Comparison of the Curative Efficacy of Elderly Patients with High-Risk MDS and MDS-Transformed AML between Decitabine Combined with Low-Dose CEG Regimen and Decitabine Combined with Low-Dose CAG Regimen / 中国实验血液学杂志

OBJECTIVE@#To evaluate the efficacy of decitabine combined with low-dose CEG regimen (DCEG) and decitabine combined with low-dose CAG regimen (DCAG) in the treatment of elderly patients with MDS and MDS-transformed acute myeloid leukemia (AML).@*METHODS@#A prospective study was conducted in 7 medical centers, 45 patients with MDS (≥ 60 years old) and MDS-transformed AML from October 2016 to January 2019 were enrolled, with the median age of 68.5 years old. The risk stratification of patients was poor or very poor, according to IPSS-R score. The treament results of decitabine combined with CEG and decitabine combined with CAG were compared.@*RESULTS@#The comparison of the two regiem showed that the DCEG regimen had advantages on total effective rate (ORR, 86.4% vs 47.8%, respectively), overall survival time (OS) (10.0 months vs 6.0 months, respectively) and progression-free survival time (PFS) (9.0 months vs 3.0 months, respectively). About 50% of MDS patients treated by DCEG regimen achieved PR or CR, with a median OS of 31 months. Multivariate analysis showed that patients with PR or CR after induction therapy and DCEG regimen had longer survival time (31months). The incidence of bone marrow suppression, infection and treatment-related mortality rate were similar between the two groups.@*CONCLUSION@#Decitabine combined with CEG regimen could improve the survival of patients with high-risk MDS and MDS-transformed AML. The conclusion of the reaserch needs to be validated by a larger prospective randomized clinical trial.

Application of miR-182 for Determination of Glucocorticoid-Resistant Patients with Lymphoid Malignancy / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To validate the clinical relationship between miR-182 and glucocorticoid-resistance in patients with lymphoid malignancy.</p><p><b>METHODS</b>Real-time quantitative PCR(qRT-PCR) was employed to detect the expression of miR-182 in lymphoma patients (68 cases, the specimens indluded bone marrow of 20 cases, and plasma of 48 cases), multiple myeloma patients (24 cases, the specimens included bone marrow of 14 cases and plasma of 10 cases), ALL patients (3 cases, specimen was plasma of 3 cases) and non-lymphotic system disorder patients (18 cases, specimens included bone marrow of 8 cases and plasma of 10 cases).</p><p><b>RESULTS</b>The expression of miR-182 in refractory lymphoblastic tumor patients was significantly higher than that in initial treatment group (P<0.05); the expression of miR-182 in initial treatment patients was not significantly different from the controls. The expression level of miR-182 in plasma of lymphoid malignancy patient significantly correlated with their used dosage of corticosteroids (P<0.05). When the expression level of miR-182 in bone marrow cells was 10.09, its sensitivity and specificity for diagnosis were 100% and 88.2% respectively; when the expression level of miR-182 in plasma was 1.393, its sensitivity and specificity for diagnosis of glucocorticoid resistance of lymphoblastic tumor cells were 90.9% and 51.3% respectively.</p><p><b>CONCLUSION</b>The expression of miR-182 in lymphoblastic tumor with glucocorticoid resistance is significantly up-regulated, suggesting that the glucocorticoid resistance of lymphoblastic tumor is related with the over-expression of miR-182. The miR-182 is expected as a new biomarker for the diagnosis of glucocorticoid resistant lymphoblastic tumor.</p>

Advances of studies on members of P53 family, interaction and relation with leukemia -review / 中国实验血液学杂志

The P53 gene has the important functions including induction of apoptosis, regulation of cell cycle, repair of DNA damage. The mutation of the P53 gene exists in more than 50% of human tumors and 13% of hematological malignancies. The P53 gene abnormality is closely related with the clinical course and prognosis of leukemia. The P73 or P63 gene, the member of the P53 family not only possesses similar to P53 activity of inducing apoptosis, activating transcription, but also plays different biological effects according to protein structural diversity, and even antagonises the function of the P53 gene. Researchers found that P73 or P63 gene also has the dual characteristics of the tumor suppressor and oncogene, and shows different expression and function in different types, different stages of leukemia. In this article, P53 family (P53, P73, P63) gene structure, biological function and the relationship of the three genes with the course, prognostic outcome, treatment and other clinical features of the leukemia are reviewed.

