Expansion and activation of granulocytic, myeloid-derived suppressor cells in childhood precursor B cell acute lymphoblastic leukemia.

Precursor B cell acute lymphoblastic leukemia (B-ALL) is a B cell-derived, malignant disorder with the highest incidence among children. In addition to the genetic abnormality, a dysregulated immune system also has an important role in the pathogenesis of B-ALL. Myeloid-derived suppressor cells (MDSCs) represent one of the key drivers in immune tolerance against tumor cells, including various solid tumors and hematologic malignancies.

[Effects of hermap gene on p-STAT5 kinases in signal transduction pathway during erythroid differentiation].

Objective: To study the effects of hermap gene on kinases in erythroid signal transduction pathway and investigate the mechanism of hermap on erythroid differentiation.

Methods: The K562 cells expressing hermap and hermap-siRNA respectively were established for up- and down-regulating the expression of hermap gene. These K562 cells were then induced by Ara-C to erythroid differentiation and analyzed at 0, 24, 48, 72 and 96 h, respectively, for cell morphology and biphenylamine staining positive cells, determination of CD235a, CD36, kinases p-STAT5, p-Akt, p-MAPK and p-c-JUN by FCM; and quantification of hermap gene and γ (Aγ,Gγ) globin gene by FQ-PCR.

[Establishment of the cell line K562 with stable expression of hermap and hermap-siRNA].

In order to establish K562 line with stable expressions of hermap and hermap-siRNA, amplified hermap and hermap-siRNA were cloned into pEGFP-c1 and pRNAT to acquire hermap-pEGFP-c1 and hermap-siRNA-pRNAT, respectively. These two plasmids were electrotransferred into K562 cells, then were followed by culturing with G418. The result showed that the transfer rate of hermap-pEGFP-c1-K562 and hermap-siRNA-pRNAT-K562 plasmids were 10.

[Relationship between the antileukemic activity of L-asparaginase and Asn level around leukemic cells].

Objective: To study the antileukemic activity of L-asparaginase through determining the changes of 4 kinds of amino acids (Asn, Aspa, Glu and Gln) in cell culture medium.

Methods: Following L-Asp treatment with designed concentrations and duration, the IC50 (inhibitory concentration 50%) of 8 kinds of common leukemia cell lines (U937, HL-60, Jurkat, NB4, THP-1, Namalwa, Karpass299, K562) were determined by CCK-8 assay. The changes of the 4 kinds of amino acids mentioned above were detected by high performance liquid chromatography (HPLC).

[Relationship between asparagine synthetase expression level and cell sensitivity to L-asparaginase in human leukemic cell lines].

This study was purposed to explore the relationship between asparagine synthetase (AsnS) mRNA expression level and the sensitivity of leukemic cell lines to L-asparaginase. The AsnS mRNA expression level in 8 cell lines (Jurkat, HL-60, U937, NB4, THP-1, Namalwa, Karpas299 and K562) was determined by real-time quantitative PCR (RQ-PCR) based on fluorescence dye Eva Green before and after treatment with L-Asp, and the cell proliferation rates were analyzed by CCK-8 assay. The results showed that there was a significant disparity of AsnS expression level in 8 cell lines, and there were significant increases of AsnS expression level in cells co-cultured with L-Asp (p < 0.

[Interaction between mixed-lineage leukemia and Menin proteins].

The study was aimed to investigate the subcellular localization of mixed-lineage leukemia (MLL) and Menin proteins, and to explore the interaction between these two proteins. The recombinant eukaryotic cell expression vectors of pcDNA3.1-myc-MLL and pCMV-flag-Menin were constructed respectively, and transfected into the HEK293T cells.

[Effects of human ERMAP-siRNA on erythroid differentiation of K562 cells induced by Ara-C].

