ABSTRACT
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.
Subject(s)
Humans , ATP Binding Cassette Transporter, Subfamily B , Genetics , Arthritis, Rheumatoid , Drug Therapy , Genetics , Cells, Cultured , Drug Resistance , Fibroblasts , Methotrexate , Pharmacology , Synovial Membrane , Cell BiologyABSTRACT
[Abstract ] Objective High-mobility group box-1 (HMGB1) is abundantly released in the epileptogenic brain tissue , but few reports are seen about the effect of HMGB 1 on the expression of P-glycoprotein ( P-gp) in the vascular endothelial cells of the epi-leptogenic tissue .This study is to explore whether HMGB 1 can regulate P-gp expression in the brain microvascular endothelial cells of the mouse in vitro . Methods Immortalized brain microvascular endothelial bEnd .3 cells of the mouse were cultured in vitro and al-located to different concentration groups ( treated with culture medium containing 10 , 100 , 500 , and 1000 ng/mL HMGB1 for 8 hours), treatment duration groups (treated with culture medium containing 100 ng/mL HMGB1 for 4, 8, 16, 24, and 32 hours), and a control group ( treated with culture medium without HMGB 1 ) .The mRNA expression of P-gp-encoding gene-multidrug resistance gene 1a (mdr1a) was detected by real-time qPCR, and its protein expression determined by Western blot and immunocytochemistry . Results The results of qPCR manifested that the expressions of mdr 1a mRNA were 1.646 ±0.176, 1.777 ±0.135, 1.617 ±0.043, and 1.398 ±0.182 in the 10, 100, 500, and 1000 ng/mL HMGB1 groups, respectively, significantly higher than 1.030 ±0.284 in the control group (P<0.05), and so were those in the 4, 8, 16, 24 h, and 32 h groups (2.655 ±0.112, 2.168 ±0.212, 1.823 ± 0.232, 1.418 ±0.376, and 1.445 ±0.123) than in the control (1.010 ±0.164) (P <0.05).Western blot showed a significant increase in the P-gp protein expression in all the concentration groups (P<0.05) as well as in the 8 h and 16 h treatment duration groups as compared with the control group (P<0.05).Immunocytochemis-try also revealed a higher P-gp expression in the HMGB1-treated than in the control cells (P<0.01). Conclusion HMGB1 can upregu-late the expressions of mdr1a mRNA and P-gp protein in the brain microvascular endothelial cells of the mouse , which may associated with drug resistance of central nervous system diseases , especially that of epilepsy .
ABSTRACT
Objectives To investigate the inlfuence of polymorphisms of SLC19A1 80G>A, MDR1 exon26C>T and MDR1 exon21G>T/A on curative effect and adverse reaction of high-dose methotrexate in patients with acute lymphoblastic leukemia. Methods MALDI-TOF-MS technique was used to detect the polymorphisms of SLC19A1 80G>A, MDR1 exon 26C>T and MDR1 exon21G>T/A in 108 patients with acute lymphoblastic leukemia (ALL). The relationship of genetic polymorphism, survival rate and toxicity was analyzed. Results The 36-month event-free survival was not related to any polymorphisms of MDR1 and SLC19A1. Patients with mutant types of MDR1 exon26C>T and MDR1 exon21G>T/A showed a much higher MTX plasma levels at 24 hours and higher incidence of hepatic injury (PT, MDR1 exon21G>T/A has a large inlfuence on hepatic toxicity and plasma concentra-tions of MTX.
ABSTRACT
Objective: To investigate the enhancive effect of TNF-α on transfection efficiency of adenovi-rus in mononuclear cells of mouse and the enhancive capability of protection of bone marrow by MDR1. Meth-ods: Before the mononuclear cells of mouse were infected by recombinant adenovirus encoding human MDR1 gene, they had been pretreated by TNF-α. Transfection efficiency of adenovirus was monitored by fluo-rescence microscopy, immunohistochemistry and flow cytometry (FCM). mRNA and protein levels of MDR1 in the mononuclear cells of mouse were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot before and after treatment of TNF-α. Results: After treatment of TNF-α, tansfection rates of adenovirus were obviously increased in the treated group (26.26%) compared with the untreated group (11.96%). mRNA levels and protein levels of MDR1 were obviously increased in the treated group compared with the untreated group. Conclusion: TNF-α could enhance transfection efficiency of adenovirus in mononu-clear cells of mouse and enhance capability of protection of bone marrow by MDR1.
ABSTRACT
Objective To study the influential factors of tacrolimus'dosage and concentration differences between individuals in morning periods after renal transplantation.Methods The clinical data consisted of 118 receptors in morning periods after renal transplantation,whose immune suppressions were tacrolimus,mycophenolate and hormone.At 3,7,14 and 30 d after operation,all the receptors'weight,dosage of tacrolimus,dosage of hormone,diarrhea,blood fat,liver function,renal function,albumn and erythrocrit were recorded respectively,and at the same time their concentrations of tacrolimus and genetic polymorphisms of CYP3A5,MDRl 3435,MDR1 2677 and MDRl 1236 weredetected.Multiple linear regressions were performed.Results The fitting degrees of stepwise regression equations were low.At 3,7,14 and 30 d after operation,the adjusted R2was 0.284,0.267,0.417 and 0.324,respectively.From the aspect of pharmacogenomics,the main factors rela-ted to the differences of tacrolimus'dosage and concentration included MDR1 2677,MDRl 1236 and MDR13435,which varied intensively.Age,albumn,renal function,blood fat and liver function were important factors too.Conclusions The main reasons of the differences of tacrolimus'dosage and concentration between individuals in morning periods after renal transplantation are medicines and changes of internal environment after operation.The genetic polymorphisms of MDR1,age,albumn,renal runetion.blood fat and liver function are important factors too.
