Bone marrow trephine biopsy as My Short experiences

Professor Pranab kumar Bhattacharya MD(cal) FIC Path(Ind.)

Professor & Head of Pathology, In charge & convener DCP &DLT course West Bengal University of Health sciences, School of Tropical Medicine 108 CR avenue Kolkata-20, -West Bengal

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Bone marrow aspiration – Normal Cellular Smear

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Marrow beats

WHY YOU FAILED TO ASPIRATE?

• No marrow fragments
• When there is Fibrosis as in Myelofibrosis in CML
• When there is Aplasia- Aplastic or Hypoplastic Anemia
• Cellular marrow BUt
• Poor aspiration – packed cells in the marrow in Leukemia even
• Focal distribution
• When there is Pathology in bone itself

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CARRY OUT A TREPHINE BIOPSY OF

BONE MARROW

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BONE MARROW BIOPSY

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• Islam & Jamshidi – 1970s
• Advances in Bx needles,
• specimen processing like Tissue processing and HE with Special Stains,
• IHC, molecular diagnostic techniques

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NEEDLES

• Jamshidi
• Islam
• Westermann-Jensen
• Drills
• Disposable needles

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SITES OF BM BIOPSY

• Posterior superior iliac spine-commonest I Prefer
• Anterior superior iliac spine
• Iliac crest –I Prefer it also
• Tibial tuberosity-in child
• Under fluoroscopic guidance – Spine

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BONE MARROW IMPRINT

• Please Note
• Necessary to carry out an aspiration simultaneously

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• If aspiration fails, make imprint smears- It will gives clue to diagnosis

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Make an imprint

PROCESSING BM Bx

• FIXATION

10% Buffered formalin

Zenker’s fluid

• DECALCIFICATION

Formic Acid – 6-8 hrs

Nitric acid – 2-3 hrs

EDTA

• EMBEDDING

Paraffin

Resin

• STAINS

H&E, Reticulin, Masson’s, Perls

• MICROWAVE FIXATION_ IN IHC

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YOUR EVALUATION BASED

• Must be accompanied with-
• Clinical details; Clinical History

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• You must Look at CBC and PBF findings- what it suggest?

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• Aspiration report should be available

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BEFORE INTERPRETATION…

• Look at ADEQUACY of Bx
• Length – 1.6 cms (1.5 – 2cms) – 25% shrinkage occurs during processing

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• 5 – 6 trabecular spaces
• Good quality staining

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How a Bone marrow Biopsy Looks after processing & HE stain

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Look at trabeculie of Bone marrow materials

WHILE INTERPRETATION….

LOOK CELLULARITY BUT REMEMBER

• Decreases with increase in age
• Infants 80%, Adults 50-60%, Old age 30-40%
• Is there Subcortical hypocellularity

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Bone marrow biopsy of Hypoplastic anemia

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Cellular good but fat space normal and only two/three megakaryocyte in total BX- Normal

Megakyocyte

Myeloid group cells

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Fat Space increased through out

STRUCTURE OF NORMAL MARROW

• Myeloid cells
• Paratrabecular
• Mature cells towards centre
• Erythroid cells
• Centre in colonies
• Megakaryocytes
• Centre around sinusoids
• Stroma
• Fat cells, fibroblasts, reticulin fibres

EVALUATION

• Cellularity
• Erythropoiesis
• Myelopoiesis
• Megakaryopoiesis
• Plasma cells
• Lymphocytes
• Macrophages

• Reticulin fibres
• Iron stain
• Necrosis
• Others
• Granuloma
• Metastasis
• Bony trabeculae

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FIBROSIS (Grading – Bauermeister 1971)

• Grade 0 : No Reticulin fibres demonstrable
• Grade 1 : Occ. fine fibril; foci of fine fibre network
• Grade 2 : Fine fibre network throughout
• Grade 3 : Diffuse fibre network with some thick coarse fibrils
• Grade 4 : Diffuse coarse fibre network with collagenization

INDICATIONS

• Inadequate or failed aspirate
• Need for accurate assessment of cellularity
• Suspected focal lesion
• Suspected bone marrow fibrosis
• Need to study bone marrow architecture
• Need to study bone structure or marrow blood vessels

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INDICATIONS

• Aplastic anemia
• MF/OMS
• Hodgkin’s
• NHL
• Hairy cell leukemia
• CML, other MPD
• MDS
• Multiple myeloma

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• Metastasis
• Granulomatous lesions – PUO
• HIV
• Acute leukemia
• Bone marrow transplant
• Osteopathies

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BIOPSY OR ASPIRATE ?

