Week 4 Flashcards
in adults where is the hematopeotic tissue located
BM and lymph nodes, spleen , liver and thymus
lymphoid cells develop in two places primary and secondary lymphoid tissue
the primary lymphoid tissue is the BM and thymus where B and T lymphs develop is the 2ndary
BM - has precursor cells
What is the primary site of hematopoisis in adults
BM - medurally - central cavity with yellow and red marrow
Can take place outside the BM because of infection = Extra medullary non BM
Its not always pathological because it occurs during fetal stages
What are the fetal stages of hematopoiesis
Mesoblastic -
Hepatic
Medullary
What happens at the Mesoblastic -
stage
-in mesoderm of yolk sac
-production of primitive erythroblasts in forming blood vessels
-production of hgb
What happens at the hepatic stages
-5-7 weeks
-occurs extravascular in the liver
-fetal liver is main site and continues 1/2 weeks after birth
-peak of hematopoises by 3rd month and decline at 6 months
-production of RBC, WBC, megakaryocytes mostly in liver and spleen
-spleen( make b cells) , kidney,( make b cells) thymus (make t cells) and lymph nodes help
What happens at the medullary stage
-hematopoiesis starts in BM at 5 months
-at 24 weeks it is the primary site of hematopoiesis
-all cell lines at different stages of maturation seen
What types of changes would we see in the BM
Infants have all red marrow and in 5-7 years it is replaced by adipose tissue
-red is hematopoietic reactive marrow
with developing RBC and progenitors
-active marrow
By adulthood , all long bones are yellow marrow
-sternum, vertebrae, scapulae, pelvis, ribs, skull and proximal parts of long bones are the only ones that are active
-yellow can reactivate when responding to hemolysis or bleeding
-yellow is hematopoietic but made up of up adipose tissue
-inactive marrow
How is the marrow supplied with its nutriets and oxygen
nutrient artery which enters the bone and branches out into the sinuses
how is the red marrow arranged
in extravascular chords made up of hematopoietic cells that are found in between vascular sinuses and supported by trabeculae , stroma cells, extracellular proteins, collegen
red marrow is active
yellow is adipose tissue
What are stroma cells comprised of
Macrophages - phagocytosis, cytokine secretion
fibroblasts - extracellular matrix formation
osteoclasts - absorbs/ removes bone
osteoblasts - forms bone
endothelial cell- line the sinus and control passage of hematopoietic cells into circulation
Reticular cells - forms meshwork
stroma cells secrete fluid extracellular matrix
What does the extracellular matrix secreted by stroma cells do
- anchors developing hematopoietic cells in the bone cavity
-made up of reticular fiber, collagen , growth factors and cytokines that stimulate hematopoiesis
How are hemotopoeitic cords arranged
- grouped in islands
-islands and megakaryocytes develop close to sinuses
-while myeloid cells develop deep in the cords
What do erythropoietic islands contain
- maturing erythrocytes that surround macrophages containing iron
-macrophages form the island base so they are able to be an anchor for erythroid precursors to control their release into circulation via stromal cells
-macrophages release cytokines to help with red cell maturation and are the source of ferritin in hgb production
-the rest of the ferritin is provided by transferrin -the most
What controls erythropoiesis
Epo- hormone and stimulatory cytokine for erythropoiesis
-produced by peritubular cells in the kidney when you are in a state of hypoxia
-this hormone fluctuates to maintain homeostasis in O2
What causes the hypoxia that induces EPO production
Low RBC count via hemolysis or bleeding
-abnormal hgb
not related to red cells - lung function or high altitude
epo binds to receptor on RBC precursors that leads to cell division, maturation with more cells entering circulation
-BM stores more RBC precursors than needed in case there is increased need because you cant store mature RBC
-when these RBC are not needed they will undergo apoptosis and macrophages will remove dead cells
What is the action of EPO
Inhibition of apoptosis
-bind receptors on erythroid progenitor cells and promotes anti -apoptotic molecule production that turns off the apoptic signal in cell
Early release of retics
-reduces adhesion receptors so immature cells can leave the erythroid islands
Reduced maturation time
- decreased cell cycle time (decreases tiem spent in mitosis) ,
-increased cell processes “short boot camp; less time maturing in marrow”
increase in EPO = larger bluer cells in circulation because EPO forces cells into circulation before theyre ready = increased poly
What are BM studies used for
diagnose and stage hematologic and non hematologic neoplasms
- NOT for anemia when you can diagnose by looking at cells and indices
-multilineage abnormalities, blasts in adults and unexplained pancytopenia can prompt marrow exam
-prohibited in pt with coagulopathies like hemophilia or Vit K deficiency
-bridging with coumadin or heparin is needed to prevent uncontrolled bleeding
What secondary conditions can warrant a BM study
-solid tumors
-infections with FUO
-hereditary and acquired histiocytosis (abnormal increase in histocytes)
-unexplained organomegaly
aspirate vs core biopsy
types of BM specimen
Aspirate
apritation of semi fluid marrow
cells not in native state- can look at morph
core biopsy
-intact bone piece is removed
-cells are in native state
cannot be taken from all sites
common sites
-posterior superior iliac crest- common in adults and fat kids - asp and core
-sternum -adult , asp only
-anterior superior iliac crest- asp and core
-spinal processes - rarely used
children 0 upper end of tibia - asp only
What types of needles used in BM collection
Trephine - for circular piece of bone
like disposable Jamshidi needle
-Gauge is 11-15 bore size
-must be sterilized after use
-EDTA
-FORMALIN
What is the procedure for BM collection
Informed consent
-iodine is the antispetic
-local anesthetic - 2% lidocaine
