Clinical Efficiency of Rituximab Combined with Autologous Hematopoietic Blood Stem Cell Transplantation for Treatment of CD20 / 中国实验血液学杂志

METHODS@#From January 2005 to December 2013, 83 patients with refractory/recurrent CD20@*RESULTS@#All the patient achieved complete response. The median follow.up time was 39 months. Both the two groups collected peripheral blood stem cells successfully, and had no difference in hematopoietic reconstitution time. Three patients in treatment group and six patients in control group relapsed and the three year overall survival and EFS in treatment group was significantly higher than that in control group, that is（93.0% vs 73.1%, P=0.037） and （89.5% vs 65.4%, P=0.034）, respectively. Subgroup analysis showed that: compared with the treatment group in which using R in the whole courses(before and after transplantation, and collection of stem cells) was superior to the control group in both OS and EFS, with the OS 97% vs 87.5% (P>0.05) and EFS 97% vs 76.2% (P=0.05) respectively. While stratified by the different courses of rituximab, the OS was 88.9% (1-2 courses, 9 cases), 93.1% (3-4 courses, 29 cases), 94.7%(more than 5 courses,19 cases), and EFS was 77.8%, 89.7% and 94.7%, respectively.@*CONCLUSION@#For the patients with refractory/recurrent CD20

Silencing HIF-1α reduces the adhesion and secretion functions of acute leukemia hBMSCs

Hypoxia inducible factor-1α (HIF-1α) is an important transcription factor, which plays a critical role in the formation of solid tumor and its microenviroment. The objective of the present study was to evaluate the expression and function of HIF-1α in human leukemia bone marrow stromal cells (BMSCs) and to identify the downstream targets of HIF-1α. HIF-1α expression was detected at both the RNA and protein levels using real-time PCR and immunohistochemistry, respectively. Vascular endothelial growth factor (VEGF) and stromal cell-derived factor-1α (SDF-1α) were detected in stromal cells by enzyme-linked immunosorbent assay. HIF-1α was blocked by constructing the lentiviral RNAi vector system and infecting the BMSCs. The Jurkat cell/BMSC co-cultured system was constructed by putting the two cells into the same suitable cultured media and conditions. Cell adhesion and secretion functions of stromal cells were evaluated after transfection with the lentiviral RNAi vector of HIF-1α. Increased HIF-1α mRNA and protein was detected in the nucleus of the acute myeloblastic and acute lymphoblastic leukemia compared with normal BMSCs. The lentiviral RANi vector for HIF-1α was successfully constructed and was applied to block the expression of HIF-1α. When HIF-1α of BMSCs was blocked, the expression of VEGF and SDF-1 secreted by stromal cells were decreased. When HIF-1α was blocked, the co-cultured Jurkat cell’s adhesion and migration functions were also decreased. Taken together, these results suggest that HIF-1α acts as an important transcription factor and can significantly affect the secretion and adhesion functions of leukemia BMSCs.

Intracerebral infiltration by monoclonal plasmacytoid cells in Waldenstrom's macroglobulinemia: a case report and review of the literature / 中华血液学杂志

<p><b>OBJECTIVE</b>To sum up the clinical experience of the diagnosis and treatment of intracerebral infiltration by monoclonal plasmacytoid cells in Waldenstrom's macroglobulinemia(Bing-Neel syndrome).</p><p><b>METHODS</b>The clinical data of the diagnosis and treatment of a case of Bing-Neel syndrome was analyzed.</p><p><b>RESULTS</b>A 56-year-old male was diagnosed as Waldenstrom's macroglobulinemia one year ago, and presented with persistent headache during the treatment period. Magnetic resonance imaging showed a high intensity area on T2-weighed images in the right frontal lobe which was well enhanced by gadolinium-diethylenetriaminepenta-acetic acid. Infiltration of neoplastic cells was confirmed by biopsy. Immunohistochemical examination showed that mature plasmacytoid cells in the cerebral parenchyma were immunoglobulin M positive.</p><p><b>CONCLUSION</b>Infiltration in CNS (Bing-Neel syndrome) is uncommon in Waldenstrom's macroglobulinemia. As there is no effective therapy for this Bing-Neel syndrome, combination of radiation and chemotherapy should be considered for this situation.</p>

