The Relationship between Reducing of Peripheral Blood Treg in Primary Diffuse Large B-Cell Lymphoma Patients and Poor Prognosis / 中国实验血液学杂志

OBJECTIVE@#To explore the relationship between Treg cells level in peripheral blood and prognosis of patients with diffuse large B-cell lymphoma (DLBCL).@*METHODS@#The percentage and absolute value of Treg cells in peripheral blood of DLBCL patients were detected by flow lytometry, and their correlation to prognosis was analyzed by survival analysis. The absolute count of Treg cells was detected by using maximally selected Log-rank statistic, and it was used as cutoff point to distinguish difference survival. The new group of Treg based on cutoff point was combined with age, sex, pathological subtype, risk stratification, treatment plan, and other indicators to include in the single factor survival analysis of Kaplan-Meier. Finally, the COX proportional risk model was used to verify the effect of the above indicators on progression-free survival.@*RESULTS@#The absolute count of Treg cells in DLBCL patients was significantly lower in the disease progressed group than those in the remission group. The cutoff point of absolute value of the Treg cell was 19 cells /μl. The absolute count of Treg cells was an independent prognostic factor of the risk stratification.@*CONCLUSION@#At the beginning of diagnosis, the reduction of the absolute count of Treg cells in peripheral blood of DLBCL patients show a poor prognosis.

Expression and Clinical Significance of CXCR3 in Mantle Cell Lymphoma / 中国实验血液学杂志

OBJECTIVE@#To investigate the expression and clinical significance of chemokine receptor CXCR3 in mantle cell lymphoma (MCL).@*METHODS@#Flow cytometry was used to detect CXCR3 in lymph nodes and extranodal tissues in 25 newly diagnosed MCL patients. The correlation of the expression of CXCR3 level with clinical features and prognostic factors was analyzed.@*RESULTS@#Twenty-five tumor submitted specimens all expressed CXCR3 at varied degrees. The expression levels of CXCR3 in lymph nodes (LN) and bone marrow (BM) were higher than those in peripheral blood (PB), and the expression intensity in BM positively correlated with the involved tumor numbers. The absolute values of lymphocytes and hemoglobins level in PB of CXCR3high group were significantly lower than those in CXCR3low group (all P0.05). The overall response rate (ORR) in CXCR3low group was significantly higher than that in CXCR3high group (P=0.001). The expression level of CXCR3 in MCL cells of the effective group was significantly lower than that before treatment (P=0.038), and the CXCR3 expression in the ineffective group was significantly higher than that before treatment (P=0.002). After following up, it was found that the 3-year overall survival (OS) time and progression-free survival (PFS) time in CXCR3high group were significantly shorter than those in CXCR3low group (all P<0.05).@*CONCLUSION@#The expression level of CXCR3 in MCL closely relates with early metastasis and prognosis. CXCR3 can be used as one of the indicators for clinical efficacy and prognosis evaluation.

Relationship of the Changes of Peripheral Blood Immuno-Cell Subsets with the Prognosis of B Cell Lymphoma Patients / 中国实验血液学杂志

OBJECTIVE@#To investigate the relationship of T lymphocyte subsets, B lymphocytes and NK cells with the genesis, progression and prognosis of B cell lymphoma.@*METHODS@#The levels of T lymphocyte subsets, B lymphocytes and NK cells in peripheral blood of healthy control group and B cell lymphoma group were detected by flow cytometry (FCM). The clinical data were collected, and the relationship of these immune indexes with the general conditions, laboratory indexes, curative effect and prognosis were analyzed.@*RESULTS@#Forty-four patients entolled in this study including 24 male and 20 females with the median age of 57 years old (17-82 years), all the patients were the first visit to our hospital and diagnosed. The total counts of lymphocytes, T, B and NK cells in the peripheral blood of patients with the first treatment of B-cell lymphoma were significantly lower than those in healthy controls, and the ratio of CD3HLA-DR activated T cells was significantly higher than that of healthy controls ( 61.5 /μl was higher than that in patients with low level of NK cells.@*CONCLUSION@#The level of total lymphocytes, total T cells, total B cells, NK cells and advanced activated T cells in the patients with B cell lymphoma were significantly different from those in normal subjects. Total count of lymphocytes, T cells, B cells, CD4 cells and NK cells in peripheral blood are important prognostic indicators for BCL. The ECOG score and β2-microglobulin level are independent risk factors for prognosis. The NK cell level and FAC-1 are independent protective factors for the prognosis of B cell lymphoma.

