Knockout of LSD1 Gene by CRISPR/Cas9 System Significantly Inhibited Proliferation and Expression of CD235a in K562 Cells / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To study the effect of LSD1 knock-out on human chronic myeloid leukemia cells(K562 cells).</p><p><b>METHODS</b>The LSD1 gene in K562 cells was knocked-out specifically by using CRISPR/Cas9 system, the single cells were gained by flow cytometric sorting technique, the LSD1and LSD1cell lines were gained after amplificantion and culture, identification of Western blot and sequencing. The MTS assay was used to detect the effect of LSD1 knockout on the proliferation of K562 cells, the flow cytometry was used to examine the expression of K562 cell surface marker after LSD1 knockout.</p><p><b>RESULTS</b>The LSD1 stable knockout cell line of K562 (LSD1and LSD1)were successfully costructed. It was found that knockout of LSD1 significantly inhibited the proliferation of K562 and the expression of CD235a.</p><p><b>CONCLUSION</b>LSD1 plays a key role in the regulation of K562 cell proliferation and CD235a expression.</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>

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.

Radioiondine therapy for Graves hyperthyroidism with large goiter: feasibility, efficacy and safety / 南方医科大学学报

<p><b>OBJECTIVE</b>To evaluate the feasibility, efficacy and safety of radioiondine therapy in the treatment of Graves hyperthyroidism with large goiter.</p><p><b>METHODS</b>A total of 128 patients with Graves; hyperthyroidism with large goiter (thyroid weight>70 g) as the study group were treated with radioiondine, using 318 concurrent patients with Graves disease with a smaller goiter (thyroid weight<70 g) as the control group. The cure rate following a single-session treatment, the total cure rate and the incidence of hypothyroidism were compared between the two groups.</p><p><b>RESULTS</b>In the large goiter group, the total cure rate was 95.3%, and the cure rate following a single-session treatment was 46.9%, with the incidence of hypothyroidism of 4.7%, as compared with 90.9%, 65.7%, and 9.1% in the control group, respectively. A significant difference was noted in the cure rate following a single-session treatment (P=0.000), but not in the total cure rate or the incidence of early-onset hypothyroidism (P=0.115) between the two groups. No tracheal compression, laryngeal edema, or hyperthyroidism crisis occurred in the large goiter group after the treatment.</p><p><b>CONCLUSION</b>Radioiondine is safe and effective for treatment of Graves hyperthyroidism with large goiter, and results in a total cure rate and incidence of early-onset hypothyroidism similar to those in patients with goiters of a smaller size.</p>

Value of (18)F-FDG dual head coincidence imaging in predicting the efficacy of radioiodine therapy for papillary thyroid carcinoma with cervical lymph node metastasis / 南方医科大学学报

<p><b>OBJECTIVE</b>To assess the value of (18)F-FDG dual head coincidence imaging in the prediction of the efficacy of radioiodine therapy in patients with cervical lymph node (LN) metastasis of papillary thyroid carcinoma (PTC).</p><p><b>METHODS</b>Thirty-six patients undergoing thyroidectomy and radioiodine ablation of the residual normal thyroid tissue received (18)F-FDG dual head coincidence imaging and then therapeutic (131)I-whole body imaging ((131)I-WBI) in the same week. According to those imaging results, the patients were divided into group I with positive results of both imaging examinations and group II with positive results by (131)I-WBI but negative results by(18)F-FDG dual head coincidence imaging. All the patients were followed up for 6 months.</p><p><b>RESULTS</b>In group I (14 patients), a total of 49 lesions were diagnosed as cervical LN metastases, and the total sensitivity differed significantly between (18)F-FDG dual head coincidence imaging and (131)I- WBI (67.3% vs 89.8%, P=0.027). In both groups, the total sensitivity of (18)F-FDG dual head coincidence imaging and (131)I-WBI showed a significant difference (26.0% vs 94.5%, P<0.001). The target and non-target ratio (T/NT) was identified as one of the factors affecting the radioiodine efficacy (P<0.001). In group II (22 patients), 76 lesions were diagnosed as cervical LN metastases. The effective rates of groups I and II were 35.7% and 81.8%, respectively, showing a significant difference between them (P=0.011).</p><p><b>CONCLUSION</b>(131)I-WBI is more sensitive than (18)F-FDG dual head coincidence imaging in detecting cervical LN metastasis in patients with PTC. Patients with cervical LN metastases who have positive results in both (131)I-WBI and (18)F-FDG dual head coincidence imaging tend to have a poorer response to the therapy than the patients with negative results in (18)F-FDG dual head coincidence imaging. The T/NT of the cervical LN metastases in (18)F-FDG dual head coincidence imaging is associated with the efficacy of radioiodine therapy.</p>

