Influencing factors of iron metabolism assessment in patients with myelodysplastic syndrome: A retrospective study / 中华血液学杂志

Objective: To analyze the influencing factors of iron metabolism assessment in patients with myelodysplastic syndrome. Methods: MRI and/or DECT were used to detect liver and cardiac iron content in 181 patients with MDS, among whom, 41 received regular iron chelation therapy during two examinations. The adjusted ferritin (ASF) , erythropoietin (EPO) , cardiac function, liver transaminase, hepatitis antibody, and peripheral blood T cell polarization were detected and the results of myelofibrosis, splenomegaly, and cyclosporine were collected and comparative analyzed in patients. Results: We observed a positive correlation between liver iron concentration and ASF both in the MRI group and DECT groups (r=0.512 and 0.606, respectively, P<0.001) , only a weak correlation between the heart iron concentration and ASF in the MRI group (r=0.303, P<0.001) , and no significant correlation between cardiac iron concentration and ASF in the DECT group (r=0.231, P=0.053) . Moreover, transfusion dependence in liver and cardiac [MRI group was significantly associated with the concentration of iron in: LIC: (28.370±10.706) mg/g vs (7.593±3.508) mg/g, t=24.30, P<0.001; MIC: 1.81 vs 0.95, z=2.625, P<0.05; DECT group: liver VIC: (4.269±1.258) g/L vs (1.078±0.383) g/L, t=23.14, P<0.001: cardiac VIC: 1.69 vs 0.68, z=3.142, P<0.05]. The concentration of EPO in the severe iron overload group was significantly higher than that in the mild to moderate iron overload group and normal group (P<0.001) . Compared to the low-risk MDS group, the liver iron concentration in patients with MDS with cyclic sideroblasts (MDS-RS) was significantly elevated [DECT group: 3.80 (1.97, 5.51) g/L vs 1.66 (0.67, 2.94) g/L, P=0.004; MRI group: 13.7 (8.1,29.1) mg/g vs 11.6 (7.1,21.1) mg/g, P=0.032]. Factors including age, bone marrow fibrosis, splenomegaly, T cell polarization, use of cyclosporine A, liver aminotransferase, and hepatitis antibody positive had no obvious effect on iron metabolism. Conclusion: There was a positive correlation between liver iron concentration and ASF in patients with MDS, whereas there was no significant correlation between cardiac iron concentration and ASF. Iron metabolism was affected by transfusion dependence, EPO concentration, and RS.

Study on phenotypic and cytogenetic characteristics of bone marrow mesenchymal stem cells in myelodysplastic syndromes / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate phenotype, cell differentiation and cytogenetic properties of bone marrow (BM) mesenchymal stem cells (MSC) separated from the myelodysplastic syndrome (MDS) patients. And to analyze cytogenetic aberration of MSC derived from MDS (MDS-MSC) and its mechanism in pathogenesis of MDS.</p><p><b>METHODS</b>Adherent MSC from both myelodysplastic (n = 22) and normal (n = 7) marrow were obtained by a stromal culture procedure. Morphological features were observed by optical microscope. The cell-surface antigens were performed by flow cytometer(FCM). Adipogenic and osteogenic differentiation potential of MSC were identified under specific induction conditions. Standard cytogenetic analysis of both hematopoietic cells and MSC were performed by trypsin-Giemsa (GTG) banding. The karyotype analysis DNA content was determined by FCM to verify the results.</p><p><b>RESULTS</b>The morphology of MDS-MSC was typical slender spindle-shaped cells, MSC obtained from MDS patients had a MSC immunophenotype, lacked the expression of hematopoietic antigens-CD34, CD45 and expressed MSC markers, such as CD73, CD90, and CD105. MDS-MSC layers showed the capability to differentiate towards adipocytes, chondrocytes and osteoblasts. Cytogenetic aberrations were observed in MSC from 14 (64%) MDS patients, usually involve the loss of chromosomal material (92%), and the clonal loss (7 cases, 50%). Two cases of structural aberrations were also detected. Abnormal karyotypes in MSC were still more frequently identified in abnormal hematopoietic cells group (12 out of 13, 92% vs 3 out of 9, 33%, P < 0.05). There were not exactly the same type of chromosomal aberrations between hematopoietic cells and MSC, but different type of the aberrations in the same chromosome were involved.</p><p><b>CONCLUSION</b>MDS-MSC retains the phenotyping characteristics and differentiated function of normal MSC, but has different type of chromosomal abnormalities. A high proportion of loss of chromosomal may be a marker of chromosomal instability of MDS-MSC. Detection of abnormalities in MDS-MSC suggests enhanced genetic susceptibility of these cells in MDS. This may indicate potential involvement of MSC in the pathophysiology of MDS.</p>

