)= 0-0,3 x10^9/l
- segmented form = 3-11,5 x10^9/l
2. %
Metamyelocyte (jugend) = 0-3%
segmented form = 60-77%
General normal values for dog, cat, horse, humans:
Lymphocyte
Absolute and %
% = 12-30 %
Absolute = 1-4,8 *10^9
General normal values for dog, cat, horse, humans:
Eosinophil granulocyte
% and Absolute
% = 2-10 %
Absolute = 0,1-1,35 *10^9
General normal values for dog, cat, horse, humans:
Basophil granulocyte
%
< 1 %
General normal values for dog, cat, horse, humans:
Monocyte
% and Absolute
% = 1-5%
Absolute = 0,1-0,5 *10^9
Haematological analysis of inflammatory processes
Where do you find the White blood cell (WBC) pools in the body:
- In the bone marrow:
- mitotic- (very young cells, proliferating),
- maturation- (under differentiation) and
-
storage- pools (mature, differentiated WBCs).
2. In the blood vessels: - marginal pool, cells attached to the inner surface of the blood vessels (mostly neutrophil granulocytes),
- Cells can be mobilized very quickly from this pool!
3. In the circulating blood: - we take our sample from here,
- the circulating pool.
- In the tissues of different organs: tissue pool.
What is Physiological leukocytosis and what happens?
Physiological leukocytosis develops due to acute or chronic stress (epinephrine, norepinephrine, glucocorticoids).
- The effects of catecholamines are visible within seconds (neutrophilia, lymphocytosis- as cells are mobilized from the marginal pool /cells attached to the vessels, lymphatic vessels, and organs/).
- The effects of ACTH or glucocorticoids can be detectable after hours (or in some instances minutes) (neutrophilia, lymphopenia, eosinopenia).
- The older neutrophils are mobilized from the bone marrow and marginal pools, they do not die too early due to the stabilisation of the lysosomal membranes, meanwhile, there is a severe lympholysis, and the sequestration of lymphocytes to the lymphatic organs, eosinophil granulocytes are NOT released to the circulation from the bone marrow.
In cats, stress can cause extreme neutrophilia, as the marginal pool is bigger (2/3 of the total neutrophilic count) in cats.
Cause of
Pathologic leukocytosis/leukopenia
Acute inflammation
The very first laboratory sign of inflammation is the:
1. increase of positive acute-phase proteins (APP) in the blood and the
2. decrease of negative acute-phase proteins. (see later in details)
Neutropenia
When does it happen
What is it caused by
- During the first period of the inflammatory process, or widespread severe inflammation = NEUTROPENIA.
- It is caused by the migrating factors (i.e. leukotriens, interleukins etc.) produced by tissue cells, macrophages.
Neutrophils are migrating out of the blood vessels to the site of inflammation (into the tissues), and this can lead to a transient decrease in the total WBC count.
We usually do not take blood samples in this very early phase of inflammation, that is why many people are convinced, that inflammation always causes a high WBC count!
Neutrophilia
When does it happen
What is it caused by
1. During later phases of inflammation = NEUTROPHILIA occurs due to the (G-CSF), or GM-CSF produced by the macrophages.
2. These factors stimulate WBC production in the bone marrow, which leads to an increased total WBC count.
What is “Left Shift”
When More young WBCs appear in the circulating blood.
- Young metamyelocytes (Jugend), and
- Band (stab) forms are visible in greater proportion and absolute number in the blood smear.
First, cells are mobilized from the marginal pool, then differentiated forms are migrating from the storage pool of the bone marrow.
Later younger cells enter the bloodstream from the maturation pool of bone marrow.
In case of severely increased tissue requirement (or in neoplastic diseases i.e. acute myelogenous leukemia) very young cells (myelocytes, promyelocytes) from the mitotic pool are going out.
Two different types of left shifts!
What is a regenerative left shift?
Increased WBC count, neutrophilia, and left shift (younger neutrophils).
