OBJECTIVES: Nephrotic syndrome is a common disease of the urinary system. The aim of this study is to explore the effect of astragalus polysaccharides (APS) on multidrug resistance gene 1 (MDR1) and P-glycoprotein 170 (P-gp170) in adriamycin nephropathy rats and the underlying mechanisms. METHODS: A total of 72 male Wistar rats were divided into a control group, a model group, an APS low-dose group, an APS high-dose group, an APS+micro RNA (miR)-16 antagomir group and an APS+miR-16 antagomir control group, with 12 rats in each group. Urine protein (UP) was detected by urine analyzer, and serum cholesterol (CHOL), albumin (ALB), blood urea nitrogen (BUN), and creatinine (SCr) were detected by automatic biochemical analyzer; serum interleukin-6 (IL-6), IL-1β, tumor necrosis factor α (TNF-α) levels were detected by ELISA kit; the morphological changes of kidney tissues were observed by HE staining; the levels of miR-16 and MDR1 mRNA in kidney tissues were detected by real-time RT-PCR; the expression levels of NF-κB p65, p-NF-κB p65, and P-gp170 protein in kidney tissues were detected by Western blotting; and dual luciferase was used to verify the relationship between miR-16 and NF-κB. RESULTS: The renal tissue structure of rats in the control group was normal without inflammatory cell infiltration. The renal glomeruli of rats in the model group were mildly congested, capillary stenosis or occlusion, and inflammatory cell infiltration was obvious. The rats in the low-dose and high-dose APS groups had no obvious glomerular congestion, the proliferation of mesangial cells was significantly reduced, and the inflammatory cells were reduced. Compared with the high-dose APS group and the APS+miR-16 antagomir control group, there were more severe renal tissue structure damages in the APS + miR-16 antagomir group. Compared with the control group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 in the model group were significantly increased (all P<0.05); the levels of ALB and miR-16 were significantly decreased (both P<0.05). Compared with the model group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of pNF-κB p65 and P-gp170 in the low-dose and high-dose APS groups were significant decreased (all P<0.05); and the levels of ALB and miR-16 were significantly increased (both P<0.05). Compared with APS+miR-16 antagomir control group, the UP, CHOL, BUN, SCr, IL-6, IL-1β, and TNF-α levels, MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 were significantly increased (all P<0.05). The levels of ALB and miR-16 were significantly decreased in the APS+miR-16 antagomir group compared with the APS+miR-16 antagomir control group (both P<0.05). CONCLUSIONS APS can regulate the miR-16/NF-κB signaling pathway, thereby affecting the levels of MDR1 and P-gp170, and reducing the inflammation in the kidney tissues in the adriamycin nephropathy rats.
ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics* ; Animals ; Antagomirs ; Doxorubicin/toxicity* ; Genes, MDR ; Interleukin-6/metabolism* ; Kidney Diseases/genetics* ; Male ; MicroRNAs/metabolism* ; NF-kappa B/metabolism* ; Polysaccharides/pharmacology* ; RNA, Messenger ; Rats ; Rats, Wistar ; Tumor Necrosis Factor-alpha/metabolism*