MicroRNA-223 Regulates the Differentiation of Human Embryonic Stem Cells to Dendritic Cells / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To establish a new inducing system for differentiation of human embryonic stem (ES) cells to dendritic cells (DC), and further explore how microRNA-223 (miR-223) regulates DC differentiation from ES cells.</p><p><b>METHODS</b>Human ES cells were cultured on plates coated by IV type collagen and differentiated into hematopoietic stem/progenitor cells, common myeloid progenitor cells and DC step by step via adding different hematopoietic growth factors. The differentiated cells were identified by morphology, flow cytometry and hematopoietic colony forming unit (CFU) assays. Human ES cells were transfected with lentiviral vectors to overexpress miR-223 or inhibit miR-223 expression, then initiated the differentiation to DC. The differentiated cells at the different miR-223 levels were compared by the numbers of hematopoietic CFU and the expressions of specific surface markers. Dual-luciferase reporter assay was performed to test whether miR-223 directly targets TGFBR3.</p><p><b>RESULTS</b>Human ES cells were successfully induced into DC as the percentage of CD83 was approximately 82%, and the expression of miR-223 was up-regulated during the whole process. Supplementing miR-223 level using synthetic miR-223 mimics improved the proportions of CD34CD45, CD34CD45and CD83in differentiated cells, which were significantly higher than those in synthetic miR-223 inhibitor group and negative control (P<0.05). The expressions of cell makers at each differentiated phase in miR-223 inhibitor group were significantly lower than those in negative control (P<0.05). The differentiated cells in miR-223 mimics group showed approximately 759 CFUs per 10cells, which was significantly higher than that in others (P<0.05). Compared with negative control, miR-223 substantially inhibited the luciferase activity of Tgfbr3 3'UTR construct (by 37%) (P<0.05). In addition, the luciferase activity of the mutant construct was significantly higher than that of the WT construct in the presence of miR-223 mimics (P<0.05). With DC mature, the protein level of TGFBR3 gradually decreased using miR-223 mimics, and increased in miR-223 inhibitor group due to the suppression of the endogenous miR-223.</p><p><b>CONCLUSION</b>MiR-223 promotes the differentiation of human ES cells to DC, probably through direet target to TGFBR3.</p>

Comparison of Criminal Characteristics in Depression Patients and Schizophrenics with Homicide Behavior / 法医学杂志

OBJECTIVES@#To explore the criminal characteristics of forensic psychiatry expertise in depression patients and schizophrenics with homicide behavior.@*METHODS@#A total of 40 depression （depressive episode） patients and 50 schizophrenics with homicide behavior were randomly assigned into the study group and control group, respectively. Data of demographic and criminal characteristic of the two groups were collected by a self-designed questionnaire, and then were compared.@*RESULTS@#There were no statistical differences in age, education level and career between study and control groups （P>0.05）. Compared with the control group, the victims in the study group were mainly the patient's children and parents, and most offenders had suicidal behavior after homicide （70%）. In study group, the motives of crime were mainly extended suicide and indirect suicide, and most offenders had attempted suicide （85%） and diminished capacity of criminal responsibility （70%）, which in control group had no capacity of criminal responsibility （56%）. Except for criminal site, there were statistical differences in other criminal characteristics between two groups （P<0.05）.@*CONCLUSIONS@#There are different criminal characteristics between depression patients and schizophrenics with homicide behavior in forensic psychiatry, and these characteristics should be considered when these two diagnoses are distinguished in forensic psychiatry expertise.

Inductive Effect of 5-Azacitidine on Apoptosis of Multiple Myeloma Cell lines and Its Mechanism / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the response of multiple myeloma (MM) cells to 5-azacitidine and its possible mechanisms.</p><p><b>METHODS</b>Two multiple myeloma-derived cell lines U266 and H929 were used in this study. The cell proliferation and apoptosis were assessed by CCK-8, flow cytometry and acridine orange staining. The cell cycle was assessed by flow cytometry. The expression of BCL-2, BAX was assessed by RT-PCR. The expressions of caspase-3 and p-ERK1/2 were assessed by Western blot.</p><p><b>RESULTS</b>The BCL-2/BAX ratio decreased, the activity of caspase-3 and p-ERK1/2 increased, the cell cycle was arrested in G2/M phase after treatment with 5-azacitidine. The 5-azacitidine inhibited proliferation in a dose-dependent manner.</p><p><b>CONCLUSION</b>5-azacitidine exerts apoptosis-inducing and grow-inhibiting effects on MM cell lines, its mechanism may be related with the decrease of BCL-2/BAX ratio, caspase-3 activation and the arrest of cell cycle.</p>