ABSTRACT
The prognostic significance of multidrug resistance (MDR) gene expression is controversial. We investigated whether multidrug resistance gene 1 (MDR1), multidrug resistance-related protein (MRP) and lung resistance protein (LRP) mRNA expression are associated with outcomes in acute leukemia patients. At diagnosis we examined MDR1, MRP and LRP mRNA expression in bone marrow samples from 71 acute leukemia patients (39 myeloid, 32 lymphoblastic) using nested RT-PCR. The expression of each of these genes was then expressed as a ratio in relation to beta-actin gene expression, and the three genes were categorized as being either 0, 1+, 2+ or 3+. MDR1, MRP and LRP mRNA expression was detected in 23.9%, 83.1% and 45.1 %, respectively. LRP mRNA expression was significantly associated with resistance to induction chemotherapy in acute leukemia patients, and in the AML proportion (p=0.02 and p=0.03, respectively). MRP and high MDR1 mRNA expression was associated with poorer 2-yr survival (p=0.049 and p=0.04, respectively). Patients expressing both MRP and LRP mRNA had poorer outcomes and had worse 2-yr survival. The present data suggest that MDR expression affects complete remission and survival rates in acute leukemia patients. Thus, determination of MDR gene expression at diagnosis appears likely to provide useful prognostic information for acute leukemia patients.
Subject(s)
Middle Aged , Male , Infant , Humans , Female , Child, Preschool , Child , Aged , Adult , Adolescent , Vault Ribonucleoprotein Particles/genetics , Survival Rate , RNA, Neoplasm/genetics , RNA, Messenger/genetics , Prognosis , Neoplasm Proteins/genetics , Multidrug Resistance-Associated Proteins/genetics , Leukemia, Myeloid, Acute/drug therapy , Precursor Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Leukemia/drug therapy , Genes, MDR , Gene Expression , Base SequenceABSTRACT
Aim To explore the relationship of the single nucleotide polymorphisms of multidrug resistance gene 1 and brain derived neurotrophic factor(BDNF) gene to childhood drug resistance epilepsy.Methods Two single nucleotide polymorphisms,T-129C polymorphism in multidrug resistance gene 1 and C270T polymorphism in BDNF gene,were conducted with PCR-restriction fragment length polymorphism analysis.The distribution of genotypes and allele frequencies of two single nucleotide polymorphisms in childhood drug resistance epilepsy were compared to those in drug respond epilepsy and controls.Results The distribution of TT genotype and T allele frequencies of multidrug resistance gene 1 in drug resistance epilepsy differed significantly from those in drug respond epilepsy and controls(P0.05).Conclusions The findings suggested that the T-129C polymorphism of multidrug resistance gene 1 maybe associated with childhood drug resistance epilepsy and played some role in the etiology of drug resistance epilepsy,but C270T polymorphism of BDNF gene was not confirmed to relate to childhood drug resistance epilepsy.
ABSTRACT
OBJECTIVE:To discuss the clinical significance of the expression of P- glycoprotein(P-gp),the multidrug resistance gene - 1(MDR - 1) expression product in the malignant lymphoma of nasalcavity and pharynx nasalis.METHODS: The expression of P - gp in the malignancy lymphoma of nasalcavity and pharynx nasalis of 37 cases was detected by S - P Immunohistochemical staining,and its correlations with patents' first visit,return visit,original sites,pathological type, immunohistochemical typing and prognosis were analyzed.RESULTS:Positive P- gp expression appeared in 87.5%(7/8) of the patients for return visit vs.34.5%(10/29) for first visit and 70.0%(7/10) with malignant lymphoma at pharynx nasalis vs.37.0%(10/27) with malignant lymphoma at nasal cavity,all showing significant differences.There were also significant differences in drug resistance between the patients with positive P- gp expression and those with negative P- gp expression (88.2%(15/17) vs.10.0%(2/20)).CONCLUSION:Positive P-gp expression serves as scientific basis for clinicians in predicating drug resistance and selecting drugs and establishing schemes.
ABSTRACT
Objective:To construct the recombinant adenovirus expressing human mdr1 gene.Methods:The cDNA of mdr1 gene was subcloned into the shuttle plasmid pAdTrack-CMV and then clone the homologous recombinant adenovirus genomic plasmid pAdEasy in bacteria.We tranfected the DNA of identified recombinant plasmid into 293 cells via liposome,and then did the package and amplification of adenovirus.The mononuclear cells of mouse were infected with Ad5-mdr1 and the target gene was determined by fluorescence microscope and flow cytometry.Results:Restriction endonuclease and PCR analysis confirmed that the recombinant adenovirus vector of mdr1 gene was successfully constructed.The titer of the recombinant adenovirus was 8.3?1011pfu/ml,and the infection efficiency to the mononuclear cell achieve 10%~15%.The expression of mdr1 gene was observed at 48h after the transfection of the mononuclearcell with Ad5-mdr1.Conclusion:The recombinant adenovirus expressing mdr1 gene was successfully constructed,which paves the way for the following studies related to mdr1.