Effect of the NHE-1-specific inhibitor DMA on pHi, proliferation and apoptosis of HL-60/ADM cells in vitro / 中国实验血液学杂志

The aim of this study was to evaluate the effect of dimethyl amiloride (DMA), a specific inhibitor of Na(+)/H(+) exchanger-1 (NHE-1), on intracellular pH value (pHi), proliferation and apoptosis of HL-60/ADM cells in vitro. After treatment with DMA at different doses, pHi of HL-60 and HL-60/ADM cell lines were determined by using pH-sensitive fluorescence dye BECEF-AM; the rate of growth inhibition of cells was detected with MTT assay; cell cycle was detected by flow cytometric DNA analysis; cell apoptosis was observed with terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). The results showed that pHi in HL-60/ADM cells was higher than that in HL-60 cells. After treatment with DMA at different doses, pHi decreased, the rate of growth inhibition and the rate of apoptotic cells in HL-60/ADM cells were all higher than those in HL-60 cells. Meanwhile, after treatment with DMA during 100 micromol/L to 150 micromol/L, the increase amplitude of G(0)/G(1) phase cells and the decrease amplitude of S + G(2)/M cells in HL-60/ADM cells were higher than those in HL-60 cells. It is concluded that by causing intracellular acidification, the NHE-1-specific inhibitor DMA inhibits proliferation of HL-60/ADM cells and induces apoptosis of HL-60/ADM cells, and the degree of this growth inhibition of HL-60/ADM cells is higher than that of HL-60 cells.

In vitro effect of all-trans retinoic acid on cell adhesion molecule expression and adhesion capacity of bone marrow stromal cells in patients received peripheral blood stem cell transplantation / 中国实验血液学杂志

The aim of this study was to evaluate the effect of all-trans retinoic acid (ATRA) on cell adhesion molecule expression and adhesion capacity of bone marrow stromal cells (BMSC) in patients after conditioning treatment for peripheral blood stem cell transplantation (PBSCT). BMSC of 27 patients before and after conditioning treatment for PBSCT were cultured in vitro. After treated with ATRA at 0.01, 0.1, or 1 micromol/L, expression of intercellular adhesion molecule-1 (ICAM-1) protein and vascular adhesion molecule-1 (VCAM-1) protein were detected by flow cytometry, and soluble ICAM-1 (sICAM-1) protein was determined by using radioimmunoassay. Then BMSC was co-cultured with CD34+ cells, and adhesion rate of BMSC to CD34+ cells was measured. The results showed that after pretreatment with conditioning regimen for PBSCT, the expressions of ICAM-1 and VCAM-1 proteins in BMSC and the expression level of sICAM-1 protein in supernatant of BMSC culture were down-regulated, and the adhesion rate of BMSC to CD34+ cells was decreased, after administration of ATRA, the expression of ICAM-1 protein in BMSC, sICAM-1 protein in culture medium and adhesion rate of BMSC to CD34+ cells all increased significantly, but expression of VCAM-1 protein changed no significantly. It is concluded that the ATRA can partly restore adhesion function of BMSC injured by pretreatment for PBSCT and contribute to hematopoietic reconstitution.

Influence of anti-CXCR4 monoclonal antibody 12G5 on killing effect of cytosine arabinoside to HL-60 cell / 中国实验血液学杂志

