Cytomorphology of Human Bone Marrow Subjected to Prolonged Standing That Mimic Features of Myelodysplasia

Abstract

Examination of bone marrow (BM) aspirate smears plays an important role in the diagnosis of patients with various hematological disorders particularly patients with myelodysplastic syndromes (MDS). In current practice, bone marrow samples are often collected by the clinical staff and sent to the laboratory after a variable period of delay. As a result bone marrow aspirate smears prepared from such samples are prone to show morphological changes which may mimic changes observed in MDS. To ascertain the type and extent of morphological changes that may occur due to prolonged standing we have studied BM aspirate specimens from patients diagnosed with various hematological disorders which have been left standing for 30 minutes to 24 hours. Bone marrow aspirate samples were obtained from the right or left posterior iliac crest from ten patients (seven men and three women, age range 50-70 years) presenting with various hematological disorders who underwent BM aspiration and BM trephine biopsy procedures as part of their diagnostic work-up. Clinical indications included pancytopenia (n=3), leukopenia (n=2), thrombocytopenia (n=2), thrombocytosis (n=1), lymphocytosis (n=2). The first draw of 2.0 ml of BM aspirate sample was obtained using a conventional BM aspiration needle and placed in a tube containing appropriate concentrations of dipotassium ethylenediamine tetraacetic acid (EDTA). A drop of marrow was then placed on each of the three glass slides and smears were made. The subsequent smears (three slides from each patient) were made from the collected samples at intervals of 30 minutes, one hour, two hours, four hours, eight hours, 12 hours, 18 hours, and 24 hours. The slides were stained with Romanowsky stain and examined under a light microscope. Five hundred consecutive cells in each slide were assessed and the number of cells exhibiting dysplastic changes was recorded. Striking morphological changes were observed between the smears made of fresh marrow as compared to marrows left standing for a "lengthy" period. No noticeable changes in the morphology of hematopoietic cells were observed in the films made from the marrow which was left standing for not more than an hour at room temperature and were not easily distinguishable from the films made immediately after its collection (fresh marrow). By two hours changes were discernible and by 18-24 hours they became striking, affecting the morphology of all the hematopoietic cells. Myeloid cells were affected uniformly; their nuclei stained more homogeneously than in the fresh marrow, the nuclear lobes became separated and the cytoplasmic margins appeared ragged or less well defined. Small vacuoles appeared in the cytoplasm and they became more prominent and more numerous and the quantity of cytoplasmic granules decreased the longer the samples stood. Nuclear hypolobulation (pseudo-Pelger-Huet anomaly) and bizarre nuclear shapes were also observed. Erythroid precursors were also affected by prolonged standing which led to progressive degenerative changes such as nuclear lobulation and budding, nuclear-cytoplasmic asynchrony, karyorrhexis, cytoplasmic vacuolation, and cytoplasmic blebs. Megakaryocytes were also adversely affected and showed morphological changes in the form of multi-nuclearity and less granulation. Some of the lymphocytes also showed similar changes, including cytoplasmic vacuolation. Some of the nuclei showed major budding giving the appearance of a nucleus with two or three lobes (clover leaf shape). Examination of a BM aspirate smear is a highly informative hematological tool at the hematologist's disposal. However, the preparation of a good quality BM aspirate smear and the aberrations in the morphology of hematopoietic cells caused by a delay in smear preparation have not been reported. Our findings demonstrate that the morphology of hematopoietic cells in the BM aspirate deteriorates considerably during prolonged standing. Some of the changes that were noted were akin to the alterations seen in patients with MDS. We recommend that whenever possible the smears should be made immediately after the sample is withdrawn from the patient. This approach helps ensure the ideal morphology of hematopoietic cells and their accurate analysis for hematological diagnosis and may even prevent misdiagnosis of conditions such as myelodysplastic syndromes.