-cut with scalpel after frozen
do not collect more than 1.5 ml
What is done with the aspirate collected
- put in a tube with EDTA or watch glass
-MLT looks for spicules or marrow tissue = grey bits floating in blood and fat droplets
-seperate spicules , pick up with pipette and put on slide ASAP with NO ANTICOAG
-2 types of smears
-push = like blood film
-squash = when you squash spicule between 2 slides and pull apart
label slides properly
-stain slides 4-6 with Wrights Geimsa
-keep a few that are unstained and unfixed for cytochemical staining
What is the role of the MLT during the BM collection process
making slides at bedside
-aspirate is collected in EDTA and brought back to the lab
-send the remaining for C/S, Molecular diagnostics, cytogenetics (heparin), flow cyto and histo if anything is left
How is the BM biopsy processed
- placed in B5 or 10% buffered formalin Zenkers or Carnoy
-processes by histo -core cut and stained by H&E and PPB
Advantages and disadvantages to bone marrow sampling
- Ad
represents marrow and bone structure in natural state
-demonstrates iron, reticulum and collagen
disadvantage
-fine cellular details lost in fixation and processing
-little value when trying to diagnose a leukemia
-dont use for differential
heme like asp and histo like biopsy
What does a complete BM examination include
Peripheral blood smear – CBC & complete morphological review done 24 hours before collection
Bone marrow aspirate
Bone marrow core biopsy
What are the problems that come with using an aspirate
DRY TAP
-correct site but sample cannot be aspirated can happen if marrow is too dense
-extreme hypercellular marrow
-fibrosis or marrow
need to take a BM core biopsy sample
-try and make slides from material around bone core = touch preps
What is a touch prep
-biopsy held by forceps and touched or rolled onto a slide BEFORE placing into formalin
-fixed and stained like BM aspirate
-youll be able to see enough cells to assess maturation stages and types or do a differential
When you look at BM what do you look for
Cellularity
Maturation of myeloid & erythroid series
M:E ratio
Cell distribution (Differential)
Eosinophils/basophils
Megakaryocytes
Presence of other cells
Lymphocytes
Plasma cells
Histiocytes
Osteoclasts
Fibroblasts
Iron stores
Abnormal infiltrates
Fibrosis
What to look for when evaluating a BM ASPIRATE
Relative cellularity - core is more accurate for cellularity
Differential and M:E
Iron stores
Megakaryocyte Evaluation
Metastatic tumor cells / Lymphoid aggregates- best on aspirate
start on low power 10x to detect spicules -hematopoietic and stromal cells stained dark blue / purple
-look for an intact area behind a spicule
Assess general cellularity-all you can say is HYPO, HYPER or NORMAL cellular
Abnormal cell clusters- seen on edges
Quality of aspirate using Megakaryocytes 2-10 if more or less then pt condition
What is the next step once you have found an intact area behind a spicule
- aspirate differential
-done under 100x
-count 300-500 nucleated cells to determine M:E ratio
All nucleated cells:
Myeloid series – Blasts – Segs: Neut, Eos, Baso
Erythroid series
Lymphs, Monos, Plasma cells – early cells as well
in a normal BM a normal BM consists of myeloid and erythroids , all stages should be present but within their associated distribution
-count and differentiate all
What is your myeloid to erythroid ration
-derived from diff
-normal ration with 1.5:1 - 3.3:1
-granulopoietic tissue occupies 1.5 to 3.3 times more space than erythroid precursors due to shorter survival of granulocytes in circulation
-so changes in granulocytes and erytrocytes are reflected in M:E ratio
-only take myeloid and erythroid cell #s from diff - exclude lymphs, monos, plasma
how to evaluate iron stores in the bone marrow
use perls prussion blue
-iron in BM is stored as intracellular hemosiderin in macrophages and in small amounts of normally developing erythroblasts
-hemoglobin iron does not stain
-positive iron is seen as blue deposits
-aspirate is favored over biopsy for iron stain because when you process biopsy it tends to leach iron which can give you a false decrease or negative iron
-enough spicules should be present because you need to know if the low iron is due to contamination or the pt lacks iron stores
report iron as
Absent
Decreased
Adequate
Moderately or
Markedly Increased
how is the megakaryocyte evaluation doe
- scan the number on low power 10x
-2-10 per lower power field
-too unevenly distributed to count on diff so they are reported as increased or decreased
-clusters can indicate hyperplasia
-< 2 megakaryocytes / LPF (low power field) may indicate hypoplasia or poor quality aspirate
-no spicules and no mega = low quality aspirate
how to assess BM biopsy sample
look at cellularity
The ratio of all nucleated hematopoietic cells (ANC) to fat cells
cellularity in adults is approximately 50% ( 1:1)
General rule to estimate normal cellularity for age: 100-age +/-10.
Reported as Increased, Normal or Decreased
so if youre 75 years old your cellularity is 15-35%
A normal adult bone marrow displays 50% hematopoietic tissue and 50% fat.
NORMAL CELLULARITY
What does a BM report contain
Patient history, age, sex, etc.
Description of samples received
Peripheral blood data: CBC, ESR, Reticulocyte count, Manual differential, PBS
Bone marrow differential count
Description of the cellularity, M:E ratio
Status of the iron stores
Description of histological sections of biopsy
Diagnostic conclusion
What can cause BM failure
-reduction of cessation of blood cell production that affects 1 or more cell lines
-no typical morph- just decrease in all lines : pancytopenia
-Aplastic Anemia is a general BM failure syndrome
pancytopenia, reticulocytopenia, bone marrow hypocellularity, low hetamo stem cells
Destruction of HSC
Premature senescence and apoptosis of HSC
Ineffective hematopoiesis
Disruption of BM microenvironment
Decreased production of growth factors or hormones
Loss of normal hematopoietic tissue due to infiltration of marrow by abnormal cells
