Animals ; Epilepsy ; genetics ; metabolism ; Genes, MDR ; Hippocampus ; drug effects ; metabolism ; Male ; Rats ; Rats, Wistar ; Triazines ; pharmacology

This study investigated the reversal effect of isotetrandrine, an isoquinoline alkaloid extracted from Caulis mahoniae, on P-glycoprotein-mediated multidrug resistance in human breast cancer doxorubicin-resistant (MCF-7/DOX) cells. RT-PCR assay and immunity histochemistry assay were used to determine the expression level of mdrl gene and P-gp in MCF-7/DOX cells to elucidate resistant character of MCF-7/DOX cells. The activity of isotetrandine to enhance doxorubicin cytotoxicity was tested using MTT (3-(4, 5-dimethyhthiazol)-2,5 -diphenyltetrazolium bromide) assay and was evaluated by the reversal fold (RF) values. Intracellular accumulation of doxorubicin was assessed by the determination of doxorubicin-associated fluorescence intensity. Effect of isotetrandrine on the expression level of P-gp in MCF-7/DOX cells was then determined by immunity histochemistry assay. The ability of isotetrandrine to inhibit P-gp function was evaluated by detecting the accumulation and efflux of rhodamine 123 (Rh123) with flow cytometry (FCM). Verapamil was employed as a comparative agent in whole experiment. The results indicated that MCF-7/DOX cells had phenotype of MDR and that the positive expression of P-gp was their resistant character. 10 microg x mL(-1) isotetrandrine could distinctly enhance cytotoxicity of DOX in MCF-7/DOX cells and reversal fold (RF) was significantly higher than that of verapamil (P < 0.05), but it hardly affected cytotoxicity of DOX in MCF-7 cells and the expression level of P-gp in MCF-7/DOX cells. The ability of isotetrandrine to inhibit P-gp function was reversible, because incubation of MCF-7/DOX cells with isotetrandrine caused a marked increase in uptake and a notable decrease in efflux of Rh123 and a marked increase of intracellular DOX concentrations. In conclusion, isotetrandrine exhibited potent effect on the reversal of P-gp-mediated MDR in vitro, suggesting that it might become a candidate of effective MDR reversing agent in cancer chemotherapy.
ATP-Binding Cassette, Sub-Family B, Member 1 ; metabolism ; Antineoplastic Agents, Phytogenic ; isolation & purification ; pharmacology ; Benzylisoquinolines ; isolation & purification ; pharmacology ; Breast Neoplasms ; pathology ; Cell Line, Tumor ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Gene Expression Regulation, Neoplastic ; Genes, MDR ; Humans ; Mahonia ; chemistry ; Plant Stems ; chemistry ; Plants, Medicinal ; chemistry ; Rhodamine 123 ; metabolism

Background: Acute lower respiratory tract infection, mainly pneumonia, were the main reasons cause death for children under 5 years old. Objectives: Determine the isolated rate of bacteria inpatients under 5 years old with acute lower respiratory tract infection in Ha Noi and antibiotic resistance of pneumococcal isolated form patients. Subjects and method: Patients under 5 years old with acute lower respiratory tract infection in National hospital of pediatrics and Bach Mai hospital from 01/2002. Using quantitative culturedand PCR method. Results: Out of total 164 patients with lower respiratory tract infection, there were 91 diagnosed pneumonia by chest X-ray, 73 cases of acute bronchitis. 73,6% of the pneumococcal isolated were penicillin resistance (gPRSP) with different genes such as pbp 1a+2x+ab. Most of the S.pneumoniae strains were serotype 19F or 23F. There were no statistic differences by comparison charactersistics of weight, vessel, subclinical symptoms such as: dissolved oxygen level (S\xac\xacp\xac\xac\xac\xacO\xac2\xac), the amount of leucocyte in blood. However, temperature of pneumonia patients was higher than bronchitis patients, breathing of pneumonia patients was also faster than bronchitis patients. Isolated bacteria with amount \ufffd?106 cfu/ml was H.influenzae, S.pneumoniae and Moraxell catarrhalis in pneumonia group, bronchitis group was 28,8% and control group was 17,1%. Conclusion: Penicillin, erythoromycin and co-trimoxazole resistance rate of S.pneumoniaein patients with acute lower respiratory tract infection was high. Quantitative cultured method has prognostic value in diagnosis pneumonia.
Genes ; MDR/ drug effects ; immunology ; Streptococcus pneumoniae/ growth & ; development ; Anti-Bacterial Agents

