Ultrastructure and Raman Spectral Characteristics of Two Kinds of Acute Myeloid Leukemia Cells / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the Raman spectral characteristics of leukemia cells from 4 patients with acute promyelocytic leukemia (M) and 3 patients with acute monoblastic leukemia (M), establish a novel Raman label-free method to distinguish 2 kinds of acute myeloid leukemia cells so as to provide basis for clinical research.</p><p><b>METHODS</b>Leukemia cells were collected from bone marrow of above-mentioned patients. Raman spectra were acquired by Horiba Xplora Raman spectrometer and Raman spectra of 30-50 cells from each patient were recorded. The diagnostic model was established according to principle component analysis (PCA), discriminant function analysis (DFA) and cluster analysis, and the spectra of leukemia cells from 7 patients were analyzed and classified. Characteristics of Raman spectra were analyzed combining with ultrastructure of leukemia cells.</p><p><b>RESULTS</b>There were significant differences between Raman spectra of 2 kinds of leukemia cells. Compared with acute monoblastic leukemia cells, the spectra of acute promyelocytic leukemia cells showed stronger peaks in 622, 643, 757, 852, 1003, 1033, 1117, 1157, 1173, 1208, 1340, 1551, 1581 cm. The diagnostic models established by PCA-DFA and cluster analysis could successfully classify these Raman spectra of different samples with a high accuracy of 100% (233/233). The model was evaluated by "Leave-one-out" cross-validation and reached a high accuracy of 97% (226/233).</p><p><b>CONCLUSION</b>The level of macromolecules of Mcells is higher than that of M. The diagnostic models established by PCA-DFA can classify these Raman spectra of different cells with a high accuracy. Raman spectra shows consistent result with ultrastructure by TEM.</p>

The effect of transmission electron microscopy on diagnosis of acute myeloid leukemia / 中华血液学杂志

<p><b>OBJECTIVE</b>To analyze coincidence rate of acute myeloid leukemia (AML) sub-typing between transmission electron microscopy (TEM) and clinical discharge diagnosis.</p><p><b>METHODS</b>Reviewing sub-typing results of TEM, light microscopy, flow cytometric analyzing, molecular biological detection and karyotype in 793 AML cases, comparing their coincidence rates with discharge diagnosis to reveal advantages of AML sub-typing by TEM.</p><p><b>RESULTS</b>General coincidence rates of TEM, light microscopy, flow cytometric analyzing, molecular biological detection and karyotype on AML sub-typing were 63%, 59%, 52%, 47%, 26% and 23% respectively, and clinical coincidence rates of TEM on M1, M2a, M4 and M5, M6, M7, t (8; 21) and t (15; 17) were 39%, 34%, 17%, 74%, 50%, 73%, 87% and 89% respectively.</p><p><b>CONCLUSION</b>TEM has a higher coincidence rate in general AML sub-typing, especially strong screenings on t (15; 17), t (8; 21), M7, M5 and M6, but lower coincidence rates on M1, M2a and M4 sub-typing than other methods.</p>

A mouse model of myelofibrosis induced by high dose of recombinant human erythropoietin / 中国实验血液学杂志

Tn order to set up a mouse model of myelofibrosis (MF) induced with high dose recombinant human erythropoietin (rhEPO). 60 mice were collected and divided into EPO and control groups, the former was injected with rhEPO and the latter with normal saline intraperitoneally. 5 mice from each group were executed on day 6, 30, 60, 90, 120 and 150 respectively. Their WBC count, Hb level, MCV, RDW and platelet amount were measured by automatic blood cell analyzer; CD34(+) cell ratio in bone marrow were analyzed by flow cytometry; liver and spleen coefficients were measured; pathological changes of liver, spleen, femur were observed by HE staining and reticular fibers staining; cortex thickness, femoral canal diameter and lumbar spine density were determined by computerized tomography (CT). The results indicated that as compared with normal control group in EPO induced group, WBC count was increased slightly in whole period, but without statistic significance (p > 0.05), Hb level and RDW increased at day 6 and 30 significantly (p < 0.05), MCV increased at day 6 significantly (p < 0.05), but platelet amount decreased significantly at all time points (p < 0.05). Most mice in EPO-induced group had hepatomegalia and their liver and spleen coefficient increased significantly at day 60 (p < 0.05), while most mice had splenomegaly and its coefficient was increased significantly at all time-points (p < 0.05). CD43(+) cell ratio of EPO group increased significantly in whole period (p < 0.05). CT scanning displayed femoral cortical thickening, medulla canal narrowing and lumbar spine density increasing at day 150, meanwhile, HE staining and reticular fiber staining showed the fatty degeneration or vacuolization in liver, splenomegaly with megakaryocytic proliferation, femur bone marrow fibrosis and osteosclerosis. It is concluded that the mouse induced by high dose of rhEPO displays the myelofibrosis associated with splenic extramedullary hemopoiesis, and this study is useful to establish a practical MF model, and to explore its pathological mechanism.

Characteristics and clinical significance of CD73 expression in subtypes of leukemia / 中国实验血液学杂志

The study was purposed to investigate the expression of CD73 on bone marrow nucleated cells (BMMNC) in various leukemia subtypes and its relationship with cell differentiation of leukemia. Immunocytochemistry staining and Wright-Giemsa staining of BMMNC from 75 cases of leukemia, 11 cases of myelodysplastic syndrome (MDS), 13 cases of non-leukemic patients and 9 healthy adults were performed, and the CD73(+) ratio in BMMNC and its relationship with differentiation of leukemia cells were analyzed. The results showed that the ratios of CD73(+) in BMMNC of com-B ALL, pre-B ALL and PLL were significantly higher than those in B-CLL (p < 0.05). CD73(+) ratios in AML subtypes of M(1), M(2a), t (8; 21), t (15; 17), M(4) and M(5) were markedly higher than those in MDS respectively, but in M(6) and MDS were lower and had no statistical difference between them. CD73(+) ratios in T-ALL, B-CLL, M(6), MDS, non-leukemia patients and healthy adults were close to each other and all of them were lower than those in B-ALL and other AML subtypes. It is concluded that the expression of CD73 is associated with leukemia subtype, differentiation and development. The higher differentiation of leukemia cells, the lower of CD73 expression in myeloid and B lymphoid leukemia, but T-ALL does not meet this pattern.

