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Objective@#To investigate the drug resistance of kaempferol reversed adriamycin (ADM)-resistant K562/ADM cells in chronic myelogenous leukemia (CML) and its related mechanism.@*Methods@#Methyl thiazolyl tetrazolium (MTT) method was used to detect the toxicity of ADM on K562 and K562/ADM cells for 24 h. The half inhibitory concentration (IC50) of ADM and the drug resistance multiple for 24 h were calculated. MTT method was used to detect the toxicity of kaempferol on K562/ADM cells for 24 h. The 5% inhibitory concentration (IC5) and 10% inhibitory concentration (IC10) of kaempferol for 24 h were calculated to determine the concentration of kaempferol in the subsequent experiments. And the cells untreated by the kaempferol were selected as the control group. The cell inhibition after the treatment of ADM for 24 h of the blank control group and kaempferol intervention group was detected by using MTT method. And then the cell inhibition for 24 h and ADM IC50 for 24 h in the above groups were calculated. The ratio of IC50 in the blank control group and kaempferol group was the reversal drug resistance multiple of kaempferol. The fluorescence intensity of ADM in K562/ADM cells treated by kaempferol was detected by using flow cytometry. Western blotting was used to detect the expressions of P-glycoprotein (P-gp), multidrug resistance-associated protein 1 (MRP1), phosphorylated p38 (p-p38), and total p38 (t-p38) protein in K562/ADM cells after the treatment of kaempferol, the specific inhibitor of p38-MAPK signaling pathway SB202190, and the combination of kaempferol and SB202190.@*Results@#After the treatment of ADM for 24 h, the IC50 value of K562 and K562/ADM cells was (0.9±0.6), (28.1 ±3.5) μg/ml, respectively. The drug resistance multiple of K562/ADM cells on the treatment of ADM for 24 h was 31.16 compared with the K562 cells. MTT method showed that kaempferol inhibited the proliferation of K562/ADM cells in a dose-dependent manner. According to the IC5 and IC10, 0.5 μmol/L and 1.0 μmol/L kaempferol were determined to do the subsequent experiments. After the combined interaction of kaempferol and ADM for 24 h, the ADM IC50 of K562/ADM cells in the blank control group, 0.5 μmol/L kaempferol group and 1.0 μmol/L kaempferol group was (33.7±5.7), (21.4±0.6), (15.9±1.8) μg/ml, respectively (F = 30.85, P < 0.05), and there was a statistical difference of pairwise comparison (both P < 0.05). The reversal drug resistance multiple of K562/ADM cells for 24 h in 0.5 μmol/L kaempferol group and 1.0 μmol/L kaempferol group was 1.58 and 2.12, respectively. Flow cytometry results showed that the mean fluorescence intensity (MFI) of ADM in the blank control group, 0.5 μmol/L kaempferol group and 1.0 μmol/L kaempferol group was 138.4±8.9, 154.3±2.2, 165.7±4.8, respectively, and the difference was statistically significant (F = 161.48, P < 0.05). Compared with the blank control group, after treatment of K562/ADM cells with 0.5 μmol/L and 1.0 μmol/L kaempferol for 24 h, the relative expressions of P-gp, MRP1 and p-p38 protein were decreased in K562/ADM cells (all P < 0.05), but there was no statistical difference in the expression of t-p38 protein (P > 0.05); SB202190 could reduce the relative expressions of P-gp, MRP1 and p-p38 protein (all P < 0.05); after the treatment of SB202190 combined with different concentration of kaempferol, the relative expressions of P-gp, MRP1 and p-p38 protein in K562/ADM cells did not decrease (P > 0.05).@*Conclusions@#Kaempferol can decrease the relative expressions of P-gp and MRP1 in K562/ADM cells by inhibiting p38-MAPK pathway, so as to increase the concentrations of ADM and to reverse the drug resistance of K562/ADM cells.
