Lecture 12 - Bone Marrow Disorders Flashcards
What conditions indicate that there is something going on in the bone marrow?
- Persistent, unexplained cytopenia or cytosis:
* Neutropenia
* Thrombocytopenia
* Pancytopenia
* Non-regenerative anemia - Unexplained rubricytosis (nucleated RBCs)
- Suspected neoplasia or monoclonal gammopathy
- Better classification of leukemia
- Fever of unknown origin; Looking for underlying inflammation, neoplasia
- Rule out osteomyelitis
- Staging lymphoma, mast cell tumor or others
- Monitoring chemotherapy response
Where would you collect bone marrow samples in dogs and cats?
Dogs, cats: Trochanteric fossa, humerus, ilium
Where would you collect bone marrow samples in horses, cattle, and camelids?
Horses, cattle, and camelids: the ilium, ribs, or sternum
What materials are required to collect bone marrow?
- Local or general __________
- Materials:
* Bone marrow ______ needle: ___-____ gauge
* ___cc syringe
* ______ tube or preferable make _______ asap
* air dry, _____ stain
- Local or general anesthesia
- Materials:
* Bone marrow biopsy needle: 16-22 gauge
* 12cc syringe
* EDTA tube or preferable make slides asap
* air dry, Wright’s stain
Illinois sternal aspiration biopsy needle, and a Jamshidi core biopsy needle along with its associated shepherd hook for specimen
removal.
Jamshidi Important to remember here: where should I obtain samples from. You will
go over these techniques later during clinical years
videos are FYI on how the procedure is done.
https://eclinpath.com/cytology/procedure-videos/
https://vod.video.cornell.edu/media/Canine+-+Bone+Marrow+Aspiration-
Forelimb/1_2iytqzo1/45064261
What is the point of obtaining a bone marrow sample?
What do you have to be careful of when collecting the sample?
What do you do in the event that you are unable to obtain a sample?
- Aspirate for cytology –avoid diluting sample with blood–. If can’t obtain - core biopsy for histopathology placed in formalin
- Core biopsy for histopathology
- Do NOT transport/ship these samples together
What is this image showing?
Fig a. Sampling at the craniolateral part of
the greater tubercle of the humerus.
Fig a. Placement of the marrow biopsy needle is shown into the craniolateral part of
the greater tubercle of the humerus. This site is preferred for obese and muscular
adult dogs and cats.
What is this image showing?
Fig b. Sampling at ?
Fig b This location may be most useful for ______ animals. Penetration is made just ______ to the greater trochanter and _______ to the shaft of the samples, avoiding the ______ nerve located more lateral and posterior.
Fig b. Sampling at proximal femur
Fig b The proximal femur may be most useful for small animals. Penetration is made just medial to the greater trochanter and parallel to the shaft of the samples,
avoiding the sciatic nerve located more lateral and posterior.
What is this image showing?
Fig a. Sampling at the ?
Fig.a. This location is popular in ____ or _____-_____ dogs, because it is readily accessible.
Fig a. Sampling at the dorsal iliac crest
Fig.a. The dorsal iliac crest is a popular location in thin or non obese dogs, because it is readily accessible
What is this image showing?
Fig. b. Transilial procedure
Fig. b. One or more samples may be taken via a transilial procedure. This site may be
helpful in young dogs and cats that have a dorsal ilial crest, too narrow to attempt
parallel placement of the biopsy needle.
How do you prepare film for a bone marrow sample?
- Squash or Pull Prep Technique: Bone marrow is collected
- 1- place a drop is placed toward the end of the slide.
- 2-The suspension is spread by placing a second microscope slide over
the sample perpendicular to the slide with the sample and pulling the
two slides apart. The weight of the top slide should be the only
pressure exerted on the sample.
What are the contraindications pf bone marrow sample collection?
- Bleeding disorders
- When you can achieve diagnosis with less invasive tests
- IMHA by CBC and blood smear
What do you have to interpret when analyzing a bone marrow sample?
