Prognostic significance of the histomorphometric features of bone marrow trephine biopsies in patients with chronic myeloid leukemia

OBJECTIVE: To study the correlation of histomorphometric data of bone marrow trephine biopsies at the time of initial diagnosis in chronic myeloid leukemia (CML) patients with the patient prognosis. STUDY DESIGN: A total of 40 CML patients were divided equally in group I (developed accelerated phase or blast crisis within 18 months of initial diagnosis of chronic phase of CML) and group II (developed accelerated phase or blast crisis > 30 months after initial diagnosis of chronic phase of CML). The clinical, hematologic and histomorphometric data were compared in the 2 groups of CML patients. RESULTS: The percentage of bone marrow promyelocytes was significantly increased in group I. On morphometry, the number of blasts per square millimeter, the area of reticulin fibers per square millimeter and the percentage area occupied by reticulin fibers were statistically significant. CONCLUSION: There seems to be grounds for hope for predicting prognosis of CML patients at initial diagnosis based on histomorphometric findings. The percentage area of reticulin fibers and the number of blasts per square millimeter are important prognostic predictors in histomorphometry data.     

Megakaryocyte precursors (pro- and megakaryoblasts) in bone marrow tissue from patients with reactive thrombocytosis, polycythemia vera and primary (essential) thrombocythemia. An immunomorphometric study

To determine the number of megakaryocyte precursors (pro- and megakaryoblasts), an immunomorphometric study was performed on paraffin-embedded trephine biopsies of the bone marrow using a monoclonal antibody against platelet glycoprotein IIIa. Eighteen control specimens from patients with no evidence of any hematological disorder and a normal platelet count were selected and assessed together with the same number of specimens from patients with reactive thrombocytosis, polycythemia vera rubra (P. vera) or primary (essential) thrombocythemia (PTH). A strikingly proportionate increase in early megakaryocytes occurred in all patients enrolled in this study, compared with the controls. Moreover, there were no significant correlations between counts for precursors or total megakaryocytes per square millimeter of bone marrow with the corresponding values for platelets. This indicates that despite an orderly increase in immature forms in the bone marrow, the number of platelets circulating in the blood is influenced by other additional factors, such as the expanded platelet pool in the enlarged spleen. The non-disproportionate expansion of megakaryocyte precursors extends previous findings on progenitor cells of this lineage in vitro, particularly in PTH. Histological evaluation of the bone marrow of patients with P. vera and PTH indicated that megakaryopoiesis proceeded to the production of appropriate mature forms with no obvious excess of very small or blastic elements.     

A histomorphometric analysis of trephine biopsies of bone marrow from 65 patients with chronic myeloid leukemia. Classification of patients into subgroups with different survival patterns

A histomorphometric (planimetric) study was performed on trephine biopsies of the bone marrow taken at presentation from 65 patients (31 males and 34 females, with a median age of 48 years) with chronic myeloid leukemia (CML). Specimens from 20 patients (9 males and 11 females, with a median age of 53 years) without any hematologic disorders served as controls. Of the various histologic variables tested, only the counts of neutrophilic granulocytes per 1 sq mm, the ratio of granulocytopoiesis to megakaryopoiesis and the density of reticulin (argyrophilic) fibers revealed a significant correlation with the prognosis. The CML patients were separated into two groups with different survival patterns. Group I (34 patients with a median survival of 24 months) mostly contained cases with the so-called "megakaryocytic subtype" of CML, which is accompanied by variable degrees of fibrosis; group II (31 patients with a median survival of 36 months) mainly contained cases with the "granulocytic subtype," which is not accompanied by myelofibrosis. Among the morphometric parameters, a positive correlation existed between the megakaryocyte count and the reticulin fiber density, which underlines the important role of that cell lineage in fibrillogenesis. There were multiple interrelationships between the histomorphometric variables and the laboratory data. Consequently, multivariate regression methods (using Cox's proportional hazards model) were applied to assess the relative predictive value of the patient characteristics for survival. The derived prognostic model divided the patients into two risk groups, with median survivals of 14 and 41 months, respectively. In order of their entry into the regression model, these variables were percentage of neutrophils in the differential blood count, amount of granulopoiesis, liver size, percentage of peripheral myeloblasts and density of reticulin fibers in the bone marrow. In comparing the two patient groups, based on bone marrow histomorphometric parameters, this model revealed that two of those factors (amount of granulopoiesis and density of reticulin fibers) had a significant correlation with the prognosis.     

Conversion of polycythemia vera to refractory anemia with hyperplastic bone marrow.

Clinical and morphologic findings in the conversion of treated polycythemia vera to pancytopenia with hyperplastic bone marrow (refractory anemia or pancytopenia with hyperplastic bone marrow) are described in light of our own observation. The nomenclature associated with this condition (pancytopenia, chronic erythroleukemia, preleukemia) is not uniform, whereas the morphologic findings are virtually identical. Some patients subsequently develop acute leukemia. The prognosis in cases of refractory anemia with hyperplastic bone marrow following polycythemia vera is, independent of the subsequent acute leukemia, invariably terminal.     