After the first neutropenic phase, bone marrow regenerates the utilized neutrophils in the circulation by the younger cell population from the storage, maturation, and/or the mitotic pool.
=This is the most common cause of the appearance of young neutrophils.
Regenerative left shift is a sign of a favorable prognosis.
What is Degenerative left shift?
Low, or normal WBC, and neutrophil count, and left shift (younger forms).
1. If there is a great neutrophil utilization in the tissues
(widespread or very severe inflammation, big abscesses, peritonitis, pleuritis, phlegmone, pyometra), and the
2. Rate of utilisation is bigger than the regenerative capacity of bone marrow,
= increased outflow of young neutrophils is NOT enough to increase WBC count
(or even for the maintenance of normal WBC count!).
Degenerative left shift is a sign of poor prognosis, it means that the disease is very serious and needs immediate treatment.
What happens due to Leukemoid reactions
In some cases, when there is a great stimulus for producing neutrophil granulocytes in an enormous number (WBC count can be >70x10^9/l) of neutrophils can be present in the peripheral blood due to increased effects of G-CSF, GM-CSF.
This is a typical reaction in case of big abscesses, endometritis (pyometra), and in the case of some neoplastic (but not leukaemia) diseases. During blood smear analysis, a leukemoid reaction can be easily confused with chronic myelogenous leukemia.
What is Toxic Neutrophils
In very severe inflammatory processes the granulocytopoesis can be disturbed.
When the granule production is NOT physiological, some azurophilic (orange-red) granules can be seen in the basophilic cytoplasm.
These are “toxic” neutrophils, or we can say that this process is the toxic granulation of neutrophils.
What is Döhle-bodies?
Due to toxic effects some angular, basophilic inclusion bodies can be seen in some neutrophil granulocytes.
= These are the remnants of the endoplasmatic reticulum.
The appearance of Döhle- bodies is more common in cats.
Which shift is characteristics for Chronic inflammation
Right shift
Chronic Inflamation
What is characterizing the right shift?
Chronic inflammation
Right shift:
The right shift= many segmented and hypersegmented, old neutrophils (3-4 segments are on one nucleus) are seen in the smear.
The appearance of these older forms + increased WBC count = typical chronic inflammatory process.
This phenomenon is also typical for the effects of glucocorticoids (given as a therapeutic drug for different diseases, or increased production of endogenous glucocorticoids in the adrenal glands, Cushing’s disease).
Glucocorticoids inhibit cellular proliferation (disappearance of young cells), and have membrane stabilizing effect (which let the neutrophils to grow old and become hypersegmented).
Moreover, glucocorticoids are lympholytic and inhibit the outflow of eosinophils from the bone marrow (it can cause lymphopenia and eosinopenia). This is a common finding in the case of MACROCYTOSIS of POODLES..
Right shift, leukocytosis, neutrophilia, lymphopenia, and eosinopenia together is called a stress leukogram.
Typical signs of chronic inflammation in the CBC are similar to the glucocorticoid effect (leukocytosis, neutrophilia, right shift),
- the difference is that in this case leukocytosis is sometimes associated with lymphocytosis, monocytosis, and/or eosinophilia.
Other typical changes are
- Haematology of Addison’s disease (hypoadrenocorticism):
- Pelger-Huet anomaly:
3. Cyclic neutropenia:
- Bone marrow damage:
Other typical changes
Hematology of Addison’s disease (hypoadrenocorticism):
There is no inhibitory effect of glucocorticoids, as there is hypoplasia or necrosis of adrenal gland.
The typical changes are an increase in WBC (due to the polyuria caused by increased PCV), an increase in young neutrophils, left shift (no inhibition of cell proliferation), lymphocytosis and eosinophilia.
Other typical changes
What is Pelger-Huet anomaly:
= Normocytaemia, and left shift (metamyelocytes, band forms of neutrophils)
= Inheritable problem causes diagnostic problem.
Other typical changes
What is Cyclic neutropenia?
Occur in which case?
This is an inheritable disease of Grey Collies.
Due to cyclic bone marrow activity,
= Neutropenia occurs in weekly, monthly intervals.