In order to investigate the potential role of human ERMAP gene in erythropoiesis, the ERMAP-dsDNA was designed, ERMAP-shRNA expressing plasmids was constructed, and ERMAP-shRNA/K562 cell was established. Cell morphology, biphenylamine staining, expression of cell surface antigens as well as quantitative level of human ERMAP gene were observed during K562 cells differentiating toward erythroid lineage induced by Ara-C. The results showed that at 72 hours after Ara-C treatment, ERMAP-shRNA/K562 cell size became large with increasing cytoplasm content.

[Expression of human ermap gene in umbilical cord blood mononuclear cells during differentiation and development towards erythroid lineage].

The aim of study was to explore the potential of human erythroid membrane associated protein (ERMAP) gene in erythroid cell differentiation and development, mononuclear cells (MNCs) were isolated from umbilical cord blood and induced to erythroid cell differentiation by SCF, IL-3 and EPO. The cell morphology was observed by using optical microscopy, the positive rate of cells was counted by biphenylamine staining and the ratios of CD36+/CD235a-, CD36+/CD235a+, CD36-/CD235a+ cells were detected by flow cytometry, the change of human ermap gene expression level was analyzed by using fluorescent quantitative PCR (FQ-PCR). The results showed that the ermap gene expression level increased while MNCs were induced to erythroid lineage after treatment with SCF, IL-3 and EPO.

[Detection of minimal residual disease in children leukemia patients by using PCR - review].

MRD detection in children leukemia has a potential importance to predict clinical outcome and to modify treatment protocols of the diseases. Although some patients with leukemia have achieved complete remission according to the clinical and morphological criteria, there are still very low numbers of malignant cells that can not be discriminated by morphology and remained in bone marrow, which is called minimal residual disease (MRD) and is the main reason leading to relapse. MRD detection has an important significance for designing treatment protocols.

[Expression of human ERMAP gene in fetal tissues].

Human ERMAP (hERMAP) is a novel gene coding for erythroid membrane-associated protein, which may play a role in erythropoiesis. To explore the role of hERMAP in fetal hematopoiesis, 29 fetuses of inevitable abortion with the fetal ages of 9 - 36 weeks were collected, and the total RNAs were isolated from the liver, spleen, kidney, heart, lung, thymus, brain, bone marrow and skeletal muscle, the RNAs from which at 25(+5) week fetal age were used to detect hERMAP expressions in different fetal tissues with Northern blot, and the hERMAP in various fetal tissues were quantified by FQ-PCR. As a result, Northern blot showed that hERMAP was expressed in liver and bone marrow at 25(+5) fetal age, but not in the other organ tissues.

[RNA isolation from human embryonic tissues].

To investigate the method of RNA isolation from human embryonic tissues and the factors influencing the quality of RNA, the RNA from human embryonic tissues obtained with drug-induced labor or non-drug induced labor were isolated by using grind with liquid nitrogen or homogenizer without liquid nitrogen. The results showed that the positive rates of RNA integrity in grind with liquid nitrogen group and in homogenizer without liquid nitrogen group were 68.42% and 29.

[Expression of human ERMAP gene in different cell lines].

To investigate the pattern of human ERMAP gene expression in different cell lines, 15 cell lines derived from hematopoietic tumor, somatic tumor and normal tissue were chosen and were cultured; cells were harvested after culture for 12, 24, 36, 48, 60 and 72 hours; the expression of the human ERMAP was detected by using fluorescent quantitative PCR. The results showed that human ERMAP gene expression was positive in K562 cell line at interval of 12 and 24 hours, and the expression levels were (5.092 +/- 2.

[Study on the expression of human ERMAP gene in erythropoietic and macrophage differentiation of K562 cells].

In order to investigate the potential of human ERMAP gene in erythroid cell differentiation, K562 cells were induced to erythroid lineage by Ara-C and to macrophage lineage by TPA, human ERMAP mRNA was detected by fluorescent quantitative PCR. The results showed that human ERMAP mRNA increased while K562 cells were induced to erythroid lineage after treatment with Ara-C at 2.5 x 10(-6) mmol/L/L and 1.