The aim of this study was to investigate the potential benefit of combination therapy with 5-bromotetrandrine (5-BrTet) and daunorubicin (DNR) on chronic leukemia. The apoptosis of K562/A02 cells treated by DNA, BrTet and BrTet combined with DNR for 48 hours was detected by flow cytometry; the expressions levels of survivin mRNA and protein K562/A02 cells treated by DNR, BrTet and BrTet combined with DNR and in untreated K562 cells for 48 hours were measured by RT-PCR and Western blot respectively. The results showed that the combination of BrTet with DNR increased apoptotic rate of K562/A02, down-regulated the expression levels of survivin mRNA and protein in K562/A02 cells as compared with blank control and cells treated by BrTet or DNR alone, the survivin expression in K562/A02 cells was higher than that in K562 cells. It is concluded that the combination of BrTet with DNR can effectively reverse the multidrug resistance of K562/A02 cells, promote the apoptosis of K562/A02 cells, the mechanism of which may be related with down-regulation of survivin expression. Survivin may be a target for the treatment of MDR in hematopoietic malignancies.
Apoptosis ; drug effects ; genetics ; Benzylisoquinolines ; pharmacology ; Daunorubicin ; pharmacology ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Gene Expression Regulation, Leukemic ; Humans ; Inhibitor of Apoptosis Proteins ; genetics ; K562 Cells

The aim of study was to investigate the effect of acute lymphoblastic leukemia (ALL) children bone marrow mesenchymal stem cells (MSC) on resistance of K562/A02 cells and its mechanism. MSC obtained from bone marrow of AL children were cultured and identified. The co-culture of MSC and K562/A02 and the culture of K562/A02 cell suspension alone was performed, of which 2 kinds of cells were treated with same concentration of adriamycin (ADM), and the rate of apoptosis was detected by flow cytometry, bcl-2 and bax of K562/A02 were detected by RT-PCR, while mdr1 gene level was detected by FQ-PCR. The results indicated that the MSC separation and proliferation were viable and steady. The apoptosis rate of the K562/A02 cells co-cultured with MSC was 1.97 ± 0.11%, while apoptosis rate of the K562/A02 cells cultured alone was 8.38 ± 0.29%, there was significant difference (p < 0.05). As compared with the K562/A02 cells cultured alone, the bcl-2 gene expression in K562/A02 cells co-cultured with MSC obviously increased; ratio of bcl-2/bax was obviously enhanced. The mdr1 gene level in K562/A02 co-cultured with MSC was no statistical different from K562/A02 cultured alone (p > 0.05), which suggested that adhesion co-cultured with MSC did not induce mdr1 expression higher than the culture of suspension. It is concluded that the MSC of ALL children can escape the leukemia cells from proapoptotic effect of drugs, the resistance of K562/A02 to ADM may be involved in enhancement of bcl-2 gene expression of K562/A02 cells co-cultured with MSC, but not in relation to mdr1 gene in K562/A02 cells themselves.
ATP Binding Cassette Transporter, Sub-Family B ; ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; Bone Marrow Cells ; drug effects ; Child ; Child, Preschool ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Female ; Gene Expression Regulation, Leukemic ; Humans ; K562 Cells ; Male ; Mesenchymal Stromal Cells ; drug effects ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; genetics ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; bcl-2-Associated X Protein ; genetics

This study was aimed to explore the expression of erythropoietin receptor (EPOR) on acute leukemia cells and its clinical significance. Bone marrow of 40 patients with acute leukemia (AL) and 24 patients with normal bone marrow as control group were collected. Samples came from outpatients and inpatients in our hospital. EPOR mRNA was detected by reverse transcription-PCR. The results showed that there was EPOR expression on AL cells, the expression rate was 57.5%, and the average expression level (Gray value) was 0.3549 ± 0.2800, but both were lower than that in control group (p < 0.05). There was no significant statistic difference of expression rate between acute myelogenous leukemia (AML) and acute lymphoblastic leukemia (ALL) (p > 0.05), and expression level of AML EPOR was higher than that of ALL (p < 0.05). It is concluded that there is EPOR expression on AL cells, while the expression rate and expression level are lower than those in control group (p < 0.05). There is no significant statistic difference of the expression rate between AML and ALL (p > 0.05), and the expression level of AML EPOR is higher than that of ALL (p < 0.05).
Case-Control Studies ; Humans ; Leukemia, Myeloid, Acute ; genetics ; metabolism ; RNA, Messenger ; genetics ; Receptors, Erythropoietin ; genetics ; metabolism