Correlation between Expression of CD200 and Regulatory T Cells in Multiple Myeloma and Its Significance in Prognostic Stratification / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To study the correlation between expression of CD200 and regulatory T cells (Tregs) in multiple myeloma(MM) patients and to explore its significance in prognostic stratification.</p><p><b>METHODS</b>CD200 and other immunophenotypes, including CD38, CD138, CD56, CD19, CD20, CD117, cytoplasm light chain Kappa and Lambda in bone marrow samples, and Tregs in peripheral blood were detected by flow cytometry from 78 newly diagnosed MM patients. Serum concentrations of hemoglobin(Hb), β2 microglobulin (β2-MG) and lactate dehydrogenase (LDH) were detected, respectively. The new risk stratification of patients was carried out according to international stage system (ISS) and cytogenetic characteristics. The correlation between expression of CD200 and Tregs in MM patients was analyzed and their differences in prognosis were compared.</p><p><b>RESULTS</b>The positive rate of CD200 expression was 71.79% in 78 patients (56/78). The expression of CD200 in sex and age of patients was no significant different. The expression of CD117 in CD200group was significantly higher than that of in CD200group (P=0.032). There was no significant difference in the expression of CD20, CD56 and CD19 between 2 groups. The level of Hb in CD200group was significantly lower than that in CD200group (P=0.035). The level of β2-MG in CD200group was significantly higher than that in CD200group (P=0.013). There was no significant difference in the level of LDH between 2 groups. In CD200group, 17 patients were in stageⅠ, accounting for 58.62% (17/29), 30 patients were in stageⅡ, accounting for 75% (30/40), 9 patients were in stage Ⅲ, accounting for 100% (9/9). With the increase of CD200 expression intensity, the risk of prognostic stratification went up (P=0). The brighter CD200 expressed, the worse the prognosis was. The percentage of Trges in CD200group was significantly higher than that of in CD200group (P=0.043). The content of Tregs was positively correlated with the expression of CD200 (r=0.743, P=0.044). Overall survival in CD200group was significantly higher than that in CD200group (P=0.036). Progression-free survival in CD200group was a bit higher than that of in CD200group, but it wasn't statistically significant.</p><p><b>CONCLUSION</b>The expression of CD200 positively correlates with the percentage of Tregs in MM patients. It is an important factor of poor prognosis and a reliable indicator for the evaluation by clinical efficacy in MM patients.</p>

Expression and Subcellular Distribution of Costimulatory Molecules B7-H1,B7-H3 and B7-H4 in Human Hematologic Malignancy Cell Lines / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the expression and subcellular distribution of costimulatory molecules B7-H1, B7-H3 and B7-H4 in human hematologic malignancy cell lines.</p><p><b>METHODS</b>The expression and subcellular distribution of B7-H1, B7-H3 and B7-H4 in 13 human hematologic malignancy cell lines were determined by RT-PCR, qPCR, Western blot and flow cytometry, the peripheral blood mononuclear cells (PB MNC) of 12 volunteers were used as control.</p><p><b>RESULTS</b>The mRNA of B7-H1, B7-H3 and B7-H4 was widely expressed in PB MNC and hematologic malignancy cell lines, with a lower level of B7-H4. The mRNA expression of 3 molecules was highest in Maver, Z138, and HL-60, respectively, while among them the B7-H3 and B7-H4 had no expression in CZ1. The nuclear and cytoplasmic protein of 3 costimulatory molecules abnormally overexpressed only in hematologic malignancy cell lines, with the highest level in U937, Z138, and Raji, respectively, while the B7-H3 and B7-H4 had no expression in CZ1. There were differences among mRNA expression, nuclear and cytoplasmic protein expression of 3 molecules in cell lines derived from the same type of tumor, but the differences of expression in mRNA and protein levels were not exactly the same. The B7-H3 expression abundance in membrane localization was higher in U937, Maver and Z138, while the membrane protein of B7-H1 and B7-H4 had no or low expression in 13 cell lines.</p><p><b>CONCLUSION</b>The mRNA expression of costimulatory molecules B7-H1, B7-H3 and B7-H4 can be widely detected. The protein level of 3 costimulatory molecules abnormally overexpressed only in hematologic malignancy cell lines, moreover the subcellular localizations mostly was found in nucleus and cytoplasm, while the membrane protein expresses in low level or had no expression. There are differences among the expression of 3 molecules in cell lines derived from the same type of tumor.</p>

Construction and in vitro study of eukaryotic expression vector carrying GITRL and IL-21 gene / 中国实验血液学杂志

The aim of this study was to construct the eukaryotic expression vector carrying glucocorticoid-induced tumor necrosis factor receptor ligand (GITRL) and interleukin-21 (IL-21) gene for transfection into chronic myeloid leukemia (CML) dervied dendritic cell (DC), so as to provide an effective platform for exploring the function of target gene in CML. The recombinant eukaryotic expression vector was transfected to the purified DC by Liposome-mediated method, the interleukin-2 (IL-2) and Interferon-γ (IFN-γ) expression of transfected DC were analyzed by ELISA. Further, the transfected DC with purified NK were mixed and cultured to be DC-CIK, the lactate dehydrogenase release assay was performed to measure the killing activity of DC-CIK. The results indicated that the sequence of cloned target gene was same as that in GenBank. The size of endonuclease products by restriction enzyme were same as the predict one. The concentration of Interleukin-2 (IL-2) and interferon-γ (IFN-γ) in transfected DC all increased. The NK kill activity became stronger while induced by transfected DC. It is concluded that DC transfected by IL-21 and GITRL gene has the ability of self-activation, up-regulate cytokine secretion. Further, the results would be help to provide the theoretical evidence of advanced immunotherapy for treatment of CML patients who showed no reaction to tyrosine kinase inhibitor.