Evaluation of the JAK2-V617F gene mutation in Turkish patients with essential thrombocythemia and polycythemia vera

An activating mutation of Janus kinase 2 (JAK2-V617F) was previously described in chronic myeloproliferative disorders (MPD). In previously published studies, the frequency of the JAK2-V617F mutation was determined to be 80-90 % for patients with polycythemia vera (PV) and 40-70 % for essential thrombocythemia (ET). In this study, we analyzed the relationship between the JAK2-V617F mutation and clinical-hematological parameters in Turkish patients with MPD and compared these findings with published studies from other geographic regions. A total of 148 patients were studied; of which, 70 were diagnosed with PV and 78 with ET. The mutation status of JAK2 was determined using a tetra-primer polymerase chain reaction. We found that 80 % of the PV group and 42 % of the ET group were positive for the JAK2-V617F mutation. When all patients were analyzed, the levels of white blood cells, hemoglobin and splenomegaly were significantly different in patients with the JAK2-V617F mutation (p < 0.05). To our knowledge, this study is the first to evaluate the relationship between MPD and JAK2-V617F in Turkish patients. The JAK2-V617F mutation is frequently detected in the Turkish patients with MPD, and especially in patients with PV. Hence, it would be useful to include JAK2 mutation screening in the initial evaluation of patients suspected to have MPD.     

Megakaryocyte precursors (promegakaryoblasts and megakaryoblasts) in the normal human bone marrow. An immunohistochemical and morphometric study on routinely processed trephine biopsies

On routinely processed trephine biopsies of the normal human bone marrow derived from 15 patients, immunostaining with a monoclonal antibody against platelet glycoprotein IIIa (Y2/51) and morphometric measurements were performed for the determination of megakaryocyte precursor cells. Based on cell sizes and on comparison with (1) specimens stained by the periodic acid-Schiff reaction and (2) smears, the smallest elements clearly identifiable as belonging to the megakaryocyte series were classified as promegakaryoblasts. Promegakaryoblasts had a frequency of 1.7/sq mm and 140/cu mm of bone marrow and constituted about 8% of the total positively stained megakaryocytic elements; they were characterized by a size of 41.5 sq microns, a diameter of 7.7 microns, a high nuclear-cytoplasmic ratio (0.32) and a nearly circular outline of their nuclear and cellular perimeters.     

RNA-dependent DNA polymerase from a cell line derived from the bone marrow of a patient with polycythemia vera.

A RNA-dependent DNA polymerase was isolated from a human cell line derived from the bone marrow of a patient with polycythemia vera. The purification procedure included chromatography on phosphocellulose and oligo(dT)-cellulose, and glycerol gradient centrifugation. The enzyme could be distinguished from polymerase A by salt elution from phosphocellulose, utilization of poly(rC) - oligo(dG) and its molecular size of about 70000, as determined by centrifugation. Throughout the purification procedure ribonuclease H activity was co-purified. Upon dodecylsulfate-polyacrylamide electrophoresis on microgradient gels two main bands with molecular weights of 68000 and 66000 and three minor bands were detected. The enzyme preferentially used poly(rA) - oligo(dT) as template-primer compared with poly(dA) - oligo(dT). It incorporated dGMP into polymer on poly(rC) - oligo(dG).     

A contribution to the cytostatic long-term chemotherapy of polycythemia vera (author's transl)

From 1967 to 1978 38 patients with polycythemia vera received a long term cytostatic chemotherapy. The results of 14 patients who were given Procarbacin (Natulan) during a mean treatment time of 9 1/2 years are communicated. Two patients showed secondary tumors after 9 or 11 years respectively. The advantages and disadvantages of the various therapeutic approaches in the management of polycythemia in the literature are discussed together with our own experiences.     

The use of the anti-factor VIII method on trephine biopsies of the bone marrow for the identification of immature and atypical megakaryocytes in myeloproliferative diseases and allied disorders. A morphometric study

A morphometric analysis was performed on trephine biopsies of the bone marrow to identify atypical megakaryocyte proliferation following PAS staining and the immunohistological demonstration of factor VIII. This study includes nine patients with a megakaryoblastic crisis in chronic myeloid leukemia (CML), four with acute megakaryoblastic leukemia (AM) and three with myeloid dysplasia later evolving into overt acute leukemia. Comparison and statistical evaluation of the PAS reaction with anti-factor VIII staining reveals that the latter technique not only facilitates the recognition of immature and abnormal megakaryocytes, but leads to a significantly increased count for all megakaryocytic elements in the bone marrow. Thus our retrospective investigation of routinely processed and paraffin-embedded trephine biopsies shows that the diagnosis of a megakaryoblastic crisis in CML as well as AM may be easily established with the aid of the anti-factor VIII method. In all cases of megakaryoblastic proliferation in CML and AM, the appearance of blasts was associated with moderate to pronounced myelofibrosis which could be also determined by morphometry.     