- Cellularity and morphology
- Presence of megakaryocytes
- Presence of iron stores
- Myeloid:Erythroid ratio
- Orderliness and completeness of maturation
- Presence of other cells, example: Plasma cells
- Presence of abnormal cells
- Presence of microorganisms –rare-
You collect a bone marrow sample, stain it, and are now looking at it under the microscope. What is wrong with this sample?
Inadequate cellularity
Looking at: Overall % hematopoietic cellularity compared to % of adipose tissue
In addition to evaluating the cellularity, what else are you looking at when you analyzing your bone marrow aspirate?
The number of granulocytes to nucleated erythrocytes.
Normal range is 1:1 up to 3:1
What can be seen in the image below?
Normal bone marrow from a dog. Chunks of dark brown-black iron (arrows) released from ruptured macrophages (Wright’s stain, 50x objective).
Iron content can be semi-quantified in a cellular sample (with spicules) using a
Prussian blue stain.
Arrowhead pointing to a megakaryocyte
What can be seen in the image below?
Normal bone marrow from a dog. Chunks of dark brown-black iron (arrows) released
from ruptured macrophages (Wright’s stain, 50x objective).
Iron content can be semi-quantified in a cellular sample (with spicules) using a
Prussian blue stain
Iron within the bone marrow is stored within _______.
Iron stores are depleted in marrow before the development of _____ or ______ ____ red blood cells, this is the only technique for detecting early iron
deficiency.
macrophages, anemia, microcytic, hypochromic
How do you quantify iron storage levels in bone marrow?
No established guidelines for quantifying it. However, iron content can be semi-quantified in a cellular sample (with spicules) using a
Prussian blue stain.
What is the best way to collect bone marrow samples in order to determine the iron levels?
Best from bone marrow spicules (where macrophages are usually
located)
Is evaluating the iron levels required in establishing iron deficiency?
Not required technique in an established iron deficiency for Dx of
Iron def. anemia
* Remember the hematologic characteristics?
What can be seen below? Label the cells accordingly.
Erythroid maturation
1- Proerythroblast, 2 basophilic erythroblast, 3 polychromatophilic erythroblast 4
normoblast 5 reticulocyte 6 erythrocyte
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What can be seen below? Label the cells accordingly.
When interpreting bone marrow aspirate results, what other diagnostics are necessary?
- It is necessary to have recent CBC
- The day of collection or within 24h from collection
- Blood film evaluation simultaneously
What is the difference between a cytology and a core biopsy histopathology?
What does it mean when you are analyzing your bone marrow aspirate and find that there is DECREASED cellularity? What does decreased cellularity result in?
A decrease in cellularity means that there is a severe decrease in ALL hematopoietic cells in the bone marrow. The aspirate will consist of fat with stromal cells (macrophages, dendritic cells, fibroblasts, mast cells, lymphocytes and plasma cells)
Decreased cellularity results in:
* Bone marrow aplasia or bone marrow anemia
- <5% of erythroid or myeloid precursors or no megakaryocytes
* Results in pancytopenia
What are the causes of pancytopenia?
- drugs,
- hormones (e.g. estrogen),
- infectious agents (e.g. Ehrlichia canis),
- radiation,
- immune-mediated
- idiopathic causes
What does it mean when you are analyzing your bone marrow aspirate and find that there is an INCREASE in M:E ratio?
When there is an increase in the Myeloid:Erythroid ratio, it could mean any of the following:
* Erythroid hypoplasia or Aplasia
* Granulocytic hyperplasia
* Granulocytic leukemia
What does it mean when you are analyzing your bone marrow aspirate and find that there is a DECREASE in M:E ratio?
When there is a decrease in the Myeloid:Erythroid ratio it could mean any of the following:
* Regenerative anemia
* Erythroid leukemia
* Lack of production of neutrophils
What does it mean when you are analyzing your bone marrow aspirate and find that there are PLASMA CELLS in your sample?
If there is more than >15% to 20%, this suggests plasma cell myeloma/multiple myeloma if infectious dz testing is Neg.
BUT up to 5% plasma cells dispersed as single cells is normal
What % range of Lymphocytes in a bone marrow aspirate sample is considered to be normal?