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.

Value of additional skull lateral static imaging in whole-body bone imaging for skull bone invasion evaluation in nasopharyngeal carcinoma patients: comparison with CT / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the value of the additional skull lateral static imaging in whole-body bone imaging (WBI) vs CT for evaluation of skull base invasion in patients with nasopharyngeal carcinoma.</p><p><b>METHODS</b>A total of 405 patients with pathologically confirmed NPC underwent WBI with additional static imaging of the left and right skull as well as CT examination of the nasopharynx and skull base within one week before the radiotherapy.</p><p><b>RESULTS</b>The concordance rates between WBI and CT for positive and negative diagnosis were 29.48% and 76.05% in these cases, respectively, with the total concordance rate of 81.23%. The concordance rates between skull lateral static imaging with visual judgment and CT examination for positive and negative diagnosis were 67.95% and 74.07%, respectively, showing a total concordance rate of 87.16%. Skull lateral static imaging with semi-quantitative analysis and CT examination showed concordance rates for positive and negative diagnosis of 75.64% and 74.07%, respectively, with a total rate of 88.64%. In 27 patients with negative diagnosis by CT but a positive one in skull lateral static imaging with semi-quantitative analysis, 9 had a positive diagnosis by magnetic resonance imaging.</p><p><b>CONCLUSIONS</b>Skull lateral static imaging can be of value in the diagnosis of skull base invasion in NPC patients and may serve as an effective means for screening skull base invasion in NPC.</p>

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 congenital dyserythropoietic anemia-type I / 中国实验血液学杂志

The study was aimed to investigate the ultranstructural feature and diagnostic criteria of congenital dyserythropoietic anemia-type I (CDA-type I). Nucleated red cells in bone marrow from two patients with CDA-type I were analyzed by transmission electron microscopy (TEM). The results indicated that the erythropoietic/granulopoietic ratio was markedly increased with megaloblastic morphology in all stage of erythrocyte. Most proerythroblast showed of irregular nuclei, while the Swiss-cheese-appearance of the heterochromatin was usually found in basophilic and polychromatic erythroblast. About half of orthochromatic erythroblast illustrated karyolysis and karyorrhexis. Some orthochromatic erythroblast exhibited karyolysis and plasmolysis simultaneously. The inter-nuclear chromatin bridge between separated erythroblasts was seldom found by TEM. The nuclear membrane and rough endoplasmic reticulum were destructed at all stage of erythrocytes in different degree. In conclusion, the megaloblastic erythrosis was the main characteristic of CDA-type I, and then nuclear membrane disruption in polychromatic erythroblast and karyolysis or karyorrhexis in orthochromatic erythroblast. The universal breakdown of cytoplasm membranous system was fundamental pathogenesis of CDA-type I.

Cardioprotective effects of atorvastatin preconditioning via iNOS upregulation in a rabbit ischemia/reperfusion model / 中华心血管病杂志