Immunophenotype characteristics of patients with multiple myeloma in different risk stratification / 中国实验血液学杂志

This study assessed the immunophenotype characteristics of newly diagosised patients with multiple myeloma (MM) in different risk stratification in order to find the relationship between the immunophenotype and prognosis of MM. The expressions of CD45, CD38, CD138, CD56, CD19, CD117, CD13, CD20, CD22, CD34, Kappa, Lambda in bone marrow samples from 62 newly diagnosed MM patients were detected by using flow cytometric multiparametric direct immunofluorescence technique, CD45/SSC and CD38/SSC combination gating, then the immumophemotypic characteristics of patients in different risk stratification groups were analyzed and compared. The new risk stratification of all patients was carried out according to ISS stages (Interuational Staging System) and cytogenetic characteristics. The results indicated that all the malignant plasma cells commonly expressed CD38 (100%) and CD138 (100%); CD19⁺ (6.5%) ,CD45⁺ (22.6%), CD56⁺ (59.6%) and monoclonal light chain (82%); but the expressions of CD117⁺ (27.4%) , CD13⁺ (17.7%) , CD20⁺ (16.1%) were diverse. According to risk stratification, it is found that the standard-risk and high-risk groups had lower expression of CD56 (P = 0.022) and higher expression of CD117 (P = 0.011), compared with the low-risk group. It is concluded that the immunophenotype of MM is heterogeneity, the lower expression of CD56 and higher expression of CD117 may be associate with poor prognosis.

Effects of different extracellular matrix on differentiation of human embryonic stem cells into hematopoietic progenitor cells / 中国实验血液学杂志

This study was purposed to establish a new inducing system for differentiation of human embryonic stem cells into hematopoietic progenitor cells and to explore the effect of different extracellular matrices (DEM) on production of hematopoietic cells. The 3 kinds of extracellular matrices-matrigel, fibronectin and IV type collagen (collagen IV) were chosen to package cultured plates, the direct adherent culture on extracellular matrix was used, and the hematopoietic growth factors were added into cultured plates to induce the differentiation of human embryonic stem cells into hematopoietic progenitor cells. The hematopoietic colony forming unit assay was used to determine the yielded colony forming cells, the flow cytometry and real-time quantitative PCR were used to detect the expression of markers specific to hematopoiesis and the effect of 3 extracellular matrices on production of hematopoietic progenitor cells was compared. The results showed that after being induced for 14 days, the total yield of colony forming cells, the ratio of CD34(+) cells and the expression level of SCL and CD34 on collagen IV were significantly higher than those on matrigel and fibronectin groups (P < 0.05). It is concluded that human embryonic stem cells can efficiently differentiate into hematopoietic progenitor cells by direct adherent culture on extracellular matrices, and the collagen IV can improve the hematopoietic differentiation of human embryonic stem cells.

Roles of microRNAs in lymphocytopoiesis and lymphoid malignancies - review / 中国实验血液学杂志

microRNAs (miRNAs) are small molecular non-coding RNA with 21-25 nucleotides in a variety of eukaryotic systems, and regulate gene expression at the post-transcriptional level by degrading or translational repressing target messenger RNA (mRNA). Many studies have showed the roles of miRNAs in normal lymphocytopoiesis, giving an interpretative key to the aberrant expression observed in human lymphoid malignancies. The recent advances of understanding the roles of miRNAs in lymphoid malignancies show that miRNAs as tumoral biomarkers can effectively be used for diagnosis, prognosis, and prediction of response to therapy. This review focuses the roles of miRNA in development and differentiation of lymphocytes and the relation of miRNA to lymphoid malignancies.

The application of evaluation tools for criminal responsibility in forensic psychiatric expertise / 法医学杂志

Criminal responsibility is divided into three types: full criminal responsibility, diminished criminal responsibility and criminal irresponsibility in China. In forensic psychiatric expertise, doctors often have different opinions about the responsibility in a given case because of lacking objective criteria. The evaluation of criminal responsibility is always unresolved problem in forensic psychiatric expertise. Application of these evaluation tools in forensic psychiatric expertise were reviewed in this article. The value of the tools were still controversial in the reliability and validity, but it is clear that these tools have the positive roles in ensuring the standardization and the uniformity of the forensic investigation.