OBJECTIVE: To explore the genetic etiology for a pedigree affected with progressive familial intrahepatic cholestasis (PFIC). METHODS: Target sequence capture and next generation sequencing (NGS) were applied for the proband. PCR and Sanger sequencing were used to verify the suspected mutation in his sister with similar symptoms and his parents. RESULTS: The proband and his sister manifested after birth with symptoms including jaundice, pruritus and developmental retardation. NGS has identified compound heterozygous mutations of ABCB11 gene, which encodes bile salt export pump protein (BSEP), namely c.2494C>T (p.Arg832Cys) and c.3223C>T (p.Gln1075*), in the proband, which were inherited from his father and mother respectively. His sister carried the same compound mutations. CONCLUSION Based on the phenotype and genetic testing, the patients were diagnosed as PFIC2 caused by mutation of the ABCB11 gene. The c.3223C>T is a novel nonsense mutation which may cause premature termination of translation. Above results have enriched the spectrum of ABCB11 mutations and provided new evidence for the molecular basis of PFIC, which also facilitated genetic counseling for this pedigree.
ATP Binding Cassette Transporter, Subfamily B, Member 11 ; genetics ; ATP-Binding Cassette Transporters ; Cholestasis, Intrahepatic ; genetics ; Female ; Genetic Testing ; Humans ; Male ; Mutation ; Pedigree ; Phenotype

OBJECTIVE: To study the association between the expression of the MDR3 gene and the pathogenesis of parenteral nutrition-associated cholestasis (PNAC) in preterm infants. METHODS: Among the preterm infants who were admitted to the hospital from June 2011 to November 2017 and received parenteral nutrition for more than 14 days, 80 who did not develop PNAC were enrolled as non-PNAC group, and 76 who developed PNAC were enrolled as PNAC group. On days 1, 14, 30, 60 and 90 after birth, serum hepatobiliary biochemical parameters [alanine aminotransferase (ALT), total bilirubin (TBil), direct bilirubin (DBil), total bile acid (TBA) and gamma-glutamyl transpeptidase (γ-GT)], fibrosis indices [hyaluronic acid, laminin, procollagen III N-terminal peptide and type IV collagen] and clinical manifestations were observed. Real-time quantitative PCR was used to measure the mRNA expression of MDR3 in both groups, and the correlation between the mRNA expression of MDR3 and serum hepatobiliary biochemical parameters was analyzed. RESULTS: In the PNAC group, serum levels of hepatobiliary biochemical parameters and fibrosis indices increased on day 14 after birth and reached the peak on day 30 after birth, followed by a reduction on day 60 after birth. On days 14, 30, 60 and 90 after birth, the PNAC group had significantly higher serum levels of hepatobiliary biochemical parameters and fibrosis indices than the non-PNAC group (P<0.05). The PNAC group had higher relative mRNA expression of MDR3 in peripheral blood cells than the non-PNAC group (P<0.05). In the PNAC group, the relative mRNA expression of MDR3 in peripheral blood cells was negatively correlated with serum levels of hepatobiliary biochemical parameters (ALT, TBil, DBil, TBA and γ-GT) (P<0.001). CONCLUSIONS High mRNA expression of MDR3 in preterm infants may be associated with the development of PNAC, and further studies are needed to identify the mechanism.
ATP Binding Cassette Transporter, Subfamily B ; genetics ; Cholestasis ; genetics ; Humans ; Infant, Newborn ; Infant, Premature ; Parenteral Nutrition ; RNA, Messenger

Drug transport is an important source of inter-individual variations in drug responses and is also a common site where drug-drug interactions happen. In recent years, more and more novel identified transporters have been added into the transporter super family, and this trend will continue in the future. Among the transporter members of this family, ATP-dependent efflux transporter P-glycoprotein (MDR1) and organic anion transporters (OATP) are the most important proteins involved in drug transport. MDR1 is the most well known transporter. Widely distributed in tissues such as the gastrointestinal tract, liver, kidney and so on, MDR1 plays an important role in drug absorption, distribution and excretion. Its functional genetic polymorphisms have significantly changed the pharmacokinetics of its substrate drugs, which has important clinical implications. OATP expressed in multiple tissues, and it mediated the drug excretion through the bile acid and kidney. Some genetic polymorphism of OATP genes is the cause of some abnormal drug responses.
ATP Binding Cassette Transporter, Subfamily B, Member 1 ; genetics ; Drug Interactions ; genetics ; Humans ; Membrane Transport Proteins ; genetics ; metabolism ; Organic Anion Transporters ; genetics ; Pharmaceutical Preparations ; metabolism ; Polymorphism, Genetic