BACKGROUND: The aim of the present study was to determine the frequency of six efflux pump genes in Acinetobacter clinical isolates collected from South Korean hospitals. METHODS: In this study, we used a total of 339 Acinetobacter strains, comprising 279 Acinetobacter calcoaceticus–Acinetobacter baumannii (ACB) complex and 60 non-ACB complex strains. We performed specific PCR assays to detect adeG, adeB, adeE, adeY, abeM, and adeJ, transporter genes of the multidrug efflux pumps AdeFGH, AdeABC, AdeDE, AdeXYZ, AbeM, and AdeIJK, respectively. RESULTS: Frequencies of six efflux pump genes varied according to the species of Acinetobacter. Frequencies of adeE, abeM, and adeJ between A. baumannii group and A. nosocomialis group were found to be significantly different. Significant differences were found in the frequencies of adeB, adeE, adeY, and adeJ among the susceptible A. baumannii (SAB), multidrug-resistant A. baumannii (MDRAB), and extensively drug-resistant A. baumannii (XDRAB) groups within the 154 strains of A. baumannii. The frequencies of efflux pump genes in imipenem-susceptible and imipenem-nonsusceptible groups were significantly different for adeB, adeY, and adeJ. The frequencies of efflux pump genes in ciprofloxacin-susceptible and ciprofloxacin-nonsusceptible groups were significantly different for adeB and adeY. No significant difference was found in the frequency of efflux pump genes among groups sampled from different regions of Korea, across 86 strains of A. baumannii collected in 2012. CONCLUSIONS: The frequencies of six efflux pump genes obtained in this study demonstrate the fundamental epidemiological feature of efflux pump genes in Korean Acinetobacter clinical isolates.
Acinetobacter ; Gene Frequency ; Genes, MDR ; Korea ; Polymerase Chain Reaction

BACKGROUND: Carbapenem-resistant Klebsiella pneumoniae (CRKP) is considered a serious global threat. However, little is known regarding the multidrug resistance (MDR) mechanisms of CRKP. This study investigated the phenotypes and MDR mechanisms of CRKP and identified their clonal characteristics. METHODS: PCR and sequencing were utilized to identify antibiotic resistance determinants. Integron gene cassette arrays were determined by restriction fragment length polymorphism (RFLP) analysis. Multi-locus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) were used for epidemiological analysis. Plasmids were typed by using a PCR-based replicon typing and analyzed by conjugation and transformation assays. RESULTS: Seventy-eight strains were identified as resistant to at least one carbapenem; these CRKP strains had a high prevalence rate (38.5%, 30/78) of carbapenemase producers. Additionally, most isolates harbored MDR genes, including Extended spectrum β-lactamases (ESBLs), AmpC, and quinolone and aminoglycoside resistance genes. Loss of porin genes was observed, and Class 1 integron was detected in 66.7% of the investigated isolates. PFGE and MLST results excluded the occurrence of clonal dissemination among these isolates. CONCLUSIONS: A high prevalence of NDM-1 genes encoding carbapenem resistance determinants was demonstrated among the K. pneumoniae isolates. Importantly, this is the first report of bla(NDM-1) carriage in a K. pneumoniae ST1383 clone in China and of a MDR CRKP isolate co-harboring bla(NDM-1), bla(KPC-2), bla(CTX-M), bla(SHV), acc(6′)-Ib, rmtB, qnrB, and acc(6′)-Ib-cr.
China* ; Clone Cells ; Drug Resistance, Bacterial ; Drug Resistance, Microbial ; Drug Resistance, Multiple* ; Electrophoresis, Gel, Pulsed-Field ; Genes, MDR ; Integrons ; Klebsiella pneumoniae* ; Klebsiella* ; Molecular Epidemiology ; Phenotype ; Plasmids ; Pneumonia ; Polymerase Chain Reaction ; Polymorphism, Restriction Fragment Length ; Prevalence ; Replicon

The efflux pump system has been suggested as an important mechanism in the drug resistance of Mycobacterium tuberculosis (MTB). In this study, molecular analysis of five genes in the efflux pump system of MTB isolates from Korean patients was performed in order to identify appropriate molecular targets. In this study, 35 culture-positive specimens were included. PCR was performed for five efflux genes, mmpL7, efpA, mmr, p55 and tap-like gene. In the 35 clinical isolates, molecular analysis of five kinds of efflux pump genes was performed. Only one clinical isolate showed negative PCR results for all five efflux pump genes. All the rest 34 isolates presented concurrent positive results for the five efflux pump genes. In the near future, gene expression study with quantitative PCR should be performed using these genes.
Drug Resistance ; Gene Expression ; Genes, MDR ; Humans ; Mycobacterium tuberculosis* ; Polymerase Chain Reaction