• Dry tap
• Large volume of tissue
• More reliable index of cellularity
• Architectural pattern
• Information on connective tissue
• Less chance of missing a focal lesion
• Post chemotherapy

BM Bx & BMA ARE ALWAYS COMPLIMENTARY TO EACH OTHER

WHAT ARE MARROW FAILURE DISORDERS?

• Aplastic anemia
• Fanconi
• PNH
• Hypoplastic MDS
• Hypoplastic Acute leukemia

APLASTIC ANEMIA

• >75% mature fat
• Residual hematopoietic cells normal
• Plasma cells, lymphocytes, reticulum cells
• Iron stores – normal/ increased

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Mature Fat cells

Hypoplastic But Blast cells

Hypoplastic But Blast cells

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MYELOPROLIFERATIVE DISORDER

• Is there neoplastic hyperplasia?
• What is the predominant cell line (s) involved?
• What is the degree of differentiation of the cells involved?
• Is there fibrosis?

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CHRONIC MYELOID LEUKEMIA

• Not to diagnose by BMA or BX
• But for Subtype CML
• To Assess marrow fibrosis
• To detect early blast crisis

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CML

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CML Myelocyte, Meta myelocye Immature Granulocyes, Erythriod cells

High Power View

MYELOFIBROSIS

• Agnogenic myeloid metaplasia with splenomegaly
• Megakaryocytic hyperplasia - PDGF, TGF
• Fibrosis is secondary and reactive
• Three phases
• Cellular
• Fibrotic
• Osteosclerotic

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MARROW FIBROSIS

• GENERALISED

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• IMF
• CML
• HCL
• AML M7
• MASTOCYTOSIS
• MM WITH FIBROSIS

• FOCAL

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• HODGKIN’S
• NHL
• GRANULOMAS
• METASTASIS
• SLE
• HIV

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HODGKIN’S DISEASE Bx DONE For….

• Staging
• BM involvement in West Bengal< 10% ( In India 2-32%)
• Mixed cellularity & Lymphocytic depleted
• Incidental diagnosis
• Aspiration yield is poor Hodgkin or RS cells

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CRITERIA FOR BM INVOLVEMENT

• RS cell demonstration in previous Bx
• Typical RS cell in HD background
• Mononuclear variants - atypical histiocytes in HD background
• Fibrosis & necrosis – hence not aspirable

RS Cells

RS Cells

Hodgkin Cell

NON-HODGKIN’S LYMPHOMA

• Staging
• Pattern
• Nodular, interstitial, mixed, diffuse
• Paratrabecular – follicular NHL
• Pretherapy marrow reserve
• Diagnostic, immunophenotype
• Recognition of discordance

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Mixed cell type NHL

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HAIRY CELL LEUKEMIA

• Diagnostic – dry tap
• Pancytopenia, splenomegaly, old age
• Diffuse/ focal/ interstitial
• Fried egg appearance
• Web-like reticulin meshwork

CHRONIC LYMPHOCYTIC LEUKEMIA

• Diagnosis (International CLL workshop)
• Prognosis
• Interstial / Nodular – Better prognosis
• Increased fibrosis – Bad prognosis
• Increased plasma cells – Bad prognosis
• Hematopoietic elements

ACUTE LEUKEMIA BX to be Done When

• PS, BMA, immunophenotype
• BMA dry/ inadequate
• Hypoplastic acute leukemia
• AML-M7
• Myelonecrosis
• Follow up (post Rx yield)

Low Power View Note Highly dense Cellularity

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Myeloblasts

MYELODYSPLASTIC SYNDROME

• Cellularity
• Topography – ALIP, paratrabecular megakaryocyte
• Dyspoiesis well assessed
• Blast increased
• Assess fibrosis - >30% increased

AML

PROBLEMS WITH MDS I Faced

• When it is Hypocellular MDS:

D/D Aplastic anemia?

When No dyspoiesis

Chromosomal abns Study.

• When MDS associated with fibrosis:

Myelofibrosis

CML- acclerated. phase

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MULTIPLE MYELOMA

• Early MM, smouldering MM
• Focal clustering
• Histological classification
• Pattern – Packed marrow
• Amyloid deposits

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GRANULOMAs are found in Bone marrow Biopsies Well in…

• Bacterial
• TB
• Typhoid
• Leprosy
• Brucellosis
• Syphilis
• Viral
• EBV
• HIV

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• Rickettsial
• Q fever
• Fungal
• Cryptococcus
• Histoplasma
• Parasitic
• Leishmania
• Toxoplasma
• Sarcoidosis
• Metastasis
• Lymphoma
• Carcinoma

Well circumscribed Granuloma with casiation

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Epetheloid cells Seen IN HIV Pt

Langhen giant cell

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What are these?

