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Normal histology Hematolymphoid tissues

Basic Hematopathology Course,


TMH, J une12-13, 2010
Dr. Sumeet Gujral,
Associate Professor,
Department of Pathology
Tata Memorial Hospital, Mumbai
s_gujral@hotmail.com
Hematolymphoid tissues
Peripheral blood
Bone marrow
Lymph node
Spleen
Thymus
Waldeyers ring
Elsewhere
Peripheral Blood Smear
Peripheral blood
Cells
RBCs,
Platelets
WBCs
Plasma: whole blood minus
cells
Serum: whole blood minus cells and the clotting factors
Steps for preparation of smears
Finger prick, fresh blood with no anticoagulant added
EDTA - anticoagulated blood: Film should be made within
2-4 hours (storage artifacts)
Heparinized blood to be avoided
Approach to peripheral blood
smear examination
To evaluate the quality, approximate number of WBCs
and platelets
- WBC count in cells/ml on PBS is low power x 3000,
- Platelet counts in cells/ml on PBS is oil immersion x 20,000
Detect rouleaux formation, platelet clumps, and leukocyte
clumps and other abnormalities.
Select an optimal area for evaluation at higher power
Low power (10X)
Do at least 200 WBC count, and record any abnormal
morphology of RBCs, WBCs, and platelets
Look for parasites
Oil immersion
Purplish pink
Light pale pink
Greyish pink
Purplish blue
Pale blue
Sky blue
Chromatin
Purplish blue
Sky blue
Pale blue
Granular cells
Round cells
Hypersegmented
polymorph
??
??
Downey
cells
?
Poor quality smears
Delayed staining
Stain deposits
Drying artefacts
Platelet clumps
PBS as part of the Medical Record
Preserve and store
Indian J Pathol Microbiol. 2010 J an-Mar;53(1):68-74
Importance of PBS examination
Bone marrow preparations
aspirate
touch
trephine
clot
Normal bone marrow
Bone Marrow Aspirate
Myeloid precursors
2
3
1
4
5
1
2
Promyelocyte
Neutrophil
Metamyelocyte
Myelocyte
Promyelocyte may be larger than a blast and cytoplasm contains
large black or purple granules. Nucleoli may be present
Promyelocyte
Blast
Promyelocyte may be larger than a blast and cytoplasm contains
large black or purple granules. Nucleoli may be present
Myelocyte
Metamyelocyte
Promyelocyte
Promyelocyte may be larger than a blast and cytoplasm contains
large black or purple granules. Nucleoli may be present
Monocytic precursors
Promyelocyte
Promonocyte
Lymphoid precursors
Lymphoblasts
Lymphocytes
Hematogones
Hematogones
Erythroid precursors
Erythroblasts
Megaloblast
Colony
??
Platelet precursors
Platelet precursors
Platelet clumps -
pseudothrombocytopenia
Anand M et al
Indian J Pathol Microbiol. 2005 J ul;48(3):425-6
??
Normal
cells
Dyspoietic
cells
Normal
Dyspoietic
Normal
Dyspoietic
Normal
Dyspoietic
Blasts
Acute Leukemia
>20% blasts in peripheral blood or bone marrow
What are blasts?
Morphology
Exceptions
Small size
Granular blasts
Abnormal promyelocytes in AMLM3
Promonocytes in AMLM5
Guess
Guess
Clusters in
bone marrow
Bone Marrow Biopsy
Indications of BM Biopsy
PUO
Storage diseases
Aplastic anemia
Dry tap
Myelofibrosis
Myelodysplastic syndrome
Staging of lymphomas
Acute leukemia
Procedure and processing
Trephine (Hammersmith Protocol)
Fixative (AZF)
Decalcifying agent (10% FA and 5% formaldehyde)
2-3 micron thick section
Immunohistochemistry
Adequate biopsy
Both aspirate and imprints with the biopsy (>1.6 cm)
Ideally, reporting of trephine biopsy sections should be done by an
individual who is competent in both histopathology and hematology
Cellularity
Cells: Fat cells, Hematopoietic cells (trilineage
hematopoiesis), Megakaryocytes, Blasts, Others
Fibrosis, granulomas, tumor
Low power examination
Cellularity
Aspirate and Biopsy
are complementary
Bone marrow in a 40-year-old
Types of cells
Types of cells
Regenerating bone marrow
Myeloid ++++
Routine sections
Hemorrhagic bone marrow biopsy
Pediatric bone marrow biopsy
Adequate bone marrow biopsy
Large subcortical area not truly representing
overall hematopoietic activity
Fragmanted BM biopsy
ALCL
Crushing artefacts - FL
IHC may be
useful
Follicular lymphoma
Mantle cell lymphoma
IHC may be
useful
Good trephine - J oint responsibility
Physician doing the biopsy (anesthetist)
OT Nurse
Technologist
Pathologist
Administrators
Vendors
Patients