Multidrug resistance-related proteins (MRP) are identified as ATP-dependent efflux pumps, the expression abnormality of which has close relationship with multidrug resistance (MDR) in a variety kinds of malignancies, leading to the failure of chemotherapy. The relationship between the expression of MRP and acute leukemia remains to be proved, since experiments and clinical researches are still insufficient. In this article, the structure, distribution, function, and relation of MRP to MDR and prognosis of acute leukemia are reviewed.
Acute Disease ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Genes, MDR ; Humans ; Leukemia ; genetics ; Proteins ; genetics

BACKGROUND: Multi-drug resistant (MDR) Acinetobacter baumannii has emerged as a significant infectious agent in hospitals worldwide. The purpose of this study was to determine the molecular characterization of MDR A. baumannii clinical isolates. METHODS: Two hundred eighty-five strains of non-duplicated A. baumannii collected from March to November 2011 from a university hospital laboratory located in the Wonju area of the Gangwon province of Korea were analyzed for MDR genes. RESULTS: All of the 285 imipenem-resistant A. baumannii isolates were encoded by a blaOXA-23-like gene, and all isolates with the blaOXA-23-like gene had the upstream element ISAba1. The 16S rRNA methylase gene armA was detected in 153 (50.2%) clinical isolates, but rmtA, rmtB, rmtC, rmtD and npmA were not detected in any isolates in the present study. The gene encoding aac(6')-Ib was the most prevalent aminoglycoside-modifying enzyme. The sequencing data for the quinolone resistance-determining region of gyrA and parC revealed the presence of Ser (TCA) 83 to Leu (TTA) and Ser (TCG) 80 to Leu (TTG) substitutions. All but one of the 285 A. baumannii isolates showed similar band patterns on repetitive extragenic palindromic-PCR profiles. CONCLUSION: The molecular characteristics of the resistance genes of MDR A. baumannii isolates obtained from the Wonju area of Gangwon province were similar to those of other areas in Korea.
Acinetobacter ; Acinetobacter baumannii ; beta-Lactamases ; Genes, MDR ; Imipenem ; Korea ; Laboratories, Hospital ; Methyltransferases

BACKGROUNDMyelosuppression is the main dose-related toxicity of many chemotherapeutic drugs. The human multidrug resistance (mdr1) gene is well-known for its ability to confering drug resistance. In this study, we meant to transplant the placenta mesenchymal stem cells (P-MSCs) moderated by mdr1 gene into a nude mice model radiated by γ-Co(60) and to explore the chemoprotection for bone marrow (BM) toxicity.

METHODSHuman P-MSCs were isolated from trypsin-digested term placentas and then transduced by with reconstructed retroviral vector containing mdr1 gene and green fluorescent protein (GFP) reporter gene. The integration and expression of mdr1 gene was observed indirectedly by the expression of GFP. A nude mice model was constructed after irradiation with a sublethal dosage of γ-Co(60). These irradiated mice were transplanted with mdr1-MSCs through the caudal vein and then received paclitaxel (PAC) intraperitoneal chemotherapy. The Peripheral peripheral blood (PB) of the nude mice was collected, and the PB cells counts and values were determined using an automatic analyzer.

RESULTSAfter PAC treatment, mdr1-MSCs transplanted mice showed markedly improved survival upon compared to MSCs transplanted mice (85.7% vs. 57.1%). White blood cell (WBC) and red blood cell (RBC) counts as well as the hemoglobin (Hb) values were significantly increased in PAC treated mdr1-MSCs mice compared to PAC treated control mice when PAC chemotherapy had been finished (all P < 0.05), but the difference was not found in the plateltes (PLT) count (P > 0.05).

CONCLUSIONHuman P-MSCs moderated by mdr1 gene when transplanted into nude mice may provide chemoprotection for hematopoietic toxicity.