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

BACKGROUNDThe hematopoietic microenvironment (HM) plays a critical role in malignant cell growth, patient survival, and response to chemotherapy in hematologic malignancies. However, mechanisms associated with this environmental influence remain unclear. In this study, we investigated the role of bone marrow derived mesenchymal stem cells (MSCs) in U937 cell line, to find out the relations between leukemia drug resistance and the MSCs.

METHODSU937 cells were cultured in suspension or grew adherently with MSCs. The cell growth curve was drawn and the cell cycle was measured by flow cytometry. Apoptosis and sensitivity of U937 to daunoblastina (DNR) were quantified by DNA ladder detection and trypan blue exclusion assays, respectively. The gene expression profile chip technology was used to determine and analyze the changes in apoptosis-related gene expression after adherent culture and the expression of MDR1 mRNA was assessed by reverse transcriptional polymerase chain reaction (RT-PCR) at the same time.

RESULTSIn the adherent culture, the proliferation of the U937 cells was inhibited, the G0/G1 phase cells increased (F = 64.9726, P < 0.0001), G2/M phase cells were decreased (F = 98.1361, P < 0.0001) and the natural apoptosis rate was decreased (F = 24.0866, P < 0.0001) compared with those in the suspended culture. U937 cell viability was enhanced and cell apoptosis was blocked during DNR treatment in adherent culture with MSCs. Thirty-nine differently expressed genes were screened from the 487 apoptosis related genes in the adherent culture U937 cells. Among the 37 upregulated genes, Bcl-XL was upregulated most significantly. Two genes were downregulated. Adherent culture did not induce MDR1 mRNA expression in U937 cells.

CONCLUSIONSMSCs play a role in modulating the proliferation of U937 cells and response of U937 cells to DNR, and Bcl-XL apoptosis-inhibiting gene may be most important in determining the sensitivity of leukemic cells to treatment, which is not related to MDR1.

Apoptosis ; drug effects ; Bone Marrow Cells ; physiology ; Cell Proliferation ; Daunorubicin ; pharmacology ; Drug Resistance, Neoplasm ; Genes, MDR ; Humans ; Immunophenotyping ; Mesenchymal Stromal Cells ; physiology ; U937 Cells ; drug effects

OBJECTIVETo screen effective sequences of small interfering RNA targeting MDR1 gene in human gastric cancer SGC7901/VCR cells.

METHODSFour siRNAs (MDR1si326, MDR1si1513, MDR1si2631 and MDR1si3071) targeting MDR1 gene were designed and synthesized by in vitro transcription. The siRNA duplexes were used to transfect into the human gastric cancer SGC7901/VCR cells. The expression level of MDR1 mRNA and P-gp were detected by RT-PCR and Western blotting, respectively. The accumulation of intracellular adriamycin (ADR) was examined by flow cytometry and the cell sensitivity to ADR was demonstrated by MTT.

RESULTSThe SGC7901/VCR cells treated with 4 siRNAs led to reversal effect on multidrug resistance to different extents. Among the SGC7901/VCR cells treated by siRNAs for 48 h, the expression level of MDR1 mRNA in cells of MDR1si326 or MDR1si2631 group (0.42 +/- 0.07 or 0.49 +/- 0.02) was more decreased than that in cells of MDR1si1513 or MDR1si3071 group (P < 0.05). The accumulation of ADR in cells of MDR1si326 group was the most; in cells of MDR1si2631 group, more; in cells of MDR1si3071 group, lower and in cells of MDR1si1513 group, the lowest (P < 0.05). The relative reversal efficiency of cells of MDR1si2631 group to ADR was the highest and in cells of MDR1si326 group, higher (P < 0.05). There was no significant difference in the relative reversal efficiency between the cells of MDR1si1513 and MDR1si3071 groups (P > 0.05). The expression level of P-gp in cells of MDR1si326 group was the lowest among the SGC7901/VCR cells treated by siRNAs for 72 h.