Quantitation of monoclonal plasma cells in bone marrow biopsies in plasma cell dyscrasia.

Direct measurement of monoclonal plasma cell mass in bone marrow biopsies may be a useful parameter to establish in plasma cell dyscrasia. In this study monoclonal plasma cells/mm in light chain immunoglobulin immunostained archival bone marrow sections from 22 patients in whom a diagnosis of multiple myeloma (MM) had been excluded but who had monoclonal proteins were counted by two observers at light microscopic level. There was good correlation between the counts of the two observers. The levels of monoclonal plasma cells/mm in biopsies were not related to the % counts in the aspirates taken at the same time as the biopsies. Three of seven patients with biopsy levels in excess of the polyclonal levels in patients without plasma cell dyscrasia developed progressive MM within the observation time. Monoclonal plasma cell levels/mm of bone marrow biopsies can be measured and they provide a useful parameter for the assessment of patients with low volume plasma cell dyscrasia.     

Polycythemia vera: electron microscopy of the bone marrow in 10 non-treated patients. A thin section and freeze-fracture study.

Electron microscopy (thin sections and freeze-fracture replicas) was performed on the bone marrow of ten patients with Polycythemia vera prior to any treatment. In addition to a hyperplasia of all three cell lineages and the sinuses, atypias were observed in the maturing erythroblasts. These aberrations of normal development consisted mainly of deep invaginations of the nuclear envelope in proerythroblasts and conspicuous nuclear clefts in erythro- and normoblasts. In comparison with similar changes in dyserythropoietic and aplastic anemia as well as leukemia these alterations are discussed in connection with disturbances of DNA synthesis. Further atypias involved megakaryopoiesis which displayed microforms probably as an evidence for maturation arrest. These ultrastructural abnormalities and their morphological features of a neoplastic proliferation of all three cell lineages in Polycythemia vera are in good agreement with the new concept of a transformation of a pluripotent stem cell with clonal character.     

Magnetic resonance (MR) evaluation of bone marrow in vertebral bodies.

Magnetic Resonance (MR) imaging was used to examine the hematopoietic bone marrow in the vertebral bodies of eight healthy subjects, and of 35 cancer patients who had been previously treated with radiation therapy. MR was instrumental in distinguishing viable hematopoietic tissue (red marrow) from adipose tissue (yellow marrow), whose presence reflected the extent of radiation-induced bone marrow injury. Different water content in proliferating hematopoietic tissue and adipose tissue enabled clear distinction of the two components even inside the same vertebral body. Three patterns of bone marrow viability were observed in irradiated patients: 1. Patients undergoing therapy at the time of MR study, and patients who had received low-intermediate dose several years before MR examination showed no alteration as compared with healthy controls (i.e. homogeneous presence of red marrow). 2. Patients who had received low-intermediate dose few years before MR, showed either partial re-colonization of yellow marrow or almost complete ablation of active red marrow with rare areas of re-colonization. 3. Patients who had received high dose, showed complete depletion of red marrow (fatty substitution) independently of the length of time elapsed since radiation therapy. Therefore, bone marrow recovery after radiation therapy was associate with two variables: received dose and length of time allowed for re-colonization by surviving hematopoietic tissue. In conclusion, our results provide evidence that MR can be purposively used to study composition and distribution of normal bone marrow, and to asses the extent of radiation-induced bone marrow injury; to monitor bone marrow recovery (or the lack of it); and in the general follow-up of treated cancer patients.     

Demonstration of mucinous-like carcinoma-associated antigen in bone marrow and lymph node biopsies from patients with breast carcinoma.

Two hundred and fifty bone marrow and 140 lymph nodal biopsies were analyzed immunocytochemically, using a mouse monoclonal antibody b-12 (M Ab b-12), which reacts with MCA (mucinous-like carcinoma-associated antigen). The presence of MCA in bone marrow specimens was demonstrated in 102 out of 105 (97.1%) breast cancer metastases, 5 out of 8 (62.5%) gastric cancers, 5 out of 6 (83.3%) colon cancers, 3 out of 5 (60%) prostate cancers, 11 out of 26 (42.3%) lung cancers and 25 out of 30 (83.3%) unknown primary cancers, while no positivity to anti-MCA antibody was found in 30 cases of normal bone marrow biopsies, 5 cases of non epithelial malignancies and 30 cases of hemolymphoproliferative disease. Normal lymph nodes and non-epithelial lymph node metastases did not show any reaction to M Ab b-12; on the contrary MCA positive staining was observed in 75 out of 75 (100%) lymph nodal metastases in breast cancer. These results suggest that application of M Ab b-12 in immunohistochemistry is valid for the detection of bone marrow and lymph nodal micrometastases of epithelial origin.