Function of peripheral blood Th17 cells in patients with multiple myeloma / 中国实验血液学杂志

This study was aimed to investigate the effects of peripheral blood Th17 cells and their secreting IL-17, IL-21 in the occurrence and development of multiple myeloma (MM). A total of 55 patients with MM were divided into non-remission group (group A , n = 30), remission group (group B, n = 25); healthy volunteers were used as control group (group C , n = 30). The concentration of IL-17, IL-21 and IL-6 in the peripheral blood mononuclear cell (PBMNC) culture supernatant were determined with ELISA. The ratio of Th7 cells in PBMNC was determined by flow cytometry. The results showed that IL-17, IL-21 levels and ratio of Th17 cells in group A were much higher than those in group B and C (P < 0.05), IL-17, IL-21 and the ratio of Th17 cells between group B and group C were not significantly different (P > 0.05); IL-17 level in non-remission MM group positively correlated with IL-6 level (r = 0.782, P < 0.05), IL-21 level in non-remission MM group positively correlated with IL-6 level (r = 0.778, P < 0.05). It is concluded that Th17 cells as a initiating factor may be involved in the immune pathogenesis of MM patients, promoting the progress of the disease.

Current understanding of iron overload hazard in patients with myelodysplastic syndrome / 中国实验血液学杂志

Patients with myelodysplastic syndromes (MDS) become dependent on blood transfusions and develop into transfusional iron overload, which is exacerbated by increased absorption of dietary iron in response to ineffective erythropoiesis. However, it is uncertain whether there is an association among iron accumulation, clinical complications, and decreased likelihood of survival in MDS patients. Thereby our current understanding of the effects of transfusion dependency and iron overload in MDS are discussed. Particular emphasis should be placed on further characterizing the role of redox-active forms of labile iron and oxidative stress in iron overload, decreased life expectancy and increased risk of leukemic transformation in MDS patients with iron overload.

Tissue factor expression of platelets and leukocytes in patients with acute coronary syndromes / 中国实验血液学杂志

The aim of this study was to investigate the tissue factor (TF) expression of platelets and leukocytes in patients with acute coronary syndrome (ACS), patients with stable angina (SA) and healthy subjects (as controls). 26 patients with ACS, 29 patients with SA, and 25 controls were enrolled in this study. The peripheral blood samples of above-mentioned subjects were collected and isolated to obtain the monocytes and platelet-rich plasma, the TF-mRNA expression of monocytes, and platelets among 3 groups was detected by RT-PCR, the TF expression ratio of platelets, platelet-leukocyte aggregates (PLA) and platelet-monocyte aggregates (PMP) was detected by flow cytometry among 3 groups. The results showed that the TF mRNA expression level of platelets in ACS group were significantly higher (3.11 ± 0.51 relative expression) as compared with SA and control groups (1.88 ± 0.78 and 0.7 ± 0.1, respectively) (P = 0.03). Expression of TF mRNA of monocytes was higher in ACS group (P = 0.05 versus controls) too. ACS group had a significantly higher amount of TF-positive platelets (8.8 ± 2.6) than SA (2.6 ± 0.5, P = 0.02) or control groups (2.5 ± 0.4, P = 0.02). A significantly greater number of TF positive platelet-leukocyte aggregates and platelet-monocyte aggregates were also found by flow cytometry in blood of ACS patients than in either SA patients or controls. It is concluded that the high TF expression of platelets and leukocytes in ACS patients strengthens the platelet activation, blood coagulation, and thrombus formation and may further contribute to the hypercoagulability associated with the disease. The present study further extends the proinflammatory/prothrombotic phenotype of ACS patients showing that new players on the scene.

p15(INK4B) methylation on prognosis and response to decitabine in patients with MDS / 中华血液学杂志