<p><b>OBJECTIVE</b>To observe the preconditioning cardioprotection of atorvastatin (ATV) in rabbits underwent 40 min ischemia and 240 min reperfusion and to explore related mechanisms.</p><p><b>METHODS</b>The rabbits were randomized divided into Control group, ATV group (10 mg.kg(-1).d(-1) for 3 days before ischemia), ATV plus iNOS inhibitor S-methylisothiourea sulfate group (ATV + SMT group), SMT group, ATV plus mito K(ATP) channel blocker 5-hydroxydecanoate group (ATV + 5-HD group) and 5-HD group (n = 16 each group). The infarction size, CK-MB, LDH-1, nitric oxide synthase and mitochondrial ATP synthesization capacity ([ATP] m) were determined at the end of reperfusion.</p><p><b>RESULTS</b>Infarction size, CK-MB, LDH-1 were decreased by 26.3%, 31.4%, 19.1% and iNOS, [ATP] m increased by 102.6%, 46.8% post ATV compared to control group (all P < 0.05) and these effects could be blocked by cotreatment with SMT and 5-HD except the iNOS was not affected by 5-HD.</p><p><b>CONCLUSION</b>The atorvastatin preconditioning exerted cardioprotection by upregulating iNOS and activating mito K(ATP).</p>

Ultrastructural characteristics of megakaryocytes in 11 patients with acute megakaryocytic leukemia / 中国实验血液学杂志

The purpose of study was to investigate the ultrastructural features of leukemic megakarocyte (LMK) in patients with acute megakaryocytic leukemia (M(7)). Analyzing the ultrastructure characteristics of LMK and positive ratio of platelet peroxides (PPO) in 11 patients with M(7) were analyzed on basis of transmission electron microscopic observation retrospectively. The results showed that the diameter of LMK in 7 out of 11 cases was less than 20 microm, in 2 cases of them, the LMK diameter was from 10 to 15 microm and their PPO positive ratio was more than 50%, most LMK displayed regular shape, less protrusions, irregular nucleus, high nuclear/cytoplasm ratio, tiny granules, undeveloped demarcation membrane system (DMS) and irregular tubules in cytoplasm; in 5 out of those 7 cases the diameter of LMK was about 20 microm, PPO positive cell count was from 8% to 22%, most showing round or horseshoe nuclei, more or less heterochromatin, no DMS and granules were found in LMK in 3 cases and 2 cases occasionally. In other 5 out of 11 cases, the diameter of LMK was from 20 to 40 microm and PPO positive ratio was from 16% to 80%, in which smaller LMKs were similar to those in former cases in shape, and the larger LMK had irregular protrusions, varied nuclear/cytoplasm ratio, more heterochromatin, prominent nucleolus, some of them contained developed DMS, tubules and alpha-granules. It is concluded that most patients with M(7) are predominant of LMK in stage-I and minority contained LMK in II or III stage simultaneously. The differentiation degrees of LMK are different in individual and various cases.

Therapeutic efficacy of strontium-89-chloride for bone metastatic tumors without bone pain / 南方医科大学学报

<p><b>OBJECTIVE</b>To evaluate the safety and efficacy of strontium-89-chloride for management of bone metastases in patients without bone pain.</p><p><b>METHODS</b>Fifty-four patients without painful bone metastases were given a single intravenous dose (1.48-2.22 MBq/kg) of strontium-89-chloride, which was repeated once or twice at the interval between 3 and 6 months.</p><p><b>RESULTS</b>The total response rate was 74.0% in these, and the response rate was significantly lower in patients with focal size>2 cm than in those with focal size<or=2 cm (33.3% vs 66.6%, chi2=14.9, P<0.01). The side effects of strontium-89-chloride included mainly thrombocytopenia and neutropenia, mostly mild and reversible without interventions.</p><p><b>CONCLUSION</b>Strontium-89-chloride is effective and safe for treatment of nonpainful bone metastases.</p>

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.