During these periods the animals are sensitive to infections.
Other typical changes
Bone Marrow damage
Leukopenia and neutropenia occur in case of
1. bone marrow damage or
2. decreased bone marrow function.
Thrombocytopenia and aplastic anemia is often accompanied.
When other cells suppress the hemopoietic cells in bone marrow, it is called: MYELOPHTYSIS
Description of Schilling-phases
Description of Schilling-phases are found in the lecture notes
Name Other tests to examine the inflammatory process.
- Glutaric aldehyde test
- ESR - Erythrocyte Sedimentation Rate
- CRP
Characterize Glutaric Aldehyde test
Test is used to determine what?
Used to examine the = increase of fibrinogen and globulin concentration in plasma.
- As fibrinogen = acute-phase protein, it can show a marked increase during acute inflammatory processes.
This method is used in clinical practice to indicate inflammatory processes in adult cattle (more than one year old).
- Glutaric aldehyde solution causes rapid coagulation of fibrinogen and labile globulins, so blood mixed with this solution can show coagulation within seconds in case of an acute inflammatory process
(high fibrinogen and globulin level in the blood),
Even if the blood sample contains heparin or EDTA (as an anticoagulant).. The coagulation time is inversely correlated to the concentration of fibrinogen and globulins in the blood.
We generally use heparinized blood and mix the sample (i.e. 1-2 ml) with the same amount of 1,25% glutaric aldehyde solution.
Tubes (or syringes) should be checked for coagulation every 30 seconds by turning them upside down.
If the symptoms and the results are not correlated to each other, it is necessary to repeat the test 24-48 hours later.
The test is primarily used to diagnose RETICULOPERITONITIS, SEVERE MASTITIS or ENDOMETRITIS in cattle. These animals can be extremely indolent, sometimes they show just vague clinical signs, (or no signs at all!!) even if they are suffering from very severe diseases!
Aid for the evaluation of glutaric aldehyde test in cattle
- Coagulation time
- Presence of the inflammatory proteins in the sample
- The character of the pathological process
Aid for the evaluation of glutaric aldehyde test in cattle
Coagulation time < 1 min Indicates?
Presence of the inflammatory proteins in the sample = ++++
The character of the pathological process = Severe, widespread acute inflammation
Aid for the evaluation of glutaric aldehyde test in cattle
Coagulation time 1-3 min Indicates?
Presence of the inflammatory proteins in the sample= +++
The character of the pathological process = Acute inflammation
Aid for the evaluation of glutaric aldehyde test in cattle
Coagulation time 3-8 min Indicates?
Presence of the inflammatory proteins in the sample = ++
Character of the pathological process = Subacute inflamation
Aid for the evaluation of glutaric aldehyde test in cattle
Coagulation time 8-15 min Indicates?
Presence of the inflammatory proteins in the sample= +/-
Character of the pathological process= Not severe inflammatory process
Aid for the evaluation of glutaric aldehyde test in cattle
Coagulation time >15 min Indicates?
Presence of the inflammatory proteins in the sample = -
Character of the pathological process= There is no inflammatory process
Evaluation of glutaric aldehyde is used for?
The test is primarily used to diagnose reticuloperitonitis, severe mastitis or endometritis of cattle.
- These animals can be extremely indolent, sometimes they show just vague clinical signs, (or no signs at all!!) even if they are suffering from very severe diseases!
Characteristics of
Erythrocyte sedimentation rate (ESR)
Theory
= The theory of ESR is the increased sedimentation of RBCs due to inflammatory processes, as the acute phase proteins and other globulins tend to attach to the surface of RBCs.
Normally RBCs have negatively charged albumin molecules on their surface
= prevents RBC aggregation.
In case of high globulin level in the blood
(severe inflammation or some types of neoplastic diseases can cause this),
= The surface of RBCs is covered by globulins, instead of albumins. Globulins have no negative charge= RBCs are able to attach to each other, form big aggregates and sediment quickly.