CONCLUSIONThe MDR1si326 with most, MDR1si2631 with more, MDR1si3071 with less and MDR1si1513 with least reversal effects on MDR1 gene mediated multidrug resistance were found in the human gastric cancer SGC7901/VCR cells.

ATP Binding Cassette Transporter, Sub-Family B ; ATP-Binding Cassette, Sub-Family B, Member 1 ; biosynthesis ; genetics ; metabolism ; Antibiotics, Antineoplastic ; metabolism ; pharmacology ; Antineoplastic Agents, Phytogenic ; pharmacology ; Cell Line, Tumor ; Doxorubicin ; metabolism ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Genes, MDR ; Humans ; RNA, Messenger ; biosynthesis ; genetics ; RNA, Small Interfering ; pharmacology ; Stomach Neoplasms ; metabolism ; pathology ; Transfection ; Vincristine ; pharmacology

OBJECTIVETo observe the lethal effect of multidrug resistance gene (MDR1) antisense RNA combined with oxaliplatin and 5-FU on drug-resistant rectal carcinoma cells.

METHODSPC-MDR1 plasmid including MDR1 was constructed with gene cloning techniques. The drug-resistant cancer cells (8348R) were transferred with the plasmids, and the positive neoplasm cells were selected with G418. Green fluorescent protein (GFP) gene was used as a reporting gene to monitor the gene transfer efficiency under the influence of oxaliplatin and 5-FU. The cytotoxicity and therapeutic effects of MDR1 anti-sense RNA combined with oxaliplatin and 5-FU were evaluated by colony-forming rate and MTT assay.

RESULTSA significant decrease of biological activity was observed in 8348R cells transferred with PC-MDR1, cell cycles were blocked in S phase, or in G2/M phase, and apoptosis rate of the cells increased. With treatment of oxaliplatin, the plasmid transfer efficiency in the drug-resistant cancer cells was improved about 18 times. Using an IC(50) dose of oxaliplatin and 5-FU combined with (MDR1) anti-sense RNA, 75 percent of 8348R cells were killed, which was significant higher than that of the control cells.

CONCLUSIONSCombined MDR1 antisense RNA with oxaliplatin and 5-FU has a synergistic effect of killing drug-resistant cancer cells and may be a promising method for treating drug-resistant rectal carcinoma.

Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Combined Modality Therapy ; Drug Synergism ; Fluorouracil ; pharmacology ; Genes, MDR ; genetics ; Genetic Therapy ; Humans ; Organoplatinum Compounds ; pharmacology ; RNA, Antisense ; genetics ; Rectal Neoplasms ; genetics ; pathology ; therapy ; Transfection

BACKGROUNDRNA interference using short hairpin RNA (shRNA) can mediate sequence-specific inhibition of gene expression in mammalian cells. A vector-based approach for synthesizing shRNA has been developed recently. Overexpression of P-glycoprotein (P-gp), the MDR1 gene product, confers multidrug resistance (MDR) to cancer cells. In this study, we reversed MDR using shRNA expression vectors in a multidrug-resistant human breast cancer cell line (MCF-7/AdrR).

METHODSThe two shRNA expression vectors were constructed and introduced into MCF-7/AdrR cells. Expression of MDR1 mRNA was assessed by RT-PCR, and P-gp expression was determined by Western Blot and immunocytochemistry. Apoptosis and sensitization of the breast cancer cells to doxorubicin were quantified by flow cytometry and methyl thiazolyl tetrazolium (MTT) assays, respectively. Cellular daunorubicin accumulation was assayed by laser confocal scanning microscopy (LCSM). Statistical significance of differences in mean values was evaluated by Student's t tests. P < 0.05 was considered statistically significant.