OBJECTIVE: To analyze the relation between the signle nucleotide polymorphisms (SNP) of CYP3A5 gene and MDR1 gene loci and the risk of cytogenetic relapse in chronic myeloid leukemia (CML). METHODS: The clinical data of 90 patients with CML treated with imatinib in our hospital were collected.The patients were divided into 2 groups: non-relapse and relapse according to relapse and non-relapse, then the relation between the SNP of CYP3A5 gene and MRD1 gene loci and the risk of cytogenetic relapse in CML patients. RESULTS: The grouping result showed that the patients with non cytogenetic relapse accounted for 41 cases those were enrolled in non-relapse group, and patient-with cytogenetic relapse accounted for 49 cases those were enrolled in relapse group. The follow-up time was 36 months. The detection showed that the incidence of cytogenetic relapse in the patients with CC genotype was significantly higher than that in the patients with TT+CT genotype of C3435T and C1236T at MDR1 gene loci (P<0.05).Compared with the patients with CT+CC genotype in C3435T locus of MDR1 gene, the rate of cytogenetic relapse in the patients with TT genotype decreased significantly (P<0.05). Compared with patients with CT+CC phemotype of C3435T in MDR1 gene locus, the non-relapse survival time of TT genotypes was significantly prolonged (P<0.05). Compared with non-relapse group, the incidence of neutropenia (29.27% vs 71.43%) and blood toxicity (39.02% vs 61.22%) in the relapse group increased significantly (P<0.05). The imatinib dose (OR=2 95, 95% CI:1.37~7.76) and the C3435T genotype in MDR1 genes (OR=0.09, 95% CI:0.05~0.72) were the factors affecting the cytogenetic relapse of the patients with CML (both P<0.05). CONCLUSION The therapeutic dose of imatinib and the C3435T and C1236T genotypes in MDR1 gene have a certain effect on the cytogenetic relapse of CML patients. C3435T genotypes in the.MDR1 gene showed a certain predictive value for evaluating the risk of cytogenetic relapse, which can be used as a clinical biomarker.
ATP Binding Cassette Transporter, Subfamily B ; genetics ; Cytochrome P-450 CYP3A ; genetics ; Genotype ; Humans ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; genetics ; Polymorphism, Single Nucleotide ; Recurrence

Objective To explore the role of multidrug resistance gene-1(MDR1)gene in methotrexate(MTX)resistance in patients with rheumatoid arthritis(RA).Methods Fibroblast-like synoviocytes(FLS)from RA patients were infected with recombinant adenovirus Ad-EGFP-MDR1 to obtain MDR1 over-expressed RA FLS.The transcription level of MDR1 gene and the expression level of its coding product P-glycoprotein(P-gp) rotein were detected by real-time PCR and Western blot analysis.The efflux function was verified by rhodamine 123 efflux assay.The resistance to MTX was detected by MTT assay.Results RA FLS were infected with recombinant adenovirus Ad-EGFP-MDR1;72 hours later,the particles size in MDR1 over-expressed RA FLS increased,the cell volume became larger,and the growth rate decreased.The transcription level of MDR1(1.4325±0.3924 0.0650±0.0070;=6.035,=0.004),the expression level of P-gp protein(1.8667±0.2857 0.9367±0.0551;=5.536,=0.005),and the ability of extracellular rhodamine 123(979.43±196.81 1680.06±147.04;=-4.940,=0.008) in MDR1 over-expressed RA FLS were significantly higher than those of negative virus control RA-FLS,and the survival rate of MDR1 over-expressed RA FLS was significantly increased at each concentration of MTX(<0.05).Conclusion The high expression of MDR1 can affect the efflux ability to MTX by up-regulating the expression of P-gp,thus enhancing the drug resistance to MTX in RA FLS.
ATP Binding Cassette Transporter, Subfamily B ; genetics ; Arthritis, Rheumatoid ; drug therapy ; genetics ; Cells, Cultured ; Drug Resistance ; Fibroblasts ; drug effects ; Humans ; Methotrexate ; pharmacology ; Synovial Membrane ; cytology