<p><b>OBJECTIVE</b>To detect p15(INK4B) methylation levels and the kinetics of the methylation status before and after decitabine to explore its influences on prognosis and response to decitabine in myelodysplastic syndromes (MDS) patients.</p><p><b>METHODS</b>We examined 261 MDS patients (143 male and 118 female) with the median age of 52 years (32-78). Of them, 172 cases were low-risk group (low-risk 104 cases, intermediate-1 68 cases), 89 cases high-risk group (intermediate-2 52 cases, high risk 37 cases). Collections of bone marrow mononuclear cells of MDS patients and extracted the genomic DNA, the methylation status of p15(INK4B) was detected by methylation-specific PCR (MSP) method. Survival analysis was conducted according to the level of p15(INK4B) methylation in the cohort of patients. The kinetics of the methylation levels of p15(INK4B) in 58 MDS patients before and after 2 courses of decitabine have been assessed with the method of MSP.</p><p><b>RESULTS</b>The methylation level of p15(INK4B) in low-risk group patients was significantly lower than that in high-risk group (117.22 vs 157.63, P<0.05 ). The expected 2-year survival rate of p15(INK4B) methylation positive patients was lower than that of negative ones (91.8% vs 69.8%, P<0.05); the expected 2-year survival rate of p15(INK4B) methylation positive patients was shorter than that of negative ones in low-risk group（78.2% vs 92.0%, P<0.05）, meanwhile there was no significant difference in terms of expected 2-year survival rate and median expected survival between p15(INK4B) methylation positive and negative patients in high-risk group [35.6% vs 38.5%, (17.0±9.3) month vs (18.0±5.7) month, P>0.05]. Multivariate analysis showed p15(INK4B) methylation degree was an independent prognostic factor for overall survival. No statistical difference of overall response (OR) rates were found between p15(INK4B) methylation positive patients and negative patients before decitabine（65.9% vs 76.5%, P>0.05）, and complete remission (CR) rates between these two groups also showed no statistical difference（22.0% vs 29.4%, P>0.05）. p15(INK4B) methylation levels had no obvious change before and after treatment in decitabine responders（P>0.05）.</p><p><b>CONCLUSION</b>The survival of newly diagnosed MDS patients with positive p15(INK4B) methylation was comparatively shorter, but p15(INK4B) methylation had no influence on response to decitabine.</p>

Cytogenetic study of autosomal monosomies among myelodysplastic syndrome patients / 中国实验血液学杂志

Monosomic karyotype (MK) has recently been associated with poor prognosis of myelodysplastic syndromes (MDS). The objective of the current study was to investigate the prevalence and spectrum of autosomal monosomies in an unselected cohort of patients with known or suspected MDS by using retrospective analysis. The results showed that bone marrow cytogenetic studies (1532 cases) were performed between 2004 and 2012, and an abnormal karyotype was found in 538 cases (35.1%). In the 538 cases, 202 (37.5%) cases had autosomal monosomies including sole (n = 47, 23.3%), part of two (n = 33, 21.3%) or more (n = 122, 78.7%) anomalies. Almost all 22 autosomes were involved, but monosomy 7 was by far the most frequent, constituting 66.1% of all isolated monosomies and the highest fraction of those with two or more abnormalities. Other recurrent sole monosomies included chromosomes 20 (15.0%) and 13 (8.5%). Monosomy 13 (12.5%), 18 (8.3%), 20 (6.3%), 17 (7.3%), 21 (5.2%), 5 (5.2%) and 12 (5.2%) were also recurrent in the setting of 3 abnormalities. Bone marrow histology and clinical information were reviewed in all cases with isolated monosomy; associated clinical phenotypes were found in RCMD (n = 20, 13 were -7), RAEB (n = 12, 11 were -7), RA (n = 9, 3 were -7) and chronic myelomonocytic leukemia (CMML, n = 6, 4 were -7) cases. Sole monosomy 20 (n = 7, RA 3 case and RCMD 4 cases) was not detected in RAEB or CMML cases. It is concluded that the presence of at least 1 autosomal monosomy was documented in approximately 37.5% of all abnormal cases, which has potential impact on a more than trivial fraction of patients with MDS. The preponderance of monosomy 7 implicates a pathogenetic role for haploinsufficiency of genes associated with chromosome 7. The rarity of sole monosomy involving other chromosomes other than 7, 20, and 13 suggests that haploinsufficiency involving entire chromosomes is detrimental to cell survival, unless their effect is overcome by the presence of other genetic changes that are often associated with additional chromosomal abnormalities. The observation is consistent with the usually favorable prognostic profile associated with sole monosomy 20.