RESULTSIn MCF-7/AdrA cells transfected with MDR1-A and MDR1-B shRNA expression vectors, RT-PCR showed that MDR1 mRNA expression was reduced by 40.9% (P < 0.05), 30.1% (P < 0.01) (transient transfection) and 37.6% (P < 0.05), 28.0% (P < 0.01) (stable transfection), respectively. Western Blot and immunocytochemistry showed that P-gp expression was significantly and specifically inhibited. Resistance against doxorubicin was decreased from 162-fold to 109-fold (P < 0.05), 54-fold (P < 0.01) (transient transfection) and to 108-fold (P < 0.05), 50-fold (P < 0.01) (stable transfection). Furthermore, shRNA vectors significantly enhanced the cellular daunorubicin accumulation. The combination of shRNA vectors and doxorubicin significantly induced apoptosis in MCF-7/AdrR cells.

CONCLUSIONSshRNA expression vectors effectively reduce MDR expression in a sustained fashion and can restore the sensitivity of drug-resistant cancer cells to conventional chemotherapeutic agents.

ATP-Binding Cassette, Sub-Family B, Member 1 ; analysis ; antagonists & inhibitors ; Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Survival ; drug effects ; Daunorubicin ; pharmacokinetics ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; Flow Cytometry ; Genes, MDR ; Genetic Vectors ; Humans ; RNA Interference ; RNA, Small Interfering ; genetics ; Transfection

OBJECTIVETo investigate the role of Ras antisense oligoribonucleotide (ASODN) in multidrug resistance (MDR) of pancreatic carcinoma Pc-2 cells.

METHODSRas and P-gp expression was suppressed by Ras ASODN. Sensitivity of Pc-2 cells to chemotherapy was determined by the MTT assay. MDR-1 mRNA level was detected by fluorogenic probe quantitative reverse transcription polymerase chain (RT-PCR) method. Flow cytometry (FCM) was used to detect the accumulative concentration of adriamycin (ADR) in the cells.

RESULTSRas ASODN significantly inhibited the Ras and P-gp expression (P < 0.05), increased the sensitivity of Pc-2 cells to chemotherapeutic agents (P < 0.05), decreased MDR-1 gene level in Pc-2 cells (P < 0.05), and increased the intracellular intake of ADR in Pc-2 cells (P < 0.05).

CONCLUSIONRas ASODN may enhance the sensitivity of multidrug-resistant pancreatic cancer Pc-2 cells to chemotherapeutic agents by regulating MDR-1 gene level.

ATP-Binding Cassette, Sub-Family B, Member 1 ; biosynthesis ; Cell Line, Tumor ; Down-Regulation ; Doxorubicin ; metabolism ; Drug Resistance, Multiple ; drug effects ; genetics ; Drug Resistance, Neoplasm ; drug effects ; genetics ; Genes, MDR ; drug effects ; genetics ; Humans ; Oligonucleotides, Antisense ; pharmacology ; Pancreatic Neoplasms ; genetics ; metabolism ; pathology ; ras Proteins ; biosynthesis ; genetics

Researches on MDR (multidrug resistance) of tumor presently focus on seeking chemosensitizers with more targets, high efficiency and low toxicity from traditional Chinese medicine. This paper reviews the research progress in the reversion of MDR of leukemia, hepatocarcinoma, breast carcinoma and oral epithelioid neoplasia by TDM compound, its extracts, its groups of active ingredients or its active ingredients.
ATP-Binding Cassette, Sub-Family B, Member 1 ; biosynthesis ; genetics ; Animals ; Breast Neoplasms ; metabolism ; Carcinoma, Hepatocellular ; metabolism ; DNA Topoisomerases, Type II ; metabolism ; Drug Combinations ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Genes, MDR ; Humans ; Leukemia ; metabolism ; Liver Neoplasms ; metabolism ; Medicine, Chinese Traditional ; Multidrug Resistance-Associated Proteins ; metabolism ; Plants, Medicinal ; chemistry ; RNA, Messenger ; biosynthesis ; genetics