- Increased globulin concentration causes increased ESR, and this is generally due to inflammatory or neoplastic processes.
- Decreased albumin concentration and as a consequence relatively increased globulin concentration causes increased ESR, too.
(Note: in case of chronic renal failure, or glomerulonephropathy when there is hypoalbuminemia ESR is typically increased.)
The method for ESR
Bullet points
- Type of tube
- Tube containing
- When to check the sedimentation rate?
- Phsiologically ESR?
ESR proportional too?
What type of sedimentation process? In case of which case?
Sedimentation rate in horses
1. Specific tubes (those made of glass are called: Westergreen tubes are used to perform this test. The tube contains Na-citrate, and there is a mm scale on it. The tube has a hole on the bottom (and on the top), so when it is driven through the rubber plug of another tube containing the blood sample, blood will flow up in the tube with the scale until the zero point.
The sedimentation rate of RBCs should be checked in the tube after one hour.
2. Physiologically ESR is 0,5-3 cm/hour.
3. ESR is inversely proportional with the Ht (PCV).
- The sedimentation can be biphasic when the young RBCs sediment later than the older forms.
= This is common in case of immunhemolytic anemia (regenerative anaemia, many young RBCs are produced). In this case, the older and even aggregated RBCs sediment much earlier than the younger forms.
5. Sedimentation is very fast in horses, so ESR must be evaluated after 20 minutes. It is found that the speed of the sedimentation decreases (!) due to inflammatory processes contrary to other species.
6. ESR is inversely proportional with the Ht (PCV), and proportional with the serum viscosity, total protein and fibrinogen concentration.
ESR of dogs
Ht (PCV) - ESR mm/hour
10 = 79
15 = 64
20 = 49
25 = 36
30 = 26
35 = 16
40 = 10
45 = 5
50 = 0
ESR of Horses
Ht (PCV) - ESR mm/20 min.
10 = 86+-1
15 = 80+-1
20 = 70+-2
25 = 60+-3
30 = 47+-5
35 = 28+-15
40 = 11+-8
45 = 2,5+-1
50 = 0+-1
Characteristics of CRP
C-reactive protein=CRP
(origin of the name of this acute-phase protein: it can bind non-specifically to a protein of Pneumococcus bacteria, called protein-C, and cause precipitation)
In the case of acute inflammatory processes the concentration some of acute phase proteins becomes increased in the blood plasma.
= Such acute phase protein is CRP, haptoglobin, serum amyloid A (SAA) etc.
Some „negative” acute phase proteins (i.e. transferrin, lactoferrin) show a decreased concentration due to acute inflammatory process.
CRP is produced in the liver and in the beginning of the inflammatory process cytokines (interleukins) stimulate its production.
For the measurements of CRP we use serum samples.
The determination is based upon the immunological (ELISA or immunturbidimetric) method.
Today species (dog, horse) specific ELISA methods are available.
General physiologial value: 8 mg/l
In the beginning of the acute inflammatory process, CRP values are increased, before the appearance of the clinical signs. (In case of concomitantly decreased liver function this increase might not be so marked.) Therefore it is a very useful diagnostic tool for the early detection of inflammatory processes.
This can be extremely important in immunosuppressed animals, for example during chemotherapy, or high dose glucocorticoid therapy.
= In these cases, the well-known clinical and laboratory signs of inflammation can be vague or absent!
There are the most common hemopoetic tumors known in animals?
The most common leukemic types.
1. Acute leukemic diseases:
2. Chronic leukemic diseases:
3. Lymphomas are hemopoetic tumours, too.
Characterize 1. ACUTE LEUKEMIC DISEASES
1., Acute leukemic diseases:
Typical cell types in these tumors are „blast” cells, that have coarse chromatin pattern, and have nucleoli.
The origin of these cells can be evaluated by (immune)cytochemical, and bone marrow analysis. In some cases, neoplastic cells do not appear in peripheric blood,
- but anemia, leukopenia, thrombocytopenia. In these cases bone marrow evaluation is important.
