Expression and role of MIG/CXCR3 axis in mantle cell lymphoma.

Mantle cell lymphoma (MCL) is a unique subtype of B-cell non-Hodgkin lymphoma with a generally aggressive and heterogeneous clinical course. Chemokines are one of the complex components in the tumor microenvironment (TME), and they play a vital role in tumor progression and metastasis. There is no information about the monokine induced by gamma interferon (MIG)/CXC chemokine receptor 3 (CXCR3) axis in patients with MCL. In the present study, we discovered that CXCR3 was highly expressed in MCL tissues and some cell lines including Maver, Z138, and Jeko-1, and significantly associated with clinical factors reflecting high tumor burden in MCL patients. Moreover, elevated serum MIG at diagnosis showed a close relationship with advanced disease and poor prognosis in MCL patients. Additionally, the role of CXCR3 in promoting the proliferation and inhibiting the apoptosis of primary MCL cells and Jeko-1 cells was validated by in vitro experiments. Further research indicated that the MIG/CXCR3 axis mediated MCL cell migration to the TME through the PI3K/AKT signaling pathway. Therefore, the MIG/CXCR3 axis might be a potential target with fewer off-target side effects than other targets in MCL.

Thymopentin enhances the generation of T-cell lineage derived from human embryonic stem cells in vitro.

Thymopentin is a group of biologically active peptide secreted mainly by the epithelial cells of thymic cortex and medulla. Whether it promotes T cells production from human embryonic stem cells(hESCs) in vitro remains an elusive issue. In the present study, we develop a novel strategy that enhances T-cell lineage differentiation of hESCs in collagen matrix culture by sequential cytokine cocktails treatment combined with thymopentin stimulation. We observed that approximately 30.75% cells expressed CD34 on day 14 of the cultures and expressed the surface markers of erythroid, lymphoid and myeloid lineages. The results of colony assays and gene expressions by RT-PCR analysis also demonstrated that hematopoietic progenitor cells (HPCs) derived from hESCs were capable of multi-lineage differentiation. Further study revealed that culturing with thymopentin treatment, the CD34(+)CD45RA(+)CD7(+) cells sorted from HPCs expressed T-cell-related genes, IKAROS, DNTT, TCRγ and TCRß, and T-cell surface markers, CD3, cytoplasmic CD3, CD5, CD27, TCRγδ, CD4 and CD8. The differentiated cells produced the cytokines including IFN-γ, IL-2 and TNF-α in response to stimulation, providing the evidence for T-cell function of these cells. In conclusion, thymopentin enhances T-cell lineage differentiation from hESCs in vitro by mimicking thymus peptide environment in vivo.

[The Relationship between Reducing of Peripheral Blood Treg in Primary Diffuse Large B-Cell Lymphoma Patients and Poor Prognosis].

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].

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 P<005). The CXCR3 expression in tumor cells significantly correlated with LDH level, ß2-MG level, Ki-67 index, MIPI score and the BM involvement (all P<0.05). But, there was no correlation between the CXCR3 expression and clinical stage, histomorphology (all P>0.05). 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.

[MicroRNA-223 Regulates the Differentiation of Human Embryonic Stem Cells to Dendritic Cells].

OBJECTIVE: 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. METHODS: 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. RESULTS: 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 CD34+CD45+, CD34+CD45+ and CD83+ in 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 105 cells, 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. CONCLUSION: MiR-223 promotes the differentiation of human ES cells to DC, probably through direet target to TGFBR3.

[Correlation between Expression of CD200 and Regulatory T Cells in Multiple Myeloma and Its Significance in Prognostic Stratification].

OBJECTIVE: 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. METHODS: 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), ß2 microglobulin (ß2-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. RESULTS: 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 CD200+ group was significantly higher than that of in CD200- group (P=0.032). There was no significant difference in the expression of CD20, CD56 and CD19 between 2 groups. The level of Hb in CD200+ group was significantly lower than that in CD200- group (P=0.035). The level of ß2-MG in CD200+ group was significantly higher than that in CD200- group (P=0.013). There was no significant difference in the level of LDH between 2 groups. In CD200+ group, 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 CD200+ group was significantly higher than that of in CD200- group (P=0.043). The content of Tregs was positively correlated with the expression of CD200 (r=0.743, P=0.044). Overall survival in CD200- group was significantly higher than that in CD200+ group (P=0.036). Progression-free survival in CD200- group was a bit higher than that of in CD200+ group, but it wasn't statistically significant. CONCLUSION: 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.

[Expression and Subcellular Distribution of Costimulatory Molecules B7-H1,B7-H3 and B7-H4 in Human Hematologic Malignancy Cell Lines].

OBJECTIVE: To investigate the expression and subcellular distribution of costimulatory molecules B7-H1, B7-H3 and B7-H4 in human hematologic malignancy cell lines. METHODS: 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. RESULTS: 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. CONCLUSION: 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.

[Clinical characteristics and therapeutic efficacy of immunoglobin D multiple myeloma].