Amaxa Nucleofector(TM) nuclear transfection apparatus transfers L1210 cell line / 中国实验血液学杂志

Mouse L1210 leukemia cell line is widely used as a model in the study of tumorigenesis, as well as the efficacy of chemotherapeutic drugs; however, like other suspension cell lines, the mouse L1210 cell line has lowest transfection efficiency, that many barriers exist to study about the structure, function, as well as metabolism in leukemia cells. This study was aimed to obtain higher transfection efficiency of L1210 cell line to facilitate scientific research. The transfection efficiencies of nucleofector and liposome in L1210 leukemia cells were detected by converted fluorescence microscopy and flow cytometry using EGFP (enhance green fluorescent protein); cell viability was observed by trypan blue exclusion test. The results showed that the transfection efficiency of nucleofector primarily through reporter gene pEGFP by Amaxa Nucleofector(TM) nuclear transfer apparatus was significantly higher than lipofectamine 2000 transfection, furthermore, in the same cell density (2 × 10(6)/ml) and plasmid content (10 µg), the transfection efficiency of nuclear transfer apparatus default mode A-20 was higher than that of other modes (S-18, T-20). Its survival rate was up to 50.5% after 24 hours. Cell viability of liposome transfection reached to 88% after 24 hours, but the transfection efficiency was lower (< 1%). It is concluded that the nuclear transfer apparatus A-20 transfected L1210 can reach higher transfection efficiency up to 61.6%, which is significantly higher than that of lipofectamine transfection. The survival rate is up to 50.5% well meeting the needs of scientific research. Higher transfection efficiency is helpful for in-depth research about the morphology, functions and pathogenesis in leukemia model L1210, and provides more searching space for the treatment of leukemia diseases.

Combination of dexamethasone with IL-2 selectively induces the expansion of CD4(+)CD25(+)FOXP3(+) regulatory T cells in vivo and suppresses graft versus host disease / 中华血液学杂志

<p><b>OBJECTIVE</b>To establish a method for increasing T regulatory cells (Treg) in the graft by using in vivo treatment with dexamethasone (Dex) plus IL-2 and observe its suppressing effect on graft-versus-host-disease (GVHD) in mice.</p><p><b>METHODS</b>After treatment of male C57BL/6N mice (donor) with Dex (5 mgxkg(-1)xd(-1)) combined with IL-2 (300 000 IUxmouse(-1)xday(-1)) for three days, spleen mononuclear cells were isolated for flow cytometry analysis of CD4(+)CD25(+) POXP3(+) Treg cells. The allogeneic lymphocytes were transplanted from male C57BL/6N mice to female BALB/c mice aged 8-12 weeks. GVHD and survival time were investigated after transplantation. Donor-derived hematopoiesis reconstituted in recipient mice was detected by Y-chromosome-specific PCR and H-2K(b) by flow cytometry.</p><p><b>RESULTS</b>Administration of Dex and IL-2 markedly expanded functional CD4(+)CD25(+)FOXP3(+) Treg cells in murine spleen, the number of which in treated group was (24.2 +/- 7.6)% while in control group was (4.0 +/- 0.8)% (P = 0.01). The ratio of Treg to effector T cells (Teff) increased obviously in the treated group (0.43 +/- 0.15 vs 0.14 +/- 0.01, P = 0.01). In a murine allogeneic lymphocyte transplantation model, the grafts from donor with combined treatment of Dex and IL-2 led to a longer survival time than that from the control group (median survival time > 60 d vs 12 d, P = 0.0045), while the mortality rate was decreased (29.4% vs 71.4%, P < 0.05).</p><p><b>CONCLUSION</b>Costimulation with Dex and IL-2 can selectively expand the functional CD4(+)CD25(+)FOXP3(+) Treg in vivo, which can suppress acute GVHD.</p>

Mechanism underlying IL-2 inhibition on T cell specific immune response / 中国实验血液学杂志

The study was aimed to investigate the effect of IL-2 on the acquired immune response of naive T cells, and to explore the influence of IL-2 on suppressor of cytokine signaling 3 (SOCS-3) expression of naive T cells, as well as to elucidate the role of SOCS-3 on antigen specific immune response in vitro. Naive DO11.10 T cells were pre-stimulated with IL-2 (50 U/ml), and stimulated with OVA(323 - 329) antigen after removing IL-2, then the proliferation of naive DO11.10 T cells was detected. Naive DO11.10 T cells were stimulated with IL-2 (50 U/ml), and SOCS-3 expression was detected by real-time PCR. Naive DO11.10 T cells were stimulated with OVA(323 - 329) antigen, and SOCS-3 expression was detected by means of (3)H-TdR. The results showed that after IL-2 pre-stimulation, the proliferation of naive DO11.10 T cells decreased significantly when stimulated with OVA(323 - 329) antigen; SOCS-3 expression of naive DO11.10 T cells was up-regulated significantly after IL-2 stimulation, the up-regulation began obviously at 4 hours and reached peak at 6 hours. SOCS-3 expression on naive DO11.10 T cells was down-regulated markedly after OVA(323 - 329) antigen stimulation, the expression level of SOCS-3 was lowest on day 2 and returned to normal on day 4 after stimulation. It is concluded that the antigen specific immune response of naive DO11.10 T cells is inhibited after pre-stimulation with IL-2, which may be mediated by SOCS-3.