Acute lymphoblastic leukemia
Acute myeloblastic, promyelocytic leukemia
Acute erythroblastic leukemia
Lymphoma of Stage V (metastatisis to bone marrow)
Characterize 1. CHRONIC LEUKEMIC DISEASES
2. Chronic leukemic diseases:
Typical cell types in these tumors are mature differentiated, or well differentiated cells, that appear in enormous numbers in the peripheric blood.
- This presentation does NOT cause diagnostic difficulties in the case of
lymphoid,
thrombocytes, or
erythroid forms,
but it can be mixed with a leukemoid reaction.
In order to diagnose these diseases bone marrow evaluation is essentially required.
Chronic small lymphocytic leukemia,
Chronic myeloid (neutrophil granulocytic, eosinophil granulocytic, basophil granulocytic,
monocytic leukemia)
Polycythaemia absoluta vera (overproduction of mature erythrocytes),
Essential thrombocytosis (overproduction of mature thrombocytes).
Characterize 3. LYMPHOMAS
Lymphomas are hemopoietic tumors, too.
In this disease, poorly differentiated lymphoid cells are overproduced in lymphatic organs (lymph nodes, spleen) and sometimes in other tissues (liver, skin, intestines etc.).
In some cases, tumorous lymphoid cells are overproduced in bone marrow, too.
The origin of this is proved in the case of cats and cattle, only.
In these species, viral infection is causing this disease
(Bovine Leukosis Virus /BVL/ és a Feline Leukemia Virus /FeLV/).
Bloodcells =
- Erythrocytes
- Leukocytes
- Thrombocytes
WBC =
- *WBC (leukocyte)**
- *Sample anticoagulated blood** (Na2EDTA, K2EDTA, or Na-citrate)
Normal WBC count: 6-12x 10^9 (G/l) (G=giga)
• Counting:
1. Bürker-chamber method
2. Estimated RBC-count
3. Automated cell counters
• Evaluation of the blood-smear
Blood smear to examine
● RBC morphology: spherocytes, Heinz bodies, RBC parasites.
● RBC autoagglutination
● Thr.cyt aggregation
● Thr.cyt count & morphology
● WBC differential count, morphology
● Neutrophil: left shift, toxic change
● Atypical cells / abnormal blast cells
Preparation of blood smear
Staining methods
● May-Grünwald
● Romanowsky
● Giemsa
● Diff-Quick
Neutrophil granulocytes
Neutrophil granulocyte cell line:
● Myeloblast
● Promyelocyte
● Myelocyte
● Metamyelocyte (jugend)
● Band forms (stab)
● Segmented forms
(Mobblast Prom Mye Metamye Band Segmented)
Neutrophil granulocytes
Size
● 1,5-2x RBC size
Neutrophil Granulocyte
Morphology
● Segmented nucleus into lobes
● Neutrophil, or the heterophil (birds, reptiles and some mammals) granules in CP
● Band neutrophils elongated, U‐shaped, non-lobulated nucleus
Neutrophil Granulocyte
● Major pools:
● Bone marrow
● Blood
● Tissue
Neutrophilia
Different cases
Neutrophilia
1● Left shift:
a● Regenerative left shift:
b● Degenerative left shift:
2● Right shift:
Neutrophilia
In the case of left shift
1● Left shift: release of immature neutrophils (predominantly band forms) from the bone marrow.
a● Regenerative left shift:
● Neutrophilia in which mature neutrophils exceed bands
● Good prognostic indicator
b● Degenerative left shift:
● Low or normal Ne. gr. count and band cells exceed mature neutrophils.