Impact of immunochemotherapy on prognostic factors in diffuse large B-cell lymphoma patients / 中国实验血液学杂志

The international prognostic index (IPI) has been established as one of the best predictors of outcome, and several different immunologic subtypes have been established as independent predictors of diffuse large B-cell lymphoma (DLBCL). This study was aimed to reassess and re-evaluate the useful value of these prognostic predictors in patients treated with immunochemotherapy. A retrospective analysis of clinical records of immuno-chemotherapeutic (rituximab + CHOP, R-CHOP) and route chemotherapeutic (CHOP) groups was carried out. Standard two-step method of immunohistochemical staining was used to assess the expression of CD10, MUM-1, BCL-6 and BCL-2. The different classification models (Han's algorithm and Muris model) were performed for patients with DLBCL according to the immunohistochemical staining results in both R-CHOP and CHOP regimen groups. Then the data of remission and overall survival rate in different groups were analyzed to investigate the effect of these prognostic factors. Total 126 de novo DLBCL patients were collected in this study, including 51 patients with treatment of R-CHOP and the other 75 patients with treatment of CHOP. The results showed that the R-CHOP group had higher complete remission rate (68.8) than CHOP group (58.7). The patients with IPI score ≤ 2 had significantly higher overall response rate and overall survival rates than the patients with IPI scores > 2 in both groups. The survival rates of different subtypes in Han's and Muris models were not different from each other in R-CHOP group, but were obvious different from each other in CHOP group. It is concluded that IPI is still effective and predictive for identification of different risk groups. Immunochemotherapy can improve the remission and overall survival rate of DLBCL, but weaken the effect of outcome predictor.

Clinical therapeutic efficacy of rituximab for relapsed and refractory idiopathic thrombocytopenic purpura / 中国实验血液学杂志

The aim of this study was to evaluate the effect of rituximab treatments for refractory and relapsed idiopathic thrombocytopenic purpura (ITP). 18 patients with refractory and relapsed ITP who received 22 courses of rituximab treatments from January 2007 to December 2010 were analyzed retrospectively. Rituximab was given at a dose of 375 mg/m(2) intravenously weekly for a continuous 4 weeks. The results indicated that responses were achieved in 15 of 22 (68%) courses, out of which complete responses were achieved in 10 of 22 (45%) courses, partial and minimal responses were achieved in 5 of 22 (23%) courses, and no responses were achieved in 7 of 22 (32%) courses. The median time of response was 3 weeks (1 - 17 weeks) from the start of treatment and median duration of response was 13 weeks (1 week - 42 months). The responses were mostly short-sustained and follow-up median time was 20 months (1 - 52 months). The responses of 8 patients (36%) sustained for over 6 months, 6 patients (27%) sustained for over 1 year, and 4 patients also showed sustained response at last visit of follow-up. Previous splenectomy resulted in a poor response (P = 0.037). Patients who failed in rituximab treatment and prior received multiple treatments including splenectomy, had a poor response to further therapies. It is concluded that rituximab is well tolerated by patients and is useful in some patients with relapsed and refractory ITP, however, only about 20% patients can achieve sustained remissions. The patients who failed in rituximab treatment has a poor response to further treatment.

Yield of CD34(+) cells in graft can be increased significantly by G-CSF used at appropriate time after chemotherapy for AutoPBSCT / 中国实验血液学杂志

This study was aimed to investigate the influence of timing using G-CSF after chemotherapy on graft yield of mobilized peripheral blood stem cells for autoPBSCT. 39 patients with lymphoma or multiple myeloma (MM) received the same chemotherapy mobilization regimen, including CTX 400 mg/m² d1; VLB 2 mg/m(2) d1; Ara-C 60 mg/m ²× d1-5; VP-16 60 mg/m² × d1-5; and prednisone 40 mg/m² × d1-5. The historical control group (12 cases) received G-CSF subcutaneously (filgrastim) at the first restoration after the initial nadir of the peripheral WBC count. The experimental group (27 cases) received G-CSF during the steady rise of the WBC count (end of fluctuating after initial nadir). G-CSF was given in a single daily subcutaneous dose of 5 µg/kg until the final PBSC apheresis. When the peripheral WBC and mononuclear cell (MNC) counts reached 10 × 10⁹/L and 1.0 × 10⁹/L respectively, leukapheresis was carried out using the COBE Spectra blood cell separator. The results indicated that despite there was comparable treatment with alkylating agents between 2 groups, a significantly increased yield of CD34 positive cells was observed in the experimental group (26.4 × 10⁶/kg), as compared to the historical control group (3.1 × 10⁶/kg) (p = 0.0031). It is concluded that the appropriate timing for the use G-CSF mobilization after chemotherapy is important to increase the CD34(+) cell yield in auto-graft.

