European Journal of Cardiovascular Medicine

Bone marrow examination plays a very crucial role in the diagnosis of hematologicaland non- hematological disorders.[1]The two important procedures used are bone marrow aspiration and Trephine biopsy.Both are invasive procedures. Aspiration alone is useful in patients with suspected nutritional deficiencies and acute leukemiasTrephine biopsy is essential for diagnosis when a 'dry tap 'or 'packed marrow 'occurs as a consequence of the marrow being very fibrotic or densely cellular. [2,3]Aspiration allows the assessment of cellular morphology and biopsy assessment is useful to know marrow architecture and arrangement of hematopoieticcells.[3,4] Bone marrow picture along with peripheral blood smear and clinicalfindings can help in arriving at a conclusive diagnosis.[5].

The aim of this present study is to study the clinical details and the bone marrow picture in various hematological disorders. We have included all the clinical cases which required bone marrow examination based on clinical details and peripheral smear findings. And all the participants who were willing to participate and who have given consent. We have excluded all the participants who were not willing to participate and who had not given the consent.

Clinical history was taken and detailed examination of all the patients was done. Base line hematological investigations such as complete blood count using 5Part analyzerYUMlZEN H500 (HORlBA) and MINDRAY BC-780, peripheral blood smear and biochemical tests were done as indicated. Peripheral smear was done using Leishman stain. Informed written consent was taken prior to the procedure. Bone marrow aspiration and biopsy (wherever necessary) were done under local anesthesia (Inj. 2% xylocaine) using all aseptic precautions. The most common site of aspiration is posterior superior iliac spine followed by sternum. Jamshedi needle was used for bone marrow aspiration and biopsy. Smears from the aspirate were checked macroscopically for presence of particles and stained with Leishman and Giemsa stains.

1-1.5 cm bone marrow core was taken on trephine biopsy.Trephine biopsy core was processed after decalcification using 5% EDTA in the automatic tissueprocessor and stained with Hematoxylin and Eosin stain. Perl's Prussian Blue stain for iron stores was done wherever necessary.Reticulin stain was done on marrow sections in case of Myelofibrosis.

The data acquired is categorized and analysed based on frequency and percentage. A total of 102 patients were included in this study, out of which 50 were males (49 %) and 52(51 %) were females. The patients age ranged from 04 -75 years.

Mean age of the patients is 40.75 years and maximum number of cases were seen between 31-40 years of age.

The most common indication for bone marrow examination based on peripheral smears is pancytopenia constituting about 38 cases (37.2%). Themajorclinical findingsin this studywere generalized weaknessfollowed bypallor.

Out of the total 102cases, 83 (81.3%) cases are of non-neoplastic, while 19 (18.7%) cases are of neoplastic nature. Among the non-neoplastic cases, nutritional deficiency anemias (55 cases) were the commonest disorders(Table1)Other non-neoplastic conditions were infections such asLeishmaniasis, Parvo virus, reactive plasmacytosis associated with Human Immunodeficiency Virus(HIV) and Immune Thrombocytopenic Purpura (ITP). (Table 1)Amongthe neoplasticcases, CMLis commonlydiagnosedfollowed byAML. (Table 2).

Table 1: Case distribution

Table 2:  Neoplastic cases distribution(n=19)

Among102 cases,wehavedone bonemarrow biopsyin 48cases. (table 3).The indications for doing bone marrow biopsy were for pattern of infiltration and staging of leukemias, phase determination in CML , dry tap in aspiration in case of myelofibrosis, marrow picture in multiple myeloma patients and for cases with packed marrow.

Table 3 :  Case distribution among 48 bone marrow biopsy cases

Bone marrow is the main site of hematopoiesis in adults. Bone marrow evaluation is a safe invasive procedure and plays an important role in identifying the etiology and in establishing a definitive diagnosis of hematological disorders.[⁶] They may vary from the reactive changes to hematological malignancies. Both aspiration and biopsy from bone marrow can be done with little discomfort to the patient, under adequate local anaesthesia

In this study the age group was from 4 to 75 years with mean age of 40.75 years. This result was in comparison with the studies ofKhan SP et al ^l ,Bashir N et al² Atla et a1 ³ Piplani et a1 ⁸In this study, males were 50 and female were 52. The male : female ratio was 0.9 : 1 which was compared with the other studies of Atla et a1 ³ Khan sp et alAtchyuta et a1 ⁵ ,Piplani et a1 ⁸

There was a slight female preponderance in the present study.

The commonest indication of BMA was pancytopenia. In this present study , 38 cases presented with pancytopenia.

The commonest cause of pancytopenia was megaloblastic anemia which was comparable with Bashir et a1 ² Atla et al^[3],Pipalani et al^[8], Knodke et a1¹¹and Gayatri et a1¹³. Erythroid hyperplasia is the predominant finding. Bone marrow aspiration smears of the megaloblastic anemia show megaloblasts with sieve like chromatin (Figure 1)

Figure 1: Aspirate smear shows megaloblasts with sieve like chromatin in a case of Megaloblastic anemia. (Leishman stain 400x)

In the present study there were 55 cases of nutritional deficiency anemia, among them therewere 35 cases of megaloblastic anemia, 8 cases of iron deficiency anemia and 12 cases of combined nutritional deficiency anemia.

In the present study there are 8 cases of pancytopenia which showed hypocellular marrow with decrease in all lineages and biopsy showed marrow replaced by fat.