● Poor prognostic indicator
2● Right shift:
● Mature neutrophilia
● Hypersegmented neutrophils have five or more lobes in the nucleus
What is Toxic Neutrophils
Toxic neutrophils
● Enhanced neutrophil turnover and reduced maturation time
● Severe inflammatory disease (severe bacterial infections), IMHA
● Not necessarily associated with “toxemia”
● Increased cytoplasmic basophilia, increased residual cytoplasmic RNA
● Frothy or vacuolated cytoplasm, which represents degranulation of llysosomes
● Döhle bodies: these are grey-blue intracytoplasmic inclusions representing aggregates of cytoplasmic reticulum
● Commonly found in normal cats
● Can develop as an artifact in stored samples
Give an example of an Toxic Neutrophils
and a potential disease due to it
Toxic neutrophil due to SEVERE INFLAMMATION = IMHA
Döhle bodies: Normal in cat
Causes of NEUTROPHILIA
1● Physiological neutrophilia: acute stress, adrenaline release (i.e. cat blood sampling)
2● Stress/steroid response:
● Hyperadrenocorticism, Exogenous corticosteroids
● No left shift,
● Stress Leukogram: monocytosis, lymphopenia and eosinopenia
3● Acute inflammatory response
● Bacterial infections, Immune-mediated disease, Neoplasia, Tissue necrosis
● Left shift
● Toxic changes
Causes of Neutropenia
Neutropenia:
1● Peracute bacterial infections
2● Retroviral infection (FeLV,FIV), Parvovirus
3● Bone marrow disorders: aplastic anemia, leukaemia, neoplasia, myelofibrosis, myelodysplasia
4● Cytotoxic myelosuppressive drugs
LEUKEMOID REACTION happens when
Leukemoid reaction
WBC count >70x10^9/l)
Extreme neutrophilia >50% neutrophils
Marked left shift
Increased effects of G-CSF, GM-CSF
Causes of an LEUKEMOID REACTION
● Severe localized pyogenic infections: Pyometra, Pyothorax, Peritonitis, Abscess
● Neoplasia
● Tissue necrosis
● Chronic Myelogenous Leukemia (CML)
Lymphocytes
Morphology
● Size ~ RBC
● Round nucleus
● Smooth, dense chromatin
● Narrow rim of basophilic cytoplasm
● Vary in size
Types of lymphocytes
1● Reactive lymphocytes - immunocytes
● Larger, basophilic cytoplasm
● Reflect antigenic stimulation
2● Granular lymphocytes
● Prominent pink cytoplasmic
Causes of LYMPHOCYTOSIS
• Lymphocytosis:
- Physiological - catecholamines
- Prolonged presence of antigen (vaccination)
- Hypoadrenocorticism
- Lymphoproliferative disorders
Causes of LymphoPenia
Lymphopenia:
- Lymphoprolipherative disorders
2. Stress leukogram
3. Glucocorticoids, immunosuppressants, chemotherapy
4. Viral infection, sepsis
Monocytes
Morphology
Monocytes
● Larger than neutrophils
● Extreme variable shape, no consistent species differences
● Very variable shape of the nucleus:
- *● Ameboid**
- *● Bean-shaped**
● Band-like
● lobulated,
● U-shaped
● Cytoplasm:
- vacuoles,
- granules,
- slightly grainy
Causes for monocytosis
1● Acute/chronic inflammation
2● Tissue necrosis
3● Immune-mediated diseases
4● Stress, endogenous/exogenous steroid response
5● Stress Leukogram: neutrophilia (no left shift), monocytosis, lymphopenia and eosinopenia
6● Monocytic or myelomonocytic leukemia
Eosinophils
Morphology
● Slightly larger than a neutrophil
● Segmented nucleus with 2-3 lobes
● Eosinophilic cytoplasmic granules
● Marked SPECIES VARIATION
1● Dog: granules are round, size, numbers are very variable
- Greyhounds: grey cytoplasm, and vacuolated appearance because their granules do not stain
2● Cat: small uniform, rod-shaped orange-pink granules
3● Horse: looks like a rasberry
Causes of Eosinophilia