Correlation between ultrastructural abnormality of bone marrow cells and anemia and neutropenia in myelodysplastic syndrome / 中国实验血液学杂志

The aim of study was to investigate the relationship of anemia and neutropenia with ultrastructural abnormalities of erythroblasts and young neutrophils in bone marrow of patients with myelodysplastic syndrome (MDS). Anemia parameters and peripheral neutrophil amount of 74 patients with MDS were measured by automatic hemocyte analyzer. According to Hb value and neutropenia degree, MDS patients were divided into 4 groups: normal, mild, middle and severe anemia or neutropenia. The morbid rate and apoptosis rate of erythroblasts and young neutrophils in bone marrow were measured by transmission electron microscopy (TEM). The results indicated that 68 out of 74 patients were consistent with anemia diagnostic criteria, and 51 out of 68 patients were with neutrocytopenia. TEM showed different abnormal features of erythroblasts and young neutrophils in all patients. The morbid rates of erythroblasts in normal, mild, middle and severe anemia groups were 37 ± 14.7%, 24 ± 9%, 32 ± 16% and 34 ± 21% respectively, while apoptotic rates of erythroblasts in normal, mild, middle and severe anemia groups were 2.25 ± 1.03%, 4.43 ± 2.60%, 8.78 ± 4.04% and 11.67 ± 4.57% respectively. The morbid rate and apoptotic rate of erythroblasts were correlated negatively with Hb and HCT value (p < 0.05). The apoptotic rates of bone marrow young neutrophils in 4 groups with different degree of neutropenia were 6.00 ± 2.67%, 9.50 ± 4.42%, 13.00 ± 3.54% and 17.00 ± 2.39%, which correlated negatively with peripheral neutrophil quantity (p < 0.01). Morbid rates of neutrophils in normal, mild, middle and severe anemia groups were 12.25 ± 16.31%, 13.5 ± 10.01%, 23 ± 8.59% and 51.67 ± 19.67% respectively, which positively correlated with its apoptotic rates (p < 0.01). It is concluded that anemia and neutropenia in patient with MDS are correlated with apoptosis and morbid rate of erythroblasts and young neutrophils in bone marrow, which may result in ineffective hematopoiesis.

Ultrastructural analysis of 5' nucleotides distribution in acute myeloid leukemia subtypes / 中国实验血液学杂志

5' nucleotides (5'NT), a purine degradative enzyme, is capable of hydrolyzing nucleotide and acting as a phosphotransferase simultaneously. It has critical role in maintaining nucleotide metabolism balance. The present study was aimed to investigate the expression of 5'NT in bone marrow granulocytes (BMGs) from patients with acute myeloid leukemia (AML) and healthy donors comparatively. The BMGs were isolated from bone marrow of 33 patients with AML and 6 healthy donors by using lymphocyte isolating solution. The reactivity of 5'NT was detected by electron microscope and cytochemistry of cytidine monophosphate (CMP). The positive BMG ratio and their index were calculated on the base of ultrastructural observation semiquantitatively. The results indicated that electron microscopy revealed plasma membrane reacting pattern of CMP. Most BMGs from normal donors were CMP negative or exhibited lower active degree. All cases of M(0), M(1), M(2) and t (8; 21) showed high positive percentages and high indexes of BMGs, but no statistic differences between them. APL of t (15; 17) shared lower percentages and indexes than other subtypes. There was no significant difference between APL and normal donors statistically. In conclusions, the results suggested the expression of 5'NT may be associated with BMG differentiation in AML, and APL of t (15; 17) may be a highly differentiated leukemia subtype.

Ultrastructural characteristics of malignant T cell in T cell lymphoma / 中国实验血液学杂志

In order to investigate the ultrastructural features of malignant T cell (MTC) in bona marrow aspirate (BMA) from patients with T Cell Lymphoma, the antigen expression of MTC was analyzed by flow cytometry, and the ultrastructural features of MTC in BMA from 13 T-cell lymphoma patients with bone marrow involvement (BMI) were observed by transmission electron microscopy. The results indicated that the sizes of MTC were uneven in every patient and their diameter were between 12 and 28 microm, in 6 out of 13 cases sizes of MTC were slightly uneven but in 7/13 cases sizes of MTC were significantly uneven. The heterochromatin of MTC was less than that of normal T cell and nucleolus diameter was from 2 to 8 microm in all cases. The nuclear contour of MTC was strikingly irregular in 10 out of 13 cases. The MTC had plenty of cytoplasm in 8 out of 13 cases and displayed many microvilli or processes on MTC surface in 7 out of 13 cases, while MTC in 6 out of 13 cases contained more Golgi's apparatuses, secretary vacuoles, dense granules and intermediate filaments. In 8 out of 13 cases mitochondria apparently swelled. It is concluded that the size of MTC increase unevenly in all patients. MTC nuclear contour in most cases is irregular by folding, indenting, and twisting, which often correlated with arising of paranuclear intermediate filaments. Processes and microvilli on surface and Golgi's apparatus, secretary vesicles, dense granules as well as intermediate filament in cytoplasm of MTC develop synchronously, meanwhile, mitochondria of MTC strikingly swell in most cases.