OBJECTIVE: To determine the effect of different heating Methods combined with neferine(Nef) on the proliferation and expressions of γH2AX and mdr-1/P-gp in MCF-7/Adr breast cancer cells. METHODS: MTT assay was used to determine block heating, water submerged heating, medium heating, and oven heating combined with 10 μg/mL Nef on adriamycin cultured MCF-7/Adr cell proliferation. The mdr-1mRNA expression was detected by real-time quantitative PCR. γH2AX and P-gp expressions were detected by Western blot. RESULTS: The absorbance values of MCF-7/Adr cells in different heating groups at 42 degree and 45 degree were significantly decreased, the mdr-1/P-gp expression was decreased, and γH2AX expression was upregulated compared with those of the 37 degree control group (all P<0.01). The absorbance values (P<0.01) and mdr-1/P-gp expression(P<0.05) were significantly lower and γH2AX expression(P<0.05) was significantly higher in the hyperthermia combined with 10 μg/mL Nef group than those of 10 μg/mL Nef group and hyperthermia group in MCF-7/Adr cells. The water submerged heating group had the lowest P-gp expression and the highest γH2AX expression among different heating groups at 42 degree and 45 degree in MCF-7/Adr cells (P<0.05). CONCLUSION Hyperthermia can increase the cell toxicity of adriamycin to multidrug resistant breast cancer cells. Hyperthermia significantly damages DNA of MCF-7/Adr cells and the higher temperature, the worse effect. Multidrug resistant breast cancer cells may respond differently to the different heating methods. Combined treatment of hyperthermia with Nef can increase the sensitivity in adriamycin chemotherapy.
ATP Binding Cassette Transporter, Subfamily B ; ATP Binding Cassette Transporter, Subfamily B, Member 1 ; genetics ; metabolism ; Antibiotics, Antineoplastic ; pharmacology ; Benzylisoquinolines ; pharmacology ; Breast Neoplasms ; genetics ; metabolism ; pathology ; Cell Line, Tumor ; Doxorubicin ; pharmacology ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Drugs, Chinese Herbal ; pharmacology ; Histones ; genetics ; metabolism ; Hot Temperature ; Humans ; RNA, Messenger ; genetics ; metabolism

OBJECTIVE: To investigate the suppression of MDR1 and P-glycoprotein induced by small interfering RNA and the restoration of sensitivity to chemotherapeutic drugs in multidrug-resistant hepatocellular carcinoma cell line Bel7402/5-Fu. METHODS: MDR1j targeted small interfering RNA duplexes were introduced into multidrug-resistant hepatocellular carcinoma cell line Bel7402/5-Fu. The suppression of MDR1 and its gene product P-glycoprotein was examined by RT-PCR and Western blot. MTT assay was performed to measure the reverse effect of small interfering RNA based on the results of IC50. Cell apoptosis was assessed by flow cytometry after various cell lines were treated with chemotherapeutic drugs. RESULTS: The overexpression of MDR1 and P-glycoprotein was suppressed efficiently by the introduction of small interfering RNA, which caused sequence-specific gene silence. The level of MDR1 in the transfected Bel7402/5-Fu cells reduced to 22.55% and P-glycoprotein to 25.49% compared with those of the controls. The apoptosis rate of Bel7402/5-Fu cells increased significantly in the siRNA group during the chemotherapy (P<0.01). Their resistance to 5-Fu was reversed by 14.88 folds, which indicated the restoration of sensitivity to drugs. CONCLUSION Small interfering RNA can inhibit MDR1 expression effective and reverse the multidrug resistance mediated by P-glycoprotein.
ATP Binding Cassette Transporter, Subfamily B ; ATP Binding Cassette Transporter, Subfamily B, Member 1 ; genetics ; metabolism ; Carcinoma, Hepatocellular ; genetics ; metabolism ; pathology ; Cell Line, Tumor ; Drug Resistance, Multiple ; genetics ; Drug Resistance, Neoplasm ; genetics ; Humans ; Liver Neoplasms ; genetics ; metabolism ; pathology ; RNA Interference ; RNA, Small Interfering ; genetics ; Transfection

OBJECTIVES: Fluorouracil chemotherapeutic drugs are the classic treatment drugs of gastric cancer. But the problem of drug resistance severely limits their clinical application. This study aims to investigate whether hypoxia microenvironment affects gastric cancer resistance to 5-fluorouracil (5-FU) and discuss the changes of gene and proteins directly related to drug resistance under hypoxia condition. METHODS: Gastric cancer cells were treated with 5-FU in hypoxia/normoxic environment, and were divided into a Normoxic+5-FU group and a Hypoxia+5-FU group. The apoptosis assay was conducted by flow cytometry Annexin V/PI double staining. The real-time reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting were used to detect the expression level of hypoxia inducible factor-1α (HIF-1α), multidrug resistance (MDR1) gene, P-glycoprotein (P-gp), and vascular endothelial growth factor (VEGF) which were related to 5-FU drug-resistance. We analyzed the effect of hypoxia on the treatment of gastric cancer with 5-FU. RESULTS: Compared with the Normoxic+5-FU group, the apoptosis of gastric cancer cells treated with 5-FU in the Hypoxia+5-FU group was significantly reduced (P<0.05), and the expression of apoptosis promoter protein caspase 8 was also decreased. Compared with the the Normoxic+5-FU group, HIF-1α mRNA expression in the Hypoxia+5-FU group was significantly increased (P<0.05), and the mRNA and protein expression levels of MDR1, P-gp and VEGF were also significantly increased (all P<0.05). The increased expression of MDR1, P-gp and VEGF had the same trend with the expression of HIF-1α. CONCLUSIONS Hypoxia is a direct influencing factor in gastric cancer resistance to 5-FU chemotherapy. Improvement of the local hypoxia microenvironment of gastric cancer may be a new idea for overcoming the resistance to 5-FU in gastric cancer.
Humans ; Fluorouracil/therapeutic use* ; Vascular Endothelial Growth Factor A/metabolism* ; Stomach Neoplasms/drug therapy* ; Drug Resistance, Multiple ; Vascular Endothelial Growth Factors/metabolism* ; Hypoxia ; ATP Binding Cassette Transporter, Subfamily B/genetics* ; ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics* ; Cell Line, Tumor ; Cell Hypoxia ; RNA, Messenger/metabolism* ; Hypoxia-Inducible Factor 1, alpha Subunit/genetics* ; Tumor Microenvironment