1.7-4.9% is normal
What % range of Macrophages in a bone marrow aspirate sample is considered to be normal?
Up to 0.4% is normal
What % range of Mast cells in a bone marrow aspirate sample is considered to be normal?
Seeing mast cells in your bone marrow aspirate sample is RARE.
What does it mean when you are analyzing your bone marrow aspirate and find that there are OSTEOBLASTS AND OSTEOCLASTS in your sample?
Osteoblasts and osteoclasts are the unique cells occurring in bone marrow smears in situations with high bone metabolic turnover (children, trauma, rachitis, Paget disease or tumors).
What are the arrows pointing to in the image below?
Presence of abnormal cells
arrowed red blood cell precursor has a larger nucleus than normal. Mitotic figures can be seen in erythroid and myeloid cells in marrows, but increased erythroid mitoses
were seen (arrowhead) (Wright’s stain, 50x objective).
What types of microorganisms can be found in bone marrow aspiration samples?
- Histoplasma capsulatum
- Toxoplasma gondii
- Leishmania donovani
- Red cell parasites
- Babesia
What can be seen in the image below?
Where in this world can this organism be found?
Once they enter the body, where are they typically found?
Histoplasma capsulatum
It is endemic in North and Central America. Clinical manifestations varies from
pulmonary disease to disseminated multi-organ involvement.
Organisms are seen within macrophages
How do you interpret a bone marrow sample taken from a patient you suspect is infected with a microorganism?
Explain the pathogenesis behind Erlichia canis –> pancytopenia.
Immune-mediated responses play a major role in the pathogenesis of E. canis infection. Anti-platelets antibodies (APA) have been demonstrated less than a week after experimental E. canis infection of dogs. Platelet aggregation abnormalities, anti-nuclear antibodies (ANA), RBC autoagglutination with positive Coombs’ test, and circulating immune-complexes have been shown in infected dogs and are associated with the disease process.
The decrease in platelets during canine ehrlichiosis is probably a result of several mechanisms. These mechanisms include increased consumption with vascular endothelial changes, platelet sequestration and pooling in the spleen, thrombophagocytosis with immunological destruction, a decrease in the half life time of circulating platelets possibly due to opsonization with antibodies, and production impairment due to bone marrow destruction and hypocellularity. In addition to the decrease in circulating platelet number, platelets dysfunction (thrombocytopathy) has also been implicating as an additional factor contributing to lack of platelet functionality in canine monocytic ehrlichiosis.
In a case of Acute Leukemia, what type of cells would you see in the bone marrow?
Describe the time line of disease progression.
How old are the affected patients.
See below
In a case of Chronic Leukemia, what type of cells would you see in the bone marrow?
Describe the time line of disease progression.
How old are the affected patients?
See below
In a case of acute lymphocytic leukemia, what will you see on histology?
Undifferentiated/immature lymphocytes (lymphoblasts)
Cats: most were FeLV of FIV positive prior to routine vaccination and
testing
In a case of chronic lymphocytic leukemia, what will you see on histology?
How is immunophenotyping helpful? List examples
- Can identify lymphocyte subsets by the proteins they express on their
surface
Remember which markers are expressed by T cells and which one for B cells.
Describe the process of flow cytometry.
It is a biophysical based instrument used to study cell size, cell count, etc
The study is based on on principle of fluorescence analysis and can count thousand
of cells in seconds.
immunolabelling for antibody to the cell is used to characterized them, could be
inside or outside the cell. Can look at size, shape, density, DNA, RNA, protein content,
internal or external receptors, membrane surface, apoptosis, necrosis, calcium influx,
intracellular pH, etc.
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Interpret the data below:
- What does each quadrant represent?
- What % of cells is higher? B cells or T cells?
- Upper Left Quadrant = (-) for T cells, (+) for B cells
Upper Right Quadrant = (+) for T cells, (+) for B cells
Lower Left Quadrant = (-) for T cells, (-) for B cells
Lower Right Quadrant = (+) for T cells, (-) for B cells - T cells = 60.3% > B cells = 7%