This study was aimed to explore the influence of anti-CXCR4 monoclonal antibody 12G5 on killing effect of cytosine arabinoside (Ara C) to HL-60 cell, and to assess its therapeutic value in marrow residual disease. HL-60 cells were cultured and co-cultured with leukemic stromal cells, and SDF-1 activity was inhibited with 10 microg/ml 12G5, then, killing effects of Ara C on HL-60 cells were investigated by MTT and morphology assay. Curves by MTT assay revealed that in the test group of 20 microg/ml Ara C, A(540) values decreased slowly but straightly, however, in control group A(540) values decreased markedly for the first two days, and increased from day 3 or 4. In the test group of 40 microg/ml Ara C, although increasing at constricted range of 7 - 9 days, A(540) values decreased in whole observing period of 12 days, while in control group A(540) values decreased markedly at day 0-3, and increased from day 4. Furthermore, two curves go across each other at day 5, and continue the increasing tendency. Morphology results showed that in both treated groups, the number of HL-60 cell decreased markedly and increased gradually in control group, but just contrary to test group. It is concluded that 12G5 may weaken the killing effect of Ara C on HL60 cell in earlier period, but reinforce the total killing effect and delay the occurrence of drug resistance simultaneously. Thus 12G5 has the therapeutic potential on marrow residual disease.

Expression of stromal cell derived factor-1(SDF-1) and its receptor CXCR4 in hematologic malignancies / 中国实验血液学杂志

The aim was to analyze the expression level of stromal cell derived factor-1 (SDF-1) and its functional chemokine receptor CXCR4 in the patients with hematologic malignant tumor and their clinic significance. 28 patients with hematologic malignant tumor and 12 normal controls were chosen to be experimental objects. CXCR4 expressed on the cell membrane in bone marrow was enumerated by flow cytometry and serum level of SDF-1 was determined by ELISA assay. The result showed that the expression of SDF-1 and CXCR4 in hematologic malignant tumor were higher than that in normal controls, and the expression levels of two molecules were correlated. What is more, the different hematologic malignant tumor had different CXCR4 expression. In conclusion, the high expression of SDF-1 and CXCR4 in serum and bone marrow cells can be used as detective factors to hematologic malignant tumor. A correlation exists between the high expression of CXCR4 and the infiltration of hematologic malignant cells.

Effects of RNA interference inhibiting SDF-1 expression in bone marrow stromal cells on the proliferation and apoptosis of co-cultured Jurkat cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To study the effects of RNA interference inhibiting stromal cell derived factor-1 (SDF-1) expression on the proliferation and apoptosis of co-cultured Jurkat cells.</p><p><b>METHOD</b>Inhibition of SDF-1 expression by RNA interference (RNAi) was achieved by transferring SDF-1 specific short hairpin RNA (shRNA) expressing plasmid into cultured human acute leukemic bone marrow stromal cells. Resistant clones were obtained by G418 selection (group A). The concentration of SDF-1 protein in culture supernatant was detected by enzyme-linked immunosorbent assay (ELISA). The population double time (PDT), cell cycles, apoptosis rates and the expressions of PCNA, Bcl-2/Bax, Fas/FasL of co-cultured Jurkat cells were detected by cells counting, flow cytometry. TdT-mediated dUTP nick-end labelling (TUNEL) and immunocytochemistry (ICC), respectively. The un-transfected acute leukemic (group B) and normal (group C) bone marrow stromal cells were taken as controls.</p><p><b>RESULTS</b>The content of SDF-1 protein in supernatant of group A\[(384 +/- 41) pg/ml] was significantly lower than that in group B[(2474 +/- 271) pg/ml] or group C[(1324 +/- 154) pg/ml]. As group A compared with group B and group C, the PDT of co-cultured Jurkat cells was prolonged (group A: 42 h, vs group B: 29 h, group C: 33 h), and G(0)/G(1) stage cells increased [group A: (28.47 +/- 2.39)%, vs group B: (19.43 +/- 2.80)%, group C: (27.15 +/- 2.07)%], S stage cells decreased [group A: (25.57 +/- 1.90)%, vs group B: (74.48 +/- 3.23)%, group C: (60.99 +/- 2.33)%], G(2)/M stage cells increased [group A: (45.96 +/- 3.24)%, vs group B: (6.09 +/- 1.96)%, group C: (11.86 +/- 1.98)%], the apoptosis rate increased [group A: (15.2 +/- 0.8)%, vs group B: (5.4 +/- 0.7)%, group C: (9.5 +/- 0.4)%], and the expressions of PCNA, Bcl-2, Fas decreased; whereas the expressions of Bax and FasL were increased.</p><p><b>CONCLUSION</b>The inhibition of SDF-1 expression in bone marrow stromal cells inhibits the proliferation and promotes the apoptosis of co-cultured Jurkat cells.</p>