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Aim: To investigate the effect of Nrf2 pathway on the expression of MRP1 in mildly stable COPD mice. Methods: The mild COPD mouse model was established by passive cigarette smoking. The pathological changes of lung tissues were examined by HE staining. Immunohistochemistry and Western blot were used to detect the protein expression of MRP1, Nrf2 and HO-1. Results: Compared with normal group, each lung function index of the mild-moderate COPD model group was significantly lower, but compared with wide type(WT) model group, the reduction was more significant in Nrf2-/- model group. HE results showed diffuse inflammatory reaction and alveolar bronchial structure damage in alveolar of WT and Nrf2-/- model mice, and it was more pronounced in Nrf2-/- mice. Immunohistochemistry and Western blot results showed that the expression of MRP1 in lung tissue of Nrf2-/- normal mice was significantly reduced compared with the normal WT group. After passive cigarette smoking, The expression of MRP1, Nrf2 and HO-1 in WT model group increased significantly, but compared with Nrf2-/- normal mice, there was no significant change in the expression of MRP1 in Nrf2-/- model group. Conclusions: Mildly stable COPD mice may counteract the xenobiotic damage caused by cigarette smoke through up-regulating the expression of MRP1 protein, which may be associated with Nrf2 signaling activation.
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AIM:To investigate the levels and clinical significance of microRNA-133a/b (miR-133a/b) and multidrug resistance-associated protein 1 ( MRP1 ) in peripheral blood of the patients with drug-refractory epilepsy. METHODS:Prediction of the miRNAs targeting transcriptional regulation of MRP1 was conducted by bioinformatics analy-sis.The plasmids containing wild type and mutant 3’UTR of MRP1 reporter gene were constructed.Dual luciferase report-er gene assay was used to verify this prediction.In addition, the peripheral blood samples of the epilepsy patients (37 cases were drug-refractory, the other 58 cases were nonresistant) were collected.The levels of miR-133a/b and MRP1 were measured by real-time PCR and ELISA.RESULTS:Through TargetScan database, it was predicted that miR-133a/b tran-scriptionally regulated MRP1.The results of dual luciferase report gene assay suggested that luciferase activity in experi-mental group with miR-133a/b mimics, pMIR-MRP1 and pRLTK plasmids were down-regulated by 76.9% and 64.1%compared with that in control group with scramble mimic, pMIR-MRP1 and pRLTK plasmids.The luciferase activity was up-regulated by 3.62 times and 2.04 times in mutation group with miR-133a/b mimics, pMIR-mut-MRP1 and pRLTK plasmids compared with experimental group.Before administration, the serum levels of miR-133a/b in the epilepsy patients without drug resistance was 2.18 times and 1.74 times higher than than in the epilepsy patients with drug resistance ( P<0.05), respectively, while MRP1 expression level was 3.72 times higher in the epilepsy patients with drug resistance than those in the epilepsy patients without drug resistance.After administration, the levels of miR-133a/b in the epilepsy pa-tients without drug resistance were 2.76 times and 2.95 times higher than those in the epilepsy patients with drug resistance (P<0.05), respectively, while the serum level of MRP1 in the epilepsy patients with drug resistance was 4.99 times higher than that in the epileptic patients without drug resistance (P<0.01).CONCLUSION:miR-133a/b transcriptio-nally regulates MRP1.There are lower expression levels of miR-133a/b and higher expression level of MRP1 in the epilep-sy patients with drug resistance compared with those in the epilepsy patients without drug resistance.miR-133a/b and MRP1 may be a diagnostic indicator for determining refractory epilepsy.
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Objective Multidrug resistance-associated protein 1 (MRP1) has been reported with a close correlation with tumor multi-drug resistance. Real-time quantitative PCR (QRT-PCR) was performed to detect the MRP1 gene expression in childhood acute lymphoblastic leukemia (ALL) and its clinical signiifcance was analyzed. Methods Sixty-seven denovo ALL patients and 10 healthier children as bone marrow donor were studied. The chemotherapy was given according to CCLG-2008 protocol. SPSS software was employed to analyze the data and p-value below 0.05 was regarded as statistic signiifcance. Results MRP1 expression level showed a close correlation with ALL risk, the median of MRP1 expression was 4.28 (2.75~6.12), 5.62 (4.99~8.60) and 7.56 (3.66~11.13) for standard-risk group (SR), intermediate-risk group (IR) and high-risk group (HR), respectively. MRP1 mRNA expression in T-ALL group was 7.71 (6.49~14.35), which is higher than that of B-ALL (5.18(3.89~8.46)) (P<0.01). The rate of leukemia cells’ sensitivity to prednisone on 7th day was 70.6%in high expression group (n=34), which was signiifcantly lower than that in low expression group (n=33, 90.9%, P=0.035). The complete remission rateon 33th day was 64.7%in high expression group, and 87.9%in low expression group, which showed a signiifcant difference between them (P=0.026). Conclusions In children ALL, the expression of MRP1 is closely related with immunophenotyping, treatment response, hazard level and disease relapse.