In vitro expansion of cord blood megakaryocyte progenitor / 中国医学科学院学报

<p><b>OBJECTIVE</b>To expand cord blood megakaryocyte progenitor cells in vitro.</p><p><b>METHODS</b>Cord blood CD34+ cells were selected by magnetic cell sorting (MACS), and thrombopoietin (TPO), interleukin-11 (IL-11), and heparin were used in the expansion system of megakaryocyte progenitor. The expansion efficiency was measured by fluorescence-activated cell sorting (FACS) using the megakaryocytic specific monoclonal antibodies (CD34+, CD41a+, CD61+, CD34+CD41a+, CD41a+CD61+) and colony-forming units-megakaryocyte (CFU-MK) analysis. The expanded megakaryocyte progenitor were determined by histochemistry staining using CD41a and the observation of the ultrastructure of megakaryocyte (MK) by electron microscopy. The megakaryocyte function were examined by the platelet activation in vitro and nonobese diabetic/severe combined immunodifficiency (NOD/SCID) mice transplantation in vivo.</p><p><b>RESULTS</b>CD34+CD41a+ cells was expanded (4.0 +/- 1.7) folds on day 7 in TPO (50 ng/ml) group and (10.5 +/- 4.8) fold in TPO combined with IL-11 group; after heparin was joined in on day 0, a more significantly elevated expansion was found in the heparin, TPO, and IL-11 group [(29.9 +/- 6.4) folds than the above two groups; P < 0.05]. Meanwhile, the large CFU-MK colony (> 50 cells/colony) was (106.8 +/- 26.9) folds on day 7 (P < 0.05). The megakaryocyte expanding with TPO, IL-11 and heparin for 7 days in vitro transplanted the NOD/SCID mice fasten the recovery of platelet and white blood cell account and improved the survival. Megakaryocyte under culture displayed certain development of territories membrane. Platelet activation test comfirmed that the expanding megakaryocyte progenitor had the normal function.</p><p><b>CONCLUSION</b>TPO, IL-11, and heparin combination system for ex vivo expansion is an effective expansion system of megakaryocyte progenitor.</p>

Preliminary study on extensive amplification of human dendritic cells differentiated from cord blood CD34+ progenitor cells by two-step culture / 中华血液学杂志

<p><b>OBJECTIVE</b>To Explore a two-step culture system to generate a large number of dendritic cells (DC) differentiated from cord blood (CB) CD(34)(+) cells.</p><p><b>METHODS</b>Enriched CB CD(34)(+) cells with immunoadsorption were primarily cultured in the presence of stem cell factor (SCF), Flt-3 ligand (FL), thrombopoietin (Tpo) and interleukin-3 (IL-3) for 7 (group I), 10 (group II) or 14 days (group III) respectively, and then further cultured with GM-CSF, IL-4 and TNF-alpha for 5 - 8 days to induce DC. The expansion and cell function were evaluated by flow cytometry (FCM) and mix-lymphocyte reaction (MLR), and detection of IL-12 in the supernatant by using ELISA.</p><p><b>RESULTS</b>The total nucleated cells with 53.39 +/- 20.59-, 307.17 +/- 119.59- and 1117.25 +/- 335.49-folds expansion could be respectively obtained after 7 - 14 days of expansion culture. After DC induction, CD(1a)(+) cells were 21.40 +/- 16.70-, 143.2 +/- 60.35- and 150.8 +/- 42.16-fold increase as compared to the initial nucleated cells. Comparing with that in group I, the CD(1a)(+) cells were much more in groups II and III; but there was no difference between the latter two groups (P > 0.05). The cultured cells in the three groups showed almost the same allo-stimulatory capability and IL-12 excretion when the second culture duration maintained 8 days, while the capability and excretion were greatly decreased when the duration shortened to 5 days (P < 0.05).</p><p><b>CONCLUSION</b>A plenty of functionally mature DC could be obtained from the CD(34)(+) cells in the two-step culture system of 7 - 10 days HSC expansion followed by 8 days DC induction.</p>

Clinical and laboratory study of a case of hematidrosis / 中华血液学杂志

<p><b>OBJECTIVE</b>To study the clinical characteristics and pathogenesis of hematidrosis.</p><p><b>METHODS</b>Detailed clinical manifestations and natural history of a patient with hematidrosis were presented. A series of laboratory examinations were performed, and skin pathohistologic features and ultra microscopic structures were observed.</p><p><b>RESULTS</b>The episodes of skin bleeding occurred on any site of the body spontaneously and promptly. The skin surface bloody extravasation has identical cell components as that of peripheral blood. All the results of laboratory tests were normal except a positive Trousseau's test. Skin pathohistological study revealed some intradermal bleeding and emphraxised capillaries. No abnormality was found in sweat glands, hair follicles and sebaceous glands.</p><p><b>CONCLUSION</b>The pathological basis for hematidrosis might be a distinctive vasculitis.</p>