This study was purposed to detect single nucleotide polymorphisms (SNP) of 2 pharmacokinetics-related genes in K562 and K562/A02 cell lines. Leukemia cell line K562 and its resistant line K562/A02 were cultured, the genomic DNA was isolated by QIAamp DNA Blood Mini kit, primers were designed, the related DNA fragments were amplified by PCR. The SNP genotyping of mthfr gene rs1801131, rs1801133 and rs2274976 and dpyd gene rs1801159, rs1801160 and rs17376848 was performed by means of matrix assisted laser desorption ionization-time of flight mass spectrometry method (MALDI-TOFMS). The results showed that the genotype of mthfr gene locus 1801131 was AC, rs1801133 was CC, rs2274976 was GG, genotype of dpyd gene locus 1801159 was GG, rs1801160 was GG, rs17376848 was AA in both K562 and K562/A02 cell lines. It is concluded that the above-mentioned loci of mthfr and dpyd genes in K562 and K562/A02 cell lines are not expressed differently.
DNA Mutational Analysis ; DNA Primers ; Dihydrouracil Dehydrogenase (NADP) ; genetics ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; Genotype ; Humans ; K562 Cells ; Methylenetetrahydrofolate Reductase (NADPH2) ; genetics ; Polymorphism, Single Nucleotide

The purpose of study was to investigate the cytogenetic abnormality of acute leukemias (AL), to analyze the relationship in the chromosomal abnormality and the AL FAB types, and to explore the impact of the chromosomal abnormalities on the prognostic factors of AL. The chromosome karyotypes of 397 patients with AL were analyzed by means of bone marrow short-term culture and G banding technique. The results showed that in 319 out of 397 patients, the chromosome karyotypes could be analyzed, and the chromosomal abnormality occurred in 175 patients (54.9%). In the patients with acute lymphocytic leukemia (ALL), acute myelogenous leukemia (AML) and acute mixed-lineage leukemia (AMLL), the chromosomal abnormality occurred respectively in 33 of 120 patients (27.5%), 129 of 252 patients (51.2%) and 13 of 25 patients (52.0%). Hyper-diploids, hypo-diploids and diploids occurred in 41 of 175 patients (23.4%), 22 of 175 patients (12.5%), and 112 of 175 patients (64.0%) respectively. In patients with AML the FAB type-associated chromosomal abnormality occurred in 69 of 129 patients (53.5%). It is concluded that chromosomal abnormalities exist in about 55% AL patients. Some special chromosomal abnormalities are cytogenetic characteristics of AL, and obviously correlated with AL FAB types, the combination of chromosomal detection with cytogenetics is useful for the diagnosis of AL, and the evaluation of therapeutic effects and prognosis.
Acute Disease ; Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child ; Chromosome Aberrations ; Female ; Humans ; Karyotype ; Karyotyping ; Leukemia ; genetics ; Male ; Middle Aged ; Young Adult

This study was aimed to establish a stable subline of K562 cells (K562-HMGB1) overexpressing HMGB1 protein and K562-HMGB1 sublines served as control, so as to provide a basis for exploring the role of hmgb1 gene in occurrence and development of leukemia and their mechanism. Protein-coding gene of hmgb1 was amplified by PCR with cDNA as template, which was synthesized by reverse transcription from total RNA extracted from U937 cells. The PCR-amplified hmgb1 gene was ligated into PMD18-T vector (PMD18-T-HMGB1 vector), and then transformed into E. coli strain DH5α. DH5α containing PMD18-T-HMGB1 vector were grown on LB agar plate supplemented with 100 µg/ml ampicillin overnight. The single ampicillin-selected DH5α clone was picked for culturing overnight and then harvested for plasmid extraction. The extracted plasmid was characterized to contain hmgb1 gene digested with the desired restriction enzymes of KpnI/XhoI. The correctness of hmgb1 sequence was confirmed with DNA sequencing. The insert of hmgb1 gene contained in PMD18-T-HMGB1 vector was cut out with restriction enzymes of KpnI/XhoI and then ligated into eukaryotic expression vector pcDNA3.1 to form pcDNA3.1-HMGB1 vector. 10µg of pcDNA3.1-HMGB1 or pcDNA3.1 plasmid was separately electroporated into K562 cells. At 48 hours after electroporation the cells were cultured with G418 at a final concentration of 800 µg/ml for over 2 weeks. Finally stably transfected sublines of K562 cells containing hmgb1 gene (K562-HMGB1), and of K562 containing pcDNA3.1 vector (K562-pcDNA3.1) served as a control, were obtained. The transcriptional or translational expression of hmgb1 gene was detected with RT-PCR or Western blot, respectively, to testify transfected efficiency and validity of stable subline of K562-HMGB1. The results indicated that the eukaryotic expression vector pcDNA3.1-HMGB1 plasmid was successfully constructed and was electroporated into K562 cells. The transcriptional or translational expression of hmgb1 gene in the stable subline of K562 cells containing hmgb1 gene was overexpressed. It indicated that stable subline of K562-HMGB1 cells was successfully established. It is concluded that the stable sublines of K562-HMGB1 cells or K562-pcDNA3.1 cells are successfully established, which provides a basis for exploring the roles and mechanisms of hmgb1 gene in leukemogenesis and development of leukemia.
Gene Expression ; Genes, Regulator ; Genetic Vectors ; HMGB1 Protein ; genetics ; Humans ; K562 Cells ; metabolism ; Plasmids ; Transformation, Genetic