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Objective To study the impact of establishment of lipopolysaccharide ( LPS )-induced rat model of chronic obstructive pulmonary disease (COPD)on the function of multidrug resistance-associated protein 1(MRP1)in the rat bronchial epithelium .Methods Using intratracheal instillation of LPS to establish COPD rat model .8-week old healthy male Wistar rats were divided into 3 groups (10 rats in each group ):(1) Normal control;(2) Modeling for 14 days after LPS instillation;(3) Modeling for 28 days after LPS instillation.Pulmonary function and the concentration of phenol red in bronchoalveolar lavage fluid ( BALF) and plasma were measured .The ratio of phenol red concentration in BALF/plasma was used as an index of the MRP 1 function in the rat bronchial epithelium and the expression of MRP 1 in the bronchial epi-thelium was also observed by immunohistochemistry .Results Compared with the normal group , the pulmonary functions of the rats in the model groups were significantly reduced along with the modeling progress .After intravenous administration of phenol red, the ratio of phenol red concentration in BALF/plasma was gradually reduced , and the expression of MRP1 in the bronchial epithelium was significantly decreased .Conclusions COPD rat model can be established by intratracheal LPS instillation, and the function of MRP1 in bronchial epithelium was gradually reduced along with the modeling progress .
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Multidrug resistance-associated protein 1 (MRP1/ABCC1) is the first identified member of ABCC subfamily which belongs to ATP-binding cassette (ABC) transporter superfamily.It is ubiquitously expressed in almost all human tissues and transports a wide spectrum of substrates including drugs,heavy metal anions,toxicants,and conjugates of glutathione,glucuronide and sulfate.With the advance of sequence technology,many MRP1/ABCC1 polymorphisms have been identified.Accumulating evidences show that some polymorphisms are significantly associated with drug resistance and disease susceptibility.In vitro reconstitution studies have also unveiled the mechanism for some polymorphisms.In this review,we present recent advances in understanding the role and mechanism of MRP1/ABCC1 polymorphisms in drug resistance,toxicity,disease susceptibility and severity,prognosis prediction,and methods to select and predict functional polymorphisms.
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Objective: To study the influences of Trastuzumab and IL-2 on human epidermal growth factor receptor-2 (HER2), multidrug resistance-associated protein1 (MRP1) of ACHN in vitro . Methods: ACHN cell line of RCC were cultured by cell culture technique. The tetrazolium-based colorimetric assay was used to evaluate the growth inhibitory effects of trastuzumab and IL-2. S-P method was used to determine the expression of HER2, MRP1 of the cells. Results: Trastuzumab showed the inhibitory effects on growth and multi-drug resistance in RCC from 40 ?g/L or 24 h in time-effective and dose-dependent manner. After treatment with Trastuzumab and/or IL-2, the expression of HER2, MRP1 genes of RCC was decreased significantly( P
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Objective To explore the role of multidrug resistance-associated protein 1 (Mrp1) on bilirubin metabolism in rat obstructive jaundiced (OJ) models. Methods Eighty Wistar male rats were randomly divided into three groups: sham operation group in which the common bile duct was educed but not ligated; common bile duct ligation (CBDL) group in which the common bile duct was ligated and cut off; bile reflow group in which on day 14 after common bile duct ligation operation, the internal drainage from common bile duct to duodenum by silica gel duct was performed. Serum prealbumin, serum total bilirubin and urine direct bilirubin were assayed routinely. The expressions of mrp1 mRNA and protein were detected in the liver tissues by semi-quantitative RT-PCR and indirect immunofluorescence respectively. Results Serum prealbumin was descending significantly (P