1● Parasitic
2● Allergy
- Feline asthma (fe)
- Eosinophilic bronchopneumopathy(ca)
- Eosinophilic granuloma complex (fe)
- Atopy, Food hypersensitivity
3● Inflammatory
- Eosinophilic enteritis (IBD)
- Eosinophilic myositis
4● Hypereosinophilic syndrome
5● Neoplastic Mast cell tumor
6● Hypoadrenocorticism (ca)
Causes of EOSINOPENIA
1● Stress leucogram
2● Glucocorticoids
Basophils
Morphology
● Mildly lobulated, ribbon-like nucleus
● Dark purplish cytoplasmic granules
● Canine: long, folded nucleus; grey, lavender cytoplasm
● Feline: round to oval granules, pale lavender, to deep purple
● Horse: deep purple granules
Causes of BASOPENIA and BASOPHILIA
1● BASOPENIA
● Corticosteroid
2● BASOPHILIA:
● Allergy
● Parasites
● Mast cell neoplasia
What is Acute phase proteins (APPs)
● Extremely sensitive markers of inflammation
● Extremely non-specific
● The decline in APP concentrations may indicate a response to treatment
● Triggered by proinflammatory cytokines:
- interleukin (IL)-1, IL-6 (produced by macrophages→ APPs production in hepatocytes
● Fibrinogen: in all species production is stimulated as part of the acute phase response
1● Major APPs: rapid increase (often within 24–48 hours) 5–50-fold
● Dog: C-reactive protein (CRP)
● Cat: Serum amyloid A (SAA)
2● Moderate APPs: slower increase (within 4–7 days) 2–10-fold
● Dog, cat: Haptoglobin (Hp)
3● Negative APPs: concentrations decline
- Albumin, transferrin
Acute phase proteins
CAT
- Major APPs
- Moderate APPs
MAJOR
- SAA
Moderate
- AGP
- Hp
SAA: serum amyloid A, CRP: C-reactive protein, Hp: haptoglobin, AGP: α1 acid glycoprotein Method: ELISA or immunturbidimetric
Acute phase proteins
DOG
- Major APPs
- Moderate APPs
MAJOR
- CRP
- SAA
Moderate
- Hp
- AGP
SAA: serum amyloid A, CRP: C-reactive protein, Hp: haptoglobin, AGP: α1 acid glycoprotein Method: ELISA or immunturbidimetric
Acute phase proteins
HORSE
- Major APPs
- Moderate APPs
MAJOR
- SAA
Moderate
- Hp
SAA: serum amyloid A, CRP: C-reactive protein, Hp: haptoglobin, AGP: α1 acid glycoprotein Method: ELISA or immunturbidimetric
Acute phase proteins
CATTLE
- Major APPs
- Moderate APPs
MAJOR
- Hp
- SAA
Moderate
- AGP
SAA: serum amyloid A, CRP: C-reactive protein, Hp: haptoglobin, AGP: α1 acid glycoprotein Method: ELISA or immunturbidimetric
Acute phase proteins
SWINE
- Major APPs
- Moderate APPs
MAJOR
- CRP
- MAP
- SAA
Moderate
- Hp
SAA: serum amyloid A, CRP: C-reactive protein, Hp: haptoglobin, AGP: α1 acid glycoprotein Method: ELISA or immunturbidimetric
Characterize
Erythrocyte sedimentation rate (ESR)
● Non-specific measure of inflammation
1● In normal conditions:
- RBCs are negatively charged due to albumin attached to the surfaces
and repel each other
2● In inflammatory conditions albumin↓, fibrinogen↑, globulins (positively charged):
APPs, immunglobulins↑ → RBC aggregation, rouleaux formation, sedimentation↑
3. Method:
● Fresh Na-citrate (1:4)
● Westergren tubes
● Normal values Note: mm/h. = millimeters per hour.
● Eq: 20 min - ESR slower in inflammation
Characterize
Glutaric aldehyde test
● In Adult cattle
● Fibrinogen as APPs markedly increases during acute inflammatory processes
● “Point-of-care” test
● Fresh anti-coagulated (heparinized) whole blood mixed with 1-2 ml glutaric aldehyde solution
● Rapid coagulation of fibrinogen and labile globulins
● Coagulation within seconds in case of the acute inflammatory process
● Diagnose
- reticuloperitonitis,
- severe mastitis,
- endometritis