In the present study ITP was seen in 4 cases (3.9 %) which was in concordance withKhan et al ⁴andAtchyuta et a1 ⁵On bone marrow examination there was relative increase in megakaryocytes showingreactivehyperplasia with markedvariabilityin size. Occasionalmicromegakaryocytes , few with hypolobated forms were seen.

In the present study hypersplenism was seen in 2% of cases which was inconcordance with Rajendra et al⁹ and more than Bashir et al²

We have encountered an interesting case in seven years old girl who presented as severe refractory anemia with hepato-splenomegaly which showed Gaucher cells (figure 2) with Crumpled tissue paper appearance on bone marrow aspiration and stained positive with PAS and Perl’s stain.

Figure 2: Marrow aspirate shows Gaucher cell with crumpled tissue paper appearance in a case of Gauchers Disease (Leishman stain,1000x)

The number of cases of Gaucher disorder is in concordance with Atla et a1³study.

A 38 year old male patient presented with fever, cough with expectoration, and had travel history. Peripheral smear examination showed pancytopenia. On bone marrow examination, amastigote forms of leishmaniasis were seen intracellularly and extracellularly with elevated plasma cells ( >10 %) (Figure 3). Leishmaniasis was also reported in the Khan sp et  and Parajuli et a1 ⁸(2.27%), Algazhyl et al ¹¹and Pudasaini.S et al ¹⁴

Figure 3: Aspirate smear shows LD bodies in a case of leishmaniasis (Leishman stain 1000x).

One case of parvovirus infection was seen in a patient with 35 years old male patient who presented with generalised weakness and pancytopenia. On bone marrow examination, giant proerythroblasts are seen with cytoplasmic projections like a dogear, known as dog ear proerythroblasts or giant proerythroblasts.(Figure-4) Panigrahi et al⁷ reported 1 parvovirus case in 29 cases of their study.

Figure 4:Marrow aspirate smear shows giant proerythroblasts with cytoplasmic projections in a case of Parvovirus infection (Leishman stain, 1000X)

In the present study, the most common neoplastic condition was chronic myeloid leukemia. There were 9 cases of CML. Bone marrow biopsy done and peripheral blood sent to FISH which showed BCR-ABL translocation. The number of CML cases in the present study are in concordance with the other , Atla et al³ Atchyuta et a1 ⁵and Kathik et al ^{l o}

There were 4 cases of AML, out of which two cases were APML. Bone marrow aspiration smears showed Faggot cells with Auer rods (figure 5) which were confirmed with FISHPML- RARA mutation. Both the cases were females presented with heavy per vaginal bleeding manifestations. They were diagnosed and started on treatment. The findings of the study are in comparison with Bashir et a1 ²Atchyuta et a1 ⁵

FIGURE 5: Marrow aspirate shows Faggot cells with Auer rods in a case of APML ( Leishman stain 1000x).

There were 3 cases of multiple myelomas, which showed rouleaux formation on peripheral smear, showed plasma cells on aspiration and biopsy (Figure 6) which were later confirmed by light chain restriction and Mband. In concordance with the studies of Khan et a1 ⁴ , Parajuli et a1 ⁶ and Khatik et al ^l0 and slightly lower than Atla et a1 ³ and Achyuta et a1 ⁵

Figure 6: Marrow aspirate showing plasma cells with eccentric nuclei and basophilic cytoplasm with perinuclear hof   in a case of Multiple Myeloma (Leishman stain 400x).

There was an interesting case in a 37 year old farmer who presented with severe anemia and weakness. Peripheralsmear study showed pancytopenia. Bonemarrow examination was reported as sideroblastic anemia (MDS/MPN) and Perls stain showed ringed sideroblasts (Figure 7)

Figure 7: Aspirate smear shows iron deposits (ringedsideroblasts) in a case of sideroblastic anemia (Perl's stain 400x)

There was one case of ALL in a 15 year old male child presented with weakness, anemia and thrombocytopenia, leucocytosis with lymphoblasts are seen on

peripheral smear. On bone marrow examination, marrow showedblasts .ALL had been reported. Later on cytogenetics it was confirmedas B- ALL.

In our study 8 cases of CML and 1 case of myelofibrosis showed dry tap on aspiration and later bone marrow biopsy was done.

Reticulin stain was done on trephine biopsy to know the extent of fibrosis. Figure 8 showed  increase in reticular network with coarse fibrils in a case of myelofibrosis.

Figure 8: Trephine biopsy smear shows increase in reticulin network with coarse fibrils in a case of Myelofibrosis (Reticulin stain 400X)

The dry tap or inadequate aspiration should always be accompanied by trephine biopsy to arrive at a finaldiagnosis. It might be due to packed marrow due to leukemias or metastatic deposits or marrow replacement due to fibrosis.Bashir et a1 ²and many other studies concluded this. This was also observed in present study.

There were three normal bone marrow cases which showed normal findings

Two bone marrow examinations showed inadequate marrow.

Bone marrow examination is of paramount importance in diagnosing neoplastic and non-neoplastic hematologicaldisorders. It also helps in defining the therapeutic approach, prognosis and response to thetherapy. The present study is useful to understand and correlate the clinical details andthe picture of bone marrow in hematological disorders.

Acknowledgement

I express my gratitude and sincere thanks to my guide Dr. Krishna kumari, M.D., Professor, Department of Pathology, NRI Institute of Medical Sciences, Visakhapatnam. For her valuable suggestions and unparalleled professional advice.

Conflict of Interest: There is no Conflict of Interest

Funding Source: There is no source of funding.

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