Effects of inhibiting SDF-1 expression by RNA interference on adhesion and drug sensitivity of Jurkat cells co-cultured with bone marrow stromal cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To observe the effects of inhibiting stromal cell derived factor-1 (SDF-1) expression by RNA interference (RNAi) on adhesion and drug sensitivity of Jurkat cells co-cultured with bone marrow stromal cells.</p><p><b>METHODS</b>SDF-1 specific short hairpin RNA (shRNA) expressing plasmid was transferred into cultured human acute leukemic bone marrow stromal cells, positive clones were isolated by screening G418 resistance (Group A) , SDF-1 protein level in culture supernatant was detected by enzyme-linked immunosorbent assay (ELISA). The adhesion rates to bone marrow stromal cells layer and the drug sensitivity to doxorubicin of co-cultured Jurkat cells were detected by cell counting and MTT assay, respectively. The un-transfected bone marrow stromal cells of acute leukemia patient (Group B) or normal subject (Group C) were taken as control.</p><p><b>RESULTS</b>The level of secreted SDF-1 protein (pg/10(5) cells/week) in the supernatants of Group A, B and C were 1920 +/- 205, 12,370 +/- 1355 and 6620 +/- 770, respectively. Of co-cultured Jurkat cells in Group A, B and C, the adhesion rates after 24 h co-culturing were (28.8 +/- 2.6)%, (57.4 +/- 3.8)% and (45.2 +/- 4.0)%, respectively, and the IC50 values of doxorubicin were 585, 6162 and 1758 nmol/L, respectively.</p><p><b>CONCLUSION</b>Down-regulating SDF-1 expression of bone marrow stromal cells by RNAi reduces adhesion rates and enhances drug sensitivity to doxorubicin of their co-cultured Jurkat cells.</p>

Effects of anti- CXCR4 monoclonal antibody on adhesiveness of human acute myelocytic leukemia cell line HL-60 and expression of Bcl-2, Fas proteins / 中国实验血液学杂志

To study the importance of chemokine SDF-1 in surviving of acute myelocytic leukemia cells HL-60, the adhesion ability of HL-60 and expression of Bcl-2, Fas protein when SDF-1 activity was blocked by anti-CXCR4 monoclonal antibody (12G5) were compared with those detected before MAb incubation, in this experiment, HL-60 cell were cultured and co-cultured with normal marrow stromal cell. The adhesion rate was detected while the expression of Bcl-2 and Fas proteins were assayed by immunohistochemical technique when SDF-1 activity was inhibited. The results showed that cell adhesion rate of HL-60 decreased while the expression Bcl-2 decreased and Fas increased. It is concluded that inhibition of SDF-1 activity increases cell apoptosis and thus reduces life-span of leukemia cell at certain level.

Inhibiting effects of stroma cell drived factor 1 (SDF-1) on proliferation of human acute myelocytic leukemia cell HL-60 / 中国实验血液学杂志

This study was aimed to investigate the importance of chemokine SDF-1 in maintaining proliferation ability of acute myelocytic leukemia cell line HL-60 when the effects of SDF-1 on HL-60 cell proliferation were inhibited. Marrow stromal cells were cultured and co-cultured with HL-60 cells, and SDF-1 activity was blocked with anti-CXCR4 McAb. HL-60 cell activity was detected by MTT while cell cycle and the expression of CXCR4 on HL-60 cell membrane were observed by flow cytometry meanwhile. The internal calcium ionic concentration in HL-60 cell was detected as well before and after treated with 12G5. The results showed that 12G5 down-regulated the expression of CXCR4 on HL-60 cell membrane; HL-60 cells at G(0)/G(1) phase increased, but decreased at S phase; survive rate of leukemia cells reduced; the intercellular calcium ionic concentration of HL-60 cell decreased after treated with 12G5. It was concluded that brockage of the SDF-1 activity may inhibit proliferation of leukemia cell at certain level.