OBJECTIVETo study the synergistic mechanism between PH II -7 and doxorubicin against multi-drug resistant HL-60/ADR cells and its parent HL-60 cells.

METHODSThe anti-tumor activity of doxorubicin alone and combined with PH II -7 were measured by MTT assay. RNA was extracted from the cells treated with PH II -7 for different times or doses then the expression of MRP gene was measured by RT-PCR. Confocal laser scanning microscopy and FACS were used to detect the intracellular cumulation of doxorubicin in PH II -7 treated HL-60 and HL-60/ADR cells.

RESULTSPH II -7 has anti-tumor effect with IC50 of (0.83 +/- 0.08) micromol/L and (1.74 +/- 0.56) micromol/L for HL-60 and HL-60/ADR, respectively. It could potentiate the anti-tumor effect of doxorubicin with CDI of 0.7 and 0.43 for HL-60 and HL-60/ADR, respectively. PH II -7 and doxorubicin act synergistically in inhibiting the proliferation of HL-60 and HL-60/ADR cells and down-regulating the expression of MRP gene in a dose and time dependent manner. PH II -7 restored the intracellular cumulation of doxorubicin in HL-60/ADR cells to 55% of that in HL-60 cells.

CONCLUSIONPH II -7 can significantly hasten the cytotoxicity of doxorubicin to HL-60 and HL-60/ADR cells through down-regulating the expression of MRP. The synergistic effect was more obvious in HL-60/ADR cells.

ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Drug Synergism ; HL-60 Cells ; Humans

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

This study was aimed to explore the changes of soluble resistance-related calcium binding protein (sorcin) expression in reversion of multidrug resistance of K562/A02 leukemic cell line with different concentrations of tetrandrine (Tet), so as to provide a new theoretic evidence for clinical application of Tet. The inhibition of K562/A02 cell line by daunorubicin (DNR) was assayed by MTT method. The changes of SORCIN gene expression were assayed by RT-PCR. The changes of SORCIN protein expressed were assayed by Western blot. The results showed that Tet could enhance the cytotoxicity of DNR to K562/A02 cells (the IC(50) of DNR + Tet was 11.3+/-0.17 mg/L, 5.15+/-0.10 mg/L, 3.91+/-0.06 mg/L, and 2.52+/-0.04 mg/L, when concentrations of Tet were 0 mg/L, 0.5 mg/L, 1.0 mg/L, and 2.0 mg/L respectively). The gene encoding sorcin was highly expressed in K562/A02 cells, the expression of which was firstly enhanced in Tet concentration 0.5 mg/L, then attenuated in Tet concentration of 1.0, 2.0 mg/L (p<0.05). Sorcin protein expressed lowly in K562 cells and highly in K562/A02 cells, but the expression of SORCIN protein in K562/A02 cells was enhanced in Tet concentration of 0.5 mg/L, then was attenuated in Tet concentration of 1.0, 2.0 mg/L (p<0.05). It is concluded that the effect of Tet on reversal of K562/A02 drug-resistance shows concentration dependence by MTT assay. Tet reverses multidrug-resistance of K562/A02 cells through regulation of expression of SORCIN gene and protein, but not fully correlates to the reversing effect.
Antineoplastic Agents, Phytogenic ; pharmacology ; Benzylisoquinolines ; pharmacology ; Calcium-Binding Proteins ; genetics ; metabolism ; Doxorubicin ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Humans ; K562 Cells

OBJECTIVETo determine the reversal effects of ramification of curcumin hydrolyzed on multidrug-resistant cell line K562/A02, and explore its reversal mechanism.

METHODAfter treatment with ramification of curcumin hydrolyzed, the sensitivity of K562/A02 cells to usuall chemotherapeutic drugs were determined by MTT. The expression of P-gp in K562/A02 and K562 cells was detected by immunohistochemical method. Intracellular mean fluorescence intensity (MFI) of DNR in K562 and K562/A02 cells was detected by Flow Cytometry.

RESULTAfter treatment with the ramification of curcumin, hydrolyzed (2.5 mg x L(-1)) IC50, of usuall chemotherapeutic drugs to K562/A02 decreased. The sensitivity of K562/A02 cells increased. The expression of the P-gp in K562/A02 cells decreased (P < 0.05); MFI of the DNR in K562/A02 cells more significantly increased (P < 0.05). The expression of mdrl-mRNA decreased.

CONCLUSIONThe ramification of curcumin hydrolyzed have effects on the reversal multidrug-resistant of K562/A02 cells in vitro. It could enhance the sensitivity of K562/A02 cells to chemotherapeutic drugs and the mechanism might be associated with inhibiting P-gp-mediated drug efflux and increasing of intracellular concentration of chemotherapeutic drugs.

ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Curcumin ; chemistry ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Gene Expression ; drug effects ; Humans ; K562 Cells ; Neoplasms ; drug therapy ; genetics ; metabolism

OBJECTIVETo study the reverse effect and its mechanism of tetrandrine (Tet) on human breast cancer cells resistant to tamoxifen (MCF-7/TAM).