Effect of HMBA on differentiation and apoptosis of HL-60 and U937 cells and its mechanism / 中华血液学杂志

<p><b>OBJECTIVES</b>To explore the effect of hexamethylene bisacetamide (HMBA) on the differentiation and apoptosis of HL-60 and U937 cells, and its mechanism.</p><p><b>METHODS</b>Flow cytometry was used to evaluate the expressions of cellular surface antigen CD(11b), CD(14), apoptotic marker Annexin V, cell cycle distribution and endocytic antigen cyclin D, cyclin E and p27. Changes of c-myc, Rb, Bcl-2 gene mRNA levels were detected by RT-PCR.</p><p><b>RESULTS</b>After 72 hours of HMBA treatment, CD(11b) expressions increased significantly, apoptosis increased under high-dose HMBA, cells were arrested in G(0)/G(1) phase and reduced cyclin E, increased cyclin D and p27 were significant in a dose-dependent manner in HL-60 and U937 cells. RT-PCR showed that c-myc and bcl-2 mRNA was significantly down-regulated and Rb mRNA up-regulated in HL-60 and U937 cells.</p><p><b>CONCLUSION</b>HMBA can induce the differentiation of HL-60 and U937 cells, while apoptosis of these cell is induced only by high dose of HMBA. The possible mechanism of HMBA inducing differentiation might be related to the changes of cell cycle regulators and certain proliferation and differentiation related genes.</p>

Clinical and laboratory studies of expression of cyclin A in leukemia cells / 中国实验血液学杂志

Uncontrolled cell proliferation is the basic feature of cancer. Some of the prime cell cycle regulators are involved directly in tumorigenesis. Cyclin A, one of the G(1)/S cyclin, can cause transformation. The purpose of this research was to investigate whether cyclin A overexpression was involved in leukemogenesis and proliferation of leukemia cells. The expression of cyclin A at S-phase in leukemia cell line HL-60, blast cells of acute leukemia patients, bone marrow cells of outpatients without malignant hematological disease and peripheral blood cells of healthy donors was investigated by simultaneous indirect immunofluorescence staining of intracellular antigen and DNA. To further investigate whether cyclin A played as a key molecular in cell proliferation, HL-60 cells were exposed to different concentrations of hexamethylene bisacetamide (HMBA). MTT dye absorbance of living cells and cell cycle analysis were adopted to evaluate growth arrest. Differentiation was evaluated by detection of the change of expression of CD11b and CD33 on cell surface. The results showed that overexpression of cyclin A was only found among specimens from acute leukemia and leukemia cell line. There was no elevated cyclin A detection for cyclin A among specimens from outpatients and healthy donors. In HMBA interference experiment, HMBA was able to induce growth arrest and monocytic macrophage differentiation of HL-60 cells in a dose-dependent manner, and all these changes were associated with a marked down-regulation of cyclin A expression. In conclusion, aberrant overexpression of cyclin A at S-phase was only found in leukemia cell lines and blast cells from acute leukemia. The dose-dependent effect of HMBA on cell growth and differentiation of HL-60 cell line which was consistent with the decrease of cyclin A expression in these cells suggested that the molecular mechanisms of HMBA inducement involved downregulation of cyclin A expression.

Proliferation and IFN-gamma secretion of autologous T lymphocytes stimulated by myeloid leukemia cells induced with rhGM-CSF and rhIL-4 / 中国实验血液学杂志

To observe the proliferation of T lymphocytes stimulated by CML and AML cells which were induced by rhGM-CSF and rhIL-4, and the secretion of IFN-gamma from proliferated T lymphocytes, the expression of CD80, CD86 and HLA-DR on CML and AML cells induced by GM-CSF and IL-4 was assayed by flow cytometry in vitro. Then one-way mixed lymphocyte reaction was carried out, with induced leukemia cells as stimulating cells and auto-T lymphocytes as reactive cells. The secretion of IFN-gamma from T lymphocytes was determined by double antibody sandwich ELISA. The results showed that GM-CSF and IL-4 significantly upregulated the expression of CD80, CD86 and HLA-DR on CML cells and CD80 and CD86 on AML cells, which could stimulate the T lymphocyte proliferation and high secretion of IFN-gamma (in CML group) of autologous T lymphocytes. It is concluded that the CML and AML cells induced by GM-CSF and IL-4 have the ability to present tumor specific antigen to auto-T lymphocyte.