Animals ; Bone Marrow ; Cell Differentiation ; genetics ; physiology ; Cells, Cultured ; Erythrocytes ; metabolism ; Female ; Genes, MDR ; genetics ; physiology ; Green Fluorescent Proteins ; genetics ; metabolism ; Hemoglobins ; metabolism ; Humans ; Leukocytes ; metabolism ; Mesenchymal Stromal Cells ; cytology ; metabolism ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Placenta ; cytology ; Pregnancy

OBJECTIVETo investigate whether there is intercellular transfer of functional P-glycoprotein(P-gp) from P-gp-positive cells to P-gp-negative cells in vitro.

METHODSHepG2/GFP cells, a HepG2 cell line stably expressing GFP, were co-cultured with HepG2/ADM cells, an adriamycin-resistant cell line derived from HepG2 cells. The distribution of P-gp in hepatocellular carcinoma cell was observed under laser scanning confocal microscope (LSCM). Immunomagnetic beads were used to separate HepG2/GFP cells from the mixed culture. The abundance of P-gp was analyzed by western blot, and the expression of mdr1 mRNA was detected by qRT-PCR.

RESULTSYellow fluorescence was detected in HepG2/aqMDR cells, green fluorescence was detected in HepG2/GFP cells, red fluorescence was detected in HepG2/ADM cells by LSCM. The level of P-gp protein in HepG2/aqMDR cells was lower than that in HepG2/ADM cells, but higher than that in HepG2/GFP cells (q = 35.07, P < 0.05) and HepG2 cells (q = 36.87, P < 0.05). The expression of mdr1 mRNA in HepG2/ADM cells was higher than that in HepG2/aqMDR, HepG2 and HepG2/GFP cells, but there was no significant difference in mdr1 mRNA among HepG2/aqMDR, HepG2 and HepG2/GFP cells (F = 2.30, P > 0.05).

CONCLUSIONSP-gp can transfer from drug resistant hepatocellular cells to sensitive hepatocellular carcinoma cells. This study suggests a novel mechanism of multidrug resistance in hepatocellular carcinoma.

ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; metabolism ; Carcinoma, Hepatocellular ; genetics ; metabolism ; pathology ; Coculture Techniques ; methods ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; genetics ; Genes, MDR ; Green Fluorescent Proteins ; Hep G2 Cells ; Humans ; Liver Neoplasms ; genetics ; metabolism ; pathology ; Plasmids ; Protein Transport ; RNA, Messenger ; genetics ; metabolism ; Transfection

This study was aimed to design and screen short hairpin RNA (shRNA) molecules targeting multidrug resistance gene (mdr1), as well as to investigate the effects of shRNA expression vector on K562/A02 cells. Mdr1-shRNA expression vector was transfected into K562/A02 cells by lipofectamine 2000, and G418 was added to screen and establish the stable expression cell strain. The expressions of mdr1 mRNA and protein were detected by real-time RT-PCR and Western blot respectively. The sensitivity of cells to chemodrugs after interference were tested by CCK8 assay. The function of p-glycoprotein was determined by Rhodamine 123 efflux experiment. The results showed that all of 4 mdr1-shRNA expression vectors could significantly knockdown the expression of p-glycoprotein as compared with control vector, moreover, the vector targeting 508 - 526 sites of mdr1 gene was the best one. It is concluded that the mdr1-shRNA expression vector gained by screening can significantly knockdown the expression of mdr1 gene and reverse leukemia drug resistance, paving the way for the application of RNAi in the following animal experiments.
ATP Binding Cassette Transporter, Sub-Family B ; ATP-Binding Cassette, Sub-Family B, Member 1 ; genetics ; Base Sequence ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Gene Knockdown Techniques ; Genes, MDR ; Genetic Vectors ; Humans ; K562 Cells ; Leukemia ; genetics ; RNA Interference ; RNA, Messenger ; genetics ; RNA, Small Interfering ; genetics ; Transfection

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