OBJECTIVE: To determine the effect of neferine (Nef) combined with mdr-1shRNA on the expression of mdr/P-gp in K562/A02 cell line. METHODS: MTT assay was used to observe the cell proliferation. The expression level of P-gp was determined by Western blot and the transcription of mdr-1 gene was detected by semi-quantitative RT-PCR. RESULTS: After K562/A02 cells were treated by Nef or mdr-1shRNA alone or both for 24 h, the proliferation of K562/A02 cells was significantly higher in the Nef combined with mdr-1shRNA treatment group than that of Nef or mdr-1shRNA alone group (P<0.01).The expression of mdr-1/P-gp in the Nef with mdr-1 shRNA group was significantly lower than that of Nef or mdr-1shRNA alone group. CONCLUSION Nef enhances the inhibition of mdr-1shRNA expression vector on K562/A02 cell proliferation and on P-gp protein to effectively reverse multidrug resistance induced by mdr-1 gene encoding P-gp.
ATP Binding Cassette Transporter, Subfamily B ; ATP Binding Cassette Transporter, Subfamily B, Member 1 ; genetics ; metabolism ; Benzylisoquinolines ; pharmacology ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Drug Synergism ; Drugs, Chinese Herbal ; pharmacology ; Humans ; K562 Cells ; RNA, Small Interfering ; genetics ; pharmacology

OBJECTIVE: To explore the multidrug resistance (MDR) mechanism of ABC superfamily transporters in the tumor stem cells(TSC) from human brain glioma tissues. METHODS: Samples of glioma were obtained from 30 patients undergoing microsurgical tumor resection. The CD133(+) cells and CD133(-) cells from these tumor specimens were isolated by magnetic activated cell sorting(MACS). These cells were cultured, proliferated and passaged. The protein and activity expression of multidrug-resistance protein 1(MDR1) and multidrug-resistance associated protein 1(MRP1) were analyzed between CD133(+) and CD133(-) cells by immunocytochemistry and RT-PCR respectively. RESULTS: CD133(+) cells generated free floating neurosphere like brain tumor spheres(BTS) and abnormal proliferating capacity in the serum-free medium(SFM) in vitro. Three cases from glioblastoma stem cells could form BTS in the complete medium, and could be cultured for 1-3 passages. The range of positive cell proportion for MDR1 and MRP1 expression in CD133(+) cells was 18%-67% and 23%-73% respectively. The expression levels of MDR1 and MRP1 mRNA were higher in CD133(+) glioma stem cells than those in the differentiated tumor cells(TC), the protein activity was increased to 16.1 and 19.6 times respectively compared with that of TC. The protein and activity expression were positively related to the pathological grades of tumors. MDR1 or MRP1 drug resistance was not expressed in all the tumors and there was obvious correlation between MDR1 and MRP1. CONCLUSION Only a small proportion of cells in the heterogeneous glioma is CD133(+) brain tumor stem cells which display the strong capacity of self-renewing, abnormal proliferation and intrinsic multidrug resistance to traditional chemotherapy. The high expression of MDR1 and MRP1 by the CD133(+) brain tumor stem cells is one of the main mechanisms in the chemoresistance of tumors. CD133(+) brain tumor stem cells can be served as the root of multidrug resistance and key therapeutic target for glioma chemotherapy.
AC133 Antigen ; ATP Binding Cassette Transporter, Subfamily B ; ATP Binding Cassette Transporter, Subfamily B, Member 1 ; genetics ; metabolism ; Antigens, CD ; immunology ; metabolism ; Brain Neoplasms ; metabolism ; Drug Resistance, Multiple ; Drug Resistance, Neoplasm ; Glioma ; metabolism ; Glycoproteins ; metabolism ; Humans ; Multidrug Resistance-Associated Proteins ; genetics ; metabolism ; Neoplastic Stem Cells ; drug effects ; metabolism ; Peptides ; metabolism ; Spheroids, Cellular ; drug effects ; Tumor Cells, Cultured