METHODSThe drug toxicity and the reverse effect of Tet on MCF-7/TAM cells were detected by MTT assay. The effects of multidrug resistance protein 1 (MRP1) gene of Tet on MCF-7iTAM cells were detected by real-time fluorescence quantitative PCR. The changes of MRP1 protein on MCF-7JTAM cells were detected by Western blot.

RESULTSTet had a significant reversal of drug resistance on MCF-7/TAM cells. The non-cytotoxic dose (0. 625 microg/mL) reversed the resistance by 2.0 folds. MRP1 was reduced at gene (P <0.05) and protein levels when Tet effected on MCF-7ITAM cells.

CONCLUSIONTet could reverse the drug resistance of MCF-7/TAM cells, and the reverse mechanism may be related to down-regulating MRP1 expression.

Benzylisoquinolines ; pharmacology ; Drug Resistance, Neoplasm ; drug effects ; genetics ; Female ; Humans ; MCF-7 Cells ; drug effects ; Multidrug Resistance-Associated Proteins ; metabolism ; Tamoxifen ; pharmacology

OBJECTIVETo investigate the apoptosis-induction, P-glycoprotein (P-gp) and mdr1 mRNA inhibition effects of arsenic trioxide (As2O3) and buthionine sulfoximine (BSO) on multidrug-resistant cell line K562/ADM cells, and to determine the relationship between intracellular GSH content and arsenic effect.

METHODSK562/ADM cells were treated with arsenic (0.5, 2.0, 5.0 micromol/L) alone or combined with BSO (100 micromol/L). The cell proliferating capacity was assessed with MTT assay, and cell apoptosis by Annexin V and propidium iodide (PI) staining. Intracellular GSH contents were measured using a glutathione assay kit by spectrophotometry. P-gp expression was determined by flow cytometry, and mdr1 mRNA expression by semi-quantitative RT-PCR.

RESULTSThe GSH contents in K562/ADM cell was (81.13 +/- 3.91) mg/g protein. After the GSH contents were degraded by BSO, the K562/ADM cell proliferating capacity was obviously inhibited and the cells were induced apoptosis in 24 hours by the combination of clinic dose arsenic group (0.5, 2.0 micromol/L) and BSO (100 micromol/L). The cell apoptosis rates at 48 hours in arsenic alone group and combination group were (59.29 +/- 6.01)% and (65.06 +/- 8.29)%, and at 72 hours were (82.15 +/- 9.28)% and (92.72 +/- 9.41)% retrospectively. At 48 hours, the mdr1 mRNA inhibition effect of the combination group was obviously stronger than that of high dose arsenic alone group. At 72 hours, the P-gp inhibition effect of the combination group (clinic dose arsenic group, 0.5, 2.0 micromol/L) was obviously stronger than that of high dose arsenic alone group (5.0 micromol/L).

CONCLUSIONThe intracellular GSH contents are closely correlated with the arsenic effect. The combination of conventional dose arsenic and BSO significantly induces K562/ADM cell apoptosis and inhibits P-gp and mdr1 mRNA expression in the cells.

ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Apoptosis ; drug effects ; Arsenicals ; pharmacology ; Buthionine Sulfoximine ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Genes, MDR ; drug effects ; Glutathione ; metabolism ; Humans ; K562 Cells ; Oxides ; pharmacology

OBJECTIVETo study the resistant related molecules of human leukemia drug resistant K562 cells (K562/HHT) induced by homoharringtonine (HHT).

METHODSGene expression profiles on K562/HHT, K562 and K562/HHT/RU486 (K562/HHT reversed by RU486) cells were detected by DNA microarray. The bone marrow tyrosine kinase gene in chromosome X (BMX) which changed dynamically among the three cells was confirmed by RT-PCR and Western blot. Then, BMX was transfected into K562 and K562/HHT cells, and the changes of daunorubicin (DNR) concentrations in these two cells were observed for BMX overexpression.

RESULTSAs compared with K562, there were changes in 117 gene expressions in K562/HHT, 57 of which were up-regulated and 60 down-regulated. The mdrl gene was significantly up-regulated. When compared with K562/HHT, 50 significantly differently expressed genes were screened out in the K562/HHT/RU486 cells, of which up- and down-regulated genes were 13 and 37 respectively. These genes involved in drug resistance, cell signaling, cell differentiation, cell proliferation, transcription regulator, ion transport and so on. Four genes [NM-001721 (BMX), NM-031459 (SESN2), NM-033642 (FGF13) and AL-049309 (SFRS12)] expressed significantly differently in the two group cells, BMX gene expression was higher in K562/HHT, than in K562, but lower than in K562/HHT/RU486 as confirmed by RT-PCR and Western blot. After the plasmid pCI-neo-BMX was transfected into K562 and K562/HHT cells, DNR concentration was significantly lower (79.28 +/- 4.04, 29.84 +/- 2.67) than those before transfection (158.52 +/- 8.08, 58.58 +/- 6.53).

CONCLUSIONBMX is associated with multi-drug resistance of K562/HHT cell line.