Human leukemias are considered as clonal malignancies initiated at stage as early as hematopoietic stem/progenitor cells. The drug resistance and relapse are two major causes for treatment failure of leukemia. Recently, the discovery of leukemia stem/progenitor cells (LSPC) and subsequent research have provided a cue to elucidate the pathogenesis of leukemia and to explore the strategies of targeted therapy against LSPC. This review summarizes the molecular characteristics of LSPC and some research advances of therapy targeting LSPC including therapy targeting to surface molecules of LSPC, interference of interaction between LSPC and bone marrow microenvironment, regulation mechanisms of some specific molecular and so on.
Humans ; Leukemia ; therapy ; Neoplastic Stem Cells ; Stem Cell Transplantation

Leukemic dendritic cells, compared to normal dendritic cells, have the similar surface molecules, but have poor antigen-presenting function. When inoculated in the human body, the dendritic cells can not produce enough immune response. Therefore, how to make dendritic cells mature and have its own function to play a better anti-leukemia immune response is the key problem in clinical treatment. And it is one of the hottest studies in the immunotherapy field. This review focuses the recent progress of research on the culture of dendritic cells in vitro, its influence factors and so on.
Cell Culture Techniques ; Cell Differentiation ; Dendritic Cells ; cytology ; Humans ; Immunotherapy ; Leukemia ; immunology ; therapy

Tumor Stem/Progenitor Cells are characterized by undifferentiation/hypodifferention, self-renewal, differentiation potential, which is responsible for tumor occurrence, growth and metastasis. The theory about tumor stem/progenitor cells provides a new insight to recognize the origin and essence of tumors. Gaining a better insight into the origin of tumor stem/progenitor cells and the mechanisms of tumor stem/progenitor cells resistance to chemotherapy may lead to new therapeutic targets and better anticancer strategies. This review summarizes recent progress of research on the origin of tumor stem/progenitor cells and multidrug resistance.
Cell Line, Tumor ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; Humans ; Neoplasms ; drug therapy ; Neoplastic Stem Cells ; drug effects ; Stem Cells ; drug effects

Chemotherapy remains at the first line for the treatment of leukemia. However, the multi-drug resistance of the tumor cells caused by chemotherapeutic drugs has seriously affected the effect of chemotherapy. And this is the main reason for the failure of the leukemia treatment. Therefore, to explore an effective way of reversing drug resistance has become the key of leukemia treatment. RNA interference, a system within living cells, helps to determine which genes are active and how active they are. It is a process in which translation of some cell messenger RNA (mRNA) sequences is prevented, because of the presence of (and consequent destruction of) matching double-stranded RNA sequences. RNA interference is also called post-transcriptional gene silencing (PTGS), since its effect on gene expression occurs after the production of mRNA during transcription. It is believed that RNA interference can protect the cell against viruses and other threats. The greatest advantage of RNAi is the specificity and high efficiency which can induce suppression of specific genes of interest but the unrelated genes are not affected. The selective and robust effect of RNAi on gene expression makes it a valuable research tool both in cell culture and living organisms because synthetic dsRNA introduced into cells can induce the suppression of specific genes of interest. Nowadays, the technology has been widely used in biomedical fields, especially in the diagnosis and treatment of blood system disease. However, besides the stability, targeting and biological safety in genetics, the immune response induced by exogenous RNA is also one of the key factors to limit the clinical practice of this emerging technology. In this review, the breakthrough of the technology in multi-drug resistance reversal in leukemia is summarized with the RNA interference technology as a starting point.
Cell Line, Tumor ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Gene Silencing ; Humans ; Leukemia ; genetics ; therapy ; RNA Interference