This study was aimed to evaluate the clinical characteristics of immunoglobin D multiple myeloma (IgD MM) and its curative efficacy. The clinical data of 15 cases of newly diagnosed IgD MM from April 1993 to June 2013 were analyzed retrospectively. Among 15 cases received induction treatment, the traditional chemotherapy was carried out in 9 cases, bortezomib-based therapy was performed in 6 cases. The diagnostic criteria and disease response criteria of MM were based on International Myeloma Working Group (IMWG) criteria,survival time was analyzed by using Kaplan-Meier method, the median age of patients was 57 years (range 40-72), the ratio of male to female was 2:1, the MM patients in Durie-Salmon stage III accounted for 100% (15/15) , the MM patients with λ light chain accounts for 80% (12/15) , with bone lesion 86.7% (13/15), with pleural effusion 26.7% (4/15) , with renal impairment (RI) 86.7% (13/15) ,with anemia 93.3% (14/15) , with serum album<35 g/L 26.7% (4/15). The median creatinine clearance rate (Ccr) of patients was 23.1 (6-44) ml/min, and median hemoglobin level was 82 (43-131) g/L. The results showed that in followed-up 11 cases, 8 cases died, 3 cases survived; the average duration of follow-up for these cases was 20 (0.5-138) months, the median progression-free survival time (PFS) was 7 (95%CI4.6-9.4) months, and the median overall survival (OS) time was 15 (95%CI6.6-27.4 ) months. Compared traditional chemotherapy with bortezomib regimen therapy,median OS time was 17 (95%CI6.1-28.0) months vs 15 (95%CI 0.0-33.3) months (P = 0.90) . Assessable curative effect of 14 cases was as follows: CR 33.3% (5/15); VGPR 13.3% (2/15); PR 20% (3/15); SD 20% (3/15); PD 6.7% (1/15). It is concluded that IgD MM is a rare type of MM and has a poor prognosis. Bortezomib may be beneficial for some patients with extramedullary infiltration. Autologous hematopoietic stem cell transplantation may improve survival time.

[Clinical characteristics and prognostic factors of 62 cases with Hodgkin lymphoma].

OBJECTIVE: To investigate the clinical characteristics, pathologic types and prognostic factors of Hodgkin lymphoma (HL). METHODS: Retrospective analyses of clinical data of 62 HL patients, including 45 from Peking University third hospital and 17 from Shanxi Dayi hospital. RESULTS: Sixty-two HL patients accounted for 5.66% of the total newly diagnosed patients with lymphoma (45/795) in the period with a ratio of male:female as 2.1:1. All the pathological types were classical Hodgkin lymphoma (cHL), including 33 cases (53.2%) of nodular sclerosis type (NS) and 19 cases (30.6%) of mixed cell type (MC). Of the 56 cases followed up, the total responsive rate was 92.9% , and 2- , 3- and 5- year overall survival (OS) rates were 97.7%, 92.2% and 80.5% respectively with the median OS as 45 (8-184) months. Analyses of the prognostic factors indicated that the involvement of spleen(P=0.042)and German Hodgkin Study Group(GHSG) risk score (P=0.002) were poor factors with statistic significance. CONCLUSION: HL is one of the curable malignant tumors, 5-year OS rate was 80.5% in this group. After recurrence, part of patients developed slowly and could be salvaged with second-line therapy with high remission rate. GHSG scoring system and involvement of spleen could predict the prognosis in HL setting.

[Analysis of clinical features and prognostic factors in 59 cases of follicular lymphoma].

This study was aimed to explore the clinical features and related prognostic factors of follicular lymphoma (FL). The total of 59 newly diagnosed FL patients were analyzed retrospectively in term of age, sex, clinical manifestation, laboratory data, pathological examination, clinical stage and so on, so as to find out the related prognostic factors. The results showed that the median age of the 59 patients was 55 years, among them the male-female ratio was 1: 1.36. Among the 64.4% cases with the involvement outside lymphonode , the bone marrow involvement accounted for 28.8%, while the involvement of gastrointestinal tract and skeleton was in the second place. The rate of BCL-2/JH gene rearrangement was 30.3%, III-IV of clinical stage accounted for 71.2% in these FL patients. During the follow up of the 56 patients, OR and CR rates were 96.4% and 80.3% respectively, 5 year-and 10 year overall survival rates were 88.2% and 74.1% respectively, the relapse rate was 33.9%. Univariate analysis showed that the age, the longest diameter of involved node, gastrointestinal involvement, serum lactate dehydrogenase, ECOG, IPI, aaIPI, FLIPI, FLIPI2 and whether initial therapy could achieve CR or not were influencing factors of prognosis for FL patients. Multivariant analysis showed that the gastrointestinal involvement, IPI, aaIPI and whether initial therapy could achieve CR or not were independent prognostic factors for OS. The relative mortality risk of patients without CR was 70 times of patient with CR. It is concluded that the newly diagnosed FL is more observed in middle to old aged females, displaying frequent bone marrow involvement, less frequent BCL-2/JH gene rearrangement, III-IV of clinical staging, and high relapse rate, but longer survival time. The gastrointestinal involvement, IPI, aaIPI and whether CR can be achieved or not after initial therapy are independent prognostic factors for FL patients.