Cytogenetic abnormalities in bone marrow mesenchymal stem cells from patients with myelodysplastic syndrome / 中国实验血液学杂志

This study was aimed to investigate the cytogenetic characteristics of hematopoietic cells (HC) and bone marrow mesenchymal stoma cells (BMMSC) isolated from patients with myelodysplastic syndrome (MDS) and healthy individuals as normal controls, and to clarify whether HC and BMMSC are simultaneously involved and participate in pathogenesis and development of MDS. Both marrows of 22 newly diagnosed patients with MDS and 7 healthy individuals were collected; BMMSC were isolated and amplified by using established culture system, as well as were identified according to morphologic features and surface antigens detected by flow cytometry; the characteristics of BMMSC from MDS patients were analyzed; the BMMSC karyotyping analysis of MDS patients was performed by banding of HC and BMMSC with pancreatin-Giemsa technique (GTG) and in accordance with ISCN (2005) requirements; the cytogenetic characteristics of HC and BMMSC were compared. The results showed that in vitro culture system for isolation and amplification was successfully established. The light microscopy and flow cytometry confirmed that BMMSC possessed characteristics showing long and thin spindle form and expressing CD29, CD73, CD90 antigens, and unexpressing CD34, CD45 antigens. In BMMSC of 14 MDS patients (64%), the cytogenetic abnormalities were found usually involving the loss of chromosomal material (92%), among which clonal loss was observed in 7 cases (50%). The detection indicated a random loss of chromosomal material in significant proportion of BMMSC. A high proportion of chromosomal material random loss may be a marker of chromosomal instability in BMMSC of MDS patients, the detection also showed that completely consistent aberration types did not exist between HC and BMMSC. The aberrations were observed in 3 cases of MDS with normal karyotype of HC, its aberration rate (33%) was obviously lower than that in MDS patients with abnormal karyotypes (92%). It is concluded that the cytogenetic abnormalities exist in BMMSC of MDS patients, the unbalanced aberration of chromosomal materials may be a genetic instability marker of BMMSC. The difference of aberration types in BMMSC from HC suggests that genetic susceptibility of BMMSC and HC is similar, but no completely identical, which indicates the potential involvement of BMMSC in pathogenesis of MDS. Studying this potential role of BMMSC can be helpful to further understanding of MDS biological characteristics and provide the new approaches for diagnosis and therapy of MDS.

Biological behavior of stromal cell-derived factor-1 on migration, adhesion and apoptosis in different kinds of AML cell lines / 中国实验血液学杂志

The study was aimed to investigate the biological behavior of stromal cell-derived factor-1 (SDF-1) in migration, adhesion and apoptosis as well as the related signaling transduction pathways in different kinds of acute myeloid leukemia (AML) cell lines. The expression of surface molecules on AML (KG1a, ML1 and U937) cells were analyzed by flow cytometry. The cell adhesion was detected by MTT assay. The cell migration was checked by transwell assay. Bcl-xl was checked by immunoblotting after activation of phosphionositide-3 kinase (PI3K) in AML cells treated with SDF-1. The results indicated that the expressions of the surface molecules on AML (KG1a, ML1 and U937) cells were different. The list of the expression showed CD34 (KG1a = 95.6%, ML1 = 4.6%, U937 = 4.8%), CD45 (KG1a = 98.3%, U937 = 97.5%, ML1 = 17.8%), CXCR4 (ML1 = 85.4%, U937 = 43.6%, KG1a = 3.8%), ICAM (KG1a = 75.8%, U937 = 41.8% and ML1 = 46.3%). SDF-1 could not upregulate their expression, but could trigger the establishment of polarized morphology of the cells which expressed CXCR4 high. SDF-1 promoted ML1 and U937 cell adhesion to the stroma cells (HS5, HS27), stimulated PI3K in the cells. It was also confirmed that SDF-1 could increase the leukemic cell survival by stimulate this pathway. After addition of wortmaninn or PTX, the cell death increased. It is concluded that the SDF-1 increases the leukemic cell adhesion, migration and survival by stimulating the PI3K pathway. These functions can be depressed by the PI3K inhibitor and also the inhibitor of G protein as well.