BM and lymphomas
Staging marrows
Diagnostic marrows
Different patterns in lymphomas
Patterns
Diffuse
Interstitial
Nodular
Patchy
Intrasinusoidal
Paratrabecular
Focal non paratrabecular
Focal paratrabecular
Intrasinusoidal
Diffuse, interstitial
Diffuse, solid
Focal paratrabecular
Diffuse pattern
ALL
Diffuse pattern
CLL
Interstitial
- Exclusively: BL, LL, HCL
- Combined focal and interstitial: SLL, LPL, MCL, ALCL
DLBCL patch
SLL Mixed
Nodule + Interstitial
Nodular
All MZL SLL, FL, HD
CD20
Nodule
Nodule
CD138
Hodgkins disease
Nodular, diffuse, patchy
Biopsy and aspirate are complementary
CHD - Nodule
CD3
PTCL NOS, Patch / nodule
ALCL - intrasinusoidal
Blastic lymphoma
versus
Burkitts lymphoma
Acute Leukemia
Do we need bone marrow biopsy ??
Burkitts lymphoma
ALL
ALL
CD34
Tdt Mic2
AML Non M3 AML M3
B-cell ALL
Granulomas in
HD
Reactive lymphoid proliferations in
bone marrow
1. Lymphoid aggregates
2. Hematogones
Benign Lymphoid Aggregates
1. Distribution - Usually perivascular, intertrabecular
2. Number/size - Few in number, small in size
3. Circumscription - Well circumscribed (except in AIDS)
4. Cell composition - Mature looking cells, Heterogeneous cell population
consisting of small to large sized lymphocytes, plasma cells, histiocytes
5. Germinal centres - seen in drug related or in autoimmune diseases
6. IHC - T cells predominate
Benign Lymphoid
Aggregates
Young age collagen vascular diseases
Old age
CD3
Hematogones
medium sized lymphoid cells,
scant cytoplasm,
round to irregular nuclei,
dense homogeneous chromatin
no - very small nucleoli
Parasites (in PB/BM)
MP with
satellitism
Exflagellated microgametes of Plasmodium vivax
Tembhare P et al.
Indian J Pathol Microbiol, 2009
Microfilaria
Parvovirus - BM
Parvovirus - BM
Lymph nodes
Lymph node is a dynamic structure
Primary follicles
IgM+IgD+
Secondary follicles, IgD-
Mantle zone
IgM+IgD+
T
T
T
T
T
Differentiation of B cells during their passage
through the germinal center
Secondary
B blasts
Fdc
macrophages
CC
Fdc
macrophages
CB
Fdc,
macrophages
Primary B blasts
Plasma cells Memory B cells
Mantle zone
IgM+IgD+
T zone proliferation
Identify
Identify
CD3
CD20
IHCs in a normal node
CD23
Mib1
bcl2
Various cells
Immunoblasts
Centroblasts
Plasma cells
Centrocytes
Common causes of lymphadenopathy
Infections
Malignancies
Warning signs of lymphadenopathy
suggestive of a malignant etiology include
- size >2 cm in size,
- duration >2 month,
- location - supraclavicular, and
- generalized lymphadenopathy with
hepatosplenomegaly or B-symptoms.
VIP Syndrome
Benign lymphadenopathy
Infections
viral (EBV, HIV, CMV), bacterial, parasitic,
Autoimmune disorders, Drug hypersensitivity reactions,
Kikuchis disease, Castlemans disease, SHML,
Kimuras disease, PTGC, Toxoplasmosis
Dermatopathic lymphadenitis
LN: Other patterns
(other than granulomas)
PTGC RTGC
Wierd looking
nodules
Follicular hyperplasia
versus
Follicular lymphoma
Which one is a lymphoma?
FL Grade 1 / MCL Follicular hyperplasia
FL Grade 1 / MCL Follicular hyperplasia
bcl2
FL (Grade 2) Follicular hyperplasia
FL (Grade 3) Reactive
Mimic
Gold standard
Avoid FNAC / FS
diagnosis of lymphomas
Spleen - Organ of Mystery
Spleen
2 x 1.5 x 0.2 cm, immediate processing (may /may not wait for fixation)
Congested
cords
Sinus
Red pulp
Trabeculae
PALS
SMZL
Others
Thymus
CD3
Tdt
Thymus
Tonsil
Tonsil: Plasma cell rich lesion
Conclude
A. Myeloid neoplasms
B. Precursor lymphoid neoplasms
C. Mature B cell neoplasms
D. Mature T- and NK- cell neoplasms
E. Hodgkin lymphoma
F. Immunodeficiency associated LPD
G. Histiocytic and dendritic cell neoplasms
So many subtypes,
Different treatment options
2008 WHO classification of Hematolymphoid
Neoplasms
Optimal tissue fixation, processing followed by a thin,
well stained (Haematoxylin and Eosin) section is most
important for lymphoma diagnosis.
Lack of trained hematopathologists, inadequate
sampling of the tissue and improper processing of the
specimen
Second opinion and multidisciplinary clinic
NHL, follicular type,
grade 1
Staff, Residents and Colleagues at Hematopathology
Laboratory and Department of Pathology, TMH
s_gujral@hotmail.com

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