Drug Resistance, Multiple ; drug effects ; genetics ; Drug Resistance, Neoplasm ; drug effects ; genetics ; Gene Expression Profiling ; Harringtonines ; pharmacology ; Humans ; K562 Cells ; Leukemia ; drug therapy ; genetics ; metabolism ; Protein-Tyrosine Kinases ; genetics ; metabolism

OBJECTIVETo investigate the role of Mcl-1 gene in resistance of all-trans retinoic acid (ATRA) of leukemia cells.

METHODSLong-term, intermittent and repetitive exposure of HL-60 cells to ATRA was used to establish a multidrug-resistance cell line (HL-60/ATRA). HL-60/ATRA cells were transfected with Mcl-1 small interference RNA (siRNA) by Lipofectamine 2000. Western blot was used to detect the expression of Mcl-1. The proliferation, apoptosis and differentiation were evaluated by MTT assay, in situ nick end-labeling (TUNEL) and NBT assay, respectively.

RESULTSThe HL-60/ATRA could keep its undifferentiated and proliferative status to a high concentration of ATRA (100 nmol/L) with highly expressed Mcl-1 protein (relative grey scale 0.624 +/- 0.127). Mcl-1 gene knockdown by siRNA (relative grey scale 0.267 +/- 0.086) could reverse the resistance of ATRA of HL-60/ATRA by inhibiting proliferation, and inducing differentiation and apoptosis [apoptosis rate (18.5 +/- 4.5)%].

CONCLUSIONMcl-1 gene might be involved in ATRA resistance in HL-60 cells and inhibiting its expression could be a new approach to ATRA resistance reversion.

Apoptosis ; drug effects ; genetics ; Cell Differentiation ; drug effects ; genetics ; Cell Proliferation ; drug effects ; Drug Resistance, Neoplasm ; genetics ; HL-60 Cells ; drug effects ; metabolism ; Humans ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; RNA, Small Interfering ; Tretinoin ; pharmacology

This study was purposed to investigate the effects of angelicasinensis (Oliv) Diel compound injection, vauqueline, ephedrine and strychnine on human erythroleukemia multidrug resistance (MDR) K562/A02 cell line Mdr1 gene and p-glycoprotein. The MTT and trypan blue methods were used to analyze the cytotoxic effect of above-mentioned traditional Chinese drug; the expressions of K562/A02 cells Mdr1 gene and p-glycoprotein were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and Western blot respectively. The results showed that after K562/A02 cell were treated with angelicasinensis (Oliv) diels compound injection, vauqueline and ephedrine, mRNA transcription of Mdr1 gene was reduced significantly (p < 0.01); the expression of P-gp also decreased (p < 0.01). The expression level P-gp in group treated with vauqueline was the lowest, but the Mdr1 mRNA level and expression of P-gp of K562/A02 cells treated with strychnine did not obviously changed. It is concluded that angelicasinensis (Oliv) diels compound injection, vauqueline, ephedrine can partly reverse the multidrug resistance of K562/A02 cells, the down-regulation of Mdr1 gene causing decrease of p-glycoprotein expression may be one of the MDR reversal mechanisms in K562/A02 cells.
ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Down-Regulation ; drug effects ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Humans ; K562 Cells ; RNA, Messenger ; genetics ; metabolism

BACKGROUND AND OBJECTIVECurcumin, a natural compound, is derived from the rthizom of Curcuma longa. In vitro and in vivo preclinical studies have shown its anti-inflammatory, antioxidant, anticancer activities and so on. miR-186*, which was found by microarray technology, was highly expressed in lung carcinoma cells A549/DDP. The aim of this study is to illustrate whether Curcumin could promote the apoptosis of A549/DDP cells through regulating the expression of miR-186*.

METHODSAn oligonucleotide microarray chip was used to profile microRNA (miRNA) expressions in A549/DDP cells treated with and without Curcumin. The significantly differentially expressed miRNA, which was selected from microarray chip, validated by quantitative real-time PCR. Ultimately, the remarkably expressed miRNA modulated the apoptosis assaying by flow cytometry expriments and the survival rate was measured by MTT method.

RESULTSThe microarray chip results demonstrated: Curcumin altered the expression level of miRNAs compared with untreated control in A549/DDP cell line, miR-186* was significantly down-regulated after Curcumin treatment, which confirmed by quantitative real-time PCR. Down-regulation of miR-186* expression by curcumin elevated the apoptosis, and the survival rate of A549/DDP cells decreased; but up-regulation of miR-186* expression by transfection its mimics restrained the apoptosis, the survival rate of A549/DDP cells increased, which were assayed by flow cytometry expriments and MTT method.

CONCLUSIONModulation of miRNAs expression may be an important mechanism underlying the biological roles of Curcumin.

Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; genetics ; Cell Line, Tumor ; Cisplatin ; pharmacology ; Curcumin ; pharmacology ; Drug Resistance, Neoplasm ; genetics ; Humans ; Lung Neoplasms ; drug therapy ; genetics ; MicroRNAs ; genetics