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Globi Filled with AFB in Leprosy

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What are these?

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Fungus PAS Positive

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Hyphe Fungi

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Hype Aspergillus's fungus in BM Biopsy

Gaucher’s Cell

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Gaucher Cells

METASTASIS

• PURPOSE OF BM
• For staging of documented malignancy
• To evaluate occult malignancy
• Confirm suspected cases

PRIMARY SITES

ADULTS

• Breast
• Prostate
• Lung
• Stomach
• Colon
• Kidney
• Thyroid

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CHILDREN

• Neuroblastoma
• Rhabdomyosarcoma
• Retinoblastoma
• Ewing sarcoma
• PNET

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Metastatic Deposit from Breast ca

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MICROMETASTASIS

• Secondaries < 0.3 sq cm or 10⁸ cells

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• Significance??
• Predicts distant metastasis and early relapse
• Autologous BMT
• Use of ancillary tests

NECROSIS

• Necrosis of myeloid tissue and stroma with bone infarction
• Bone pain
• Fever
• Pancytopenia
• Leucoerythroblastic reaction
• LDH and Alk Phos increased

NECROSIS

• Acute leukemia – pre and post therapy
• Lymphoma
• Metastasis
• Sickle cell disease, SLE, TB, HIV

GELATINOUS TRANSFORMATION

• Acellular myxoid material
• Acid mucopolysaccharide
• Compensatory response to mobilisation of fat
• Reversible

CAUSES

• Malnutrition – Anorexia nervosa
• Starvation
• Post chemotherapy
• Acute leukemia
• AIDS

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Geletenious Transformation in a post chemotherapy Pt

ROLE OF SPECIAL STAINS

• Giemsa
• Reticulin
• Masson’s trichrome
• Perls
• PAS, ZN, Mucicarmine

ROLE OF IHC

• Acute leukemia when aspirate is unsatisfactory / MRD
• Metastasis – to find out origin
• Lymphoid nodules (benign vs malignant)
• Early myeloma

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IRON STAIN

• Usually removed during processing
• Positive or negative report, do not say iron stores are diminished
• Zenker’s fixative
• 16 to 18 hrs
• Wash for 6 hours
• Perls stain 80^o C for 5 mins

A WORD OF CAUTION When to attempt BX…

• If Pt has Coagulation disorders, DIC

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• Liver failure and Increased PT

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• Thrombocytopenia < 15,000/cmm

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• Hemorrhage

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SUMMARY

• Investigation of suspected HD & NHL
• Staging of HD and NHL
• Diagnosis of Aplastic anemia, Hypoplastic MDS, Hypoplastic AML
• Diagnosis & follow up of HCL
• Evaluation & follow up of CLL

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SUMMARY …

• Diagnosis of metastatic Ca
• Diagnosis, staging, & follow up of small cell tumors of childhood
• Investigation of suspected myeloproliferative disorders
• Evaluation of CML
• Investigation of an unexplained leucoerythroblastic blood picture
• Investigation of PUO

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SUMMARY…

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• Investigation of suspected AML
• Investigation of suspected MDS
• Investigation of suspected MM
• Investigation of suspected Amyloidosis
• Evaluation of patients in whom an adequate BMA cannot be obtained

I’m grateful to late Professor Rathin Roy MD(cal)FISH, Ex- Professor Director Medicine, Calcutta Medical college, under whom I was first trained in hematology during 1981-83 and had preliminary knowledge in looking abnormal cells in BM aspiration and very rarely BX.

* To my MD thesis guide Doctoral teachers & professor Prof. Subir kumar Dutta- Ex National president IAPM for my teaching, �*To professor K. Mukherjee MD(BHU) who extended me the real scope to study Trephine biopsies & [ me in charge of Histology] of patients attending Hematology clinic of IPGME&R, kolkata-20; W.B India

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A Genetist- not a hematologist

• I am no more a hematologist, or a histo-pathologist- many pathologist /clinician/surgeon of state or country refers me cases.
• I work now on human Gene- Cloning, DNA Micro array signal Trasductions- since 2002 at free of cost,- my choice.
• A Recognized Lab by MCI at IPGMER