Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.6.3.44 (P-glycoprotein)
13,344 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Spontaneous and culture condition-dependent changes in P-glycoprotein expression and activity have been monitored in primary cultures of rat hepatocytes by using immunoblotting, PCR and fluorimetric techniques. In hepatocytes cultured in basal medium without addition of dexamethasone or 3-methylcholanthrene, mdr mRNA and P-glycoprotein increased progressively throughout a 72 h culture period, in concert with an enhancement in the ability to extrude the fluorescent dye Rhodamine-123. Addition of 1 microM dexamethasone to the culture medium slowed down the increase in mdr mRNA and P-glycoprotein, while inducing a significant increase in the efficiency of R-123 efflux. Addition of either 100 nM or 10 microM DEX produced different changes in mdr mRNA and protein, unrelated to the rate of Rhodamine-123 extrusion. When 50 microM 3-methylcholanthrene was added to the culture medium in the absence of any hormone supplementation, no significant changes in P-glycoprotein activity and expression took place, in comparison with control cultures. On the contrary, in the presence of dexamethasone (100 nM and 1 microM), 3-methylcholanthrene induced an increase in mdr mRNA and in the amount of immunoblottable protein during culture, without producing any concomitant increase in the efficiency to extrude Rhodamine-123. The last phenomenon resulted to be an artefact, since 3-methylcholanthrene was shown to inhibit Rhodamine-123 transport competitively. These results indicate that rat hepatocyte P-glycoprotein may be variously modulated in vitro, by supplementing culture medium with hormones and/or xenobiotics. Functional activity of the P-glycoprotein is not necessarily related with protein amount and/or mdr RNA.
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PMID:Differential modulation of P-glycoprotein expression by dexamethasone and 3-methylcholanthrene in rat hepatocyte primary cultures. 790 6

This study examined whether levels of estrogen receptor (ER), progesterone receptor (PR), and expression of estrogen regulated pS2 and/or heat shock protein (hsp) 27 were associated with drug resistance in a series of MCF-7 sublines expressing modest (i.e. 3- to 14-fold), yet clinically relevant, levels of resistance to vincristine (VCR). These sublines were variously derived following pulsed exposures to VCR, to fractionated X-irradiation, or to alternating drug and X-ray treatments. This selection procedure more closely reflects the clinical treatment of breast tumors than the use of continuous drug exposures. The drug-selected sublines exhibited the classical multidrug resistance phenotype (MDR) characterized by cross-resistance to vinblastine (VLB), etoposide (VP-16), and Adriamycin (ADR), overexpression of P-glycoprotein (Pgp), impaired accumulation of [3H]-VCR and of Rhodamine-123 (Rh 123), and altered activities of certain drug detoxification enzymes. This classic MDR phenotype was associated with a lack of mitogenic response to estrogen or antiestrogen, related to loss of detectable ER and PR; consistent with these data, neither pS2 nor hsp27 expression was detectable. In contrast, X-ray-pretreated VCR-resistant cells (MCF/DXR-10) cells exhibited a distinctive resistance phenotype proving cross-resistant to VLB and VP-16 but not to ADR, and Pgp overexpression was not detectable. Furthermore, these VCR-resistant DXR-10 cells retained parental levels of ER and PR, exhibited sensitivity to estrogen and 4-hydroxytamoxifen, and expressed detectable levels of pS2 and hsp27. Comparable characteristics to these MCF-7/DXR-10 cells were also identified in a similarly-derived X-ray-pretreated VCR-resistant subline of the ZR-75-1 human breast tumor cell line. These data therefore indicate that functional ER are frequently, but not invariably, modified in tumor cells which express resistance to multiple drugs.
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PMID:Differential expression of steroid receptors, hsp27, and pS2 in a series of drug resistant human breast tumor cell lines derived following exposure to antitumor drugs or to fractionated X-irradiation. 840 Mar 21

The physiological role of the multidrug resistance P-glycoprotein (P-gp), which is expressed by normal human T lymphocytes, is still largely unknown. To investigate whether or not P-gp is involved in the transport of cytokines, peripheral blood lymphocytes were stimulated with phytohemagglutinin (PHA) in the absence or presence of P-gp inhibitors, and concentrations of cytokines (interleukin-2 [IL-2], IL-4, IL-6, interferon-gamma [IFN-gamma]) in the supernatants of these cultures were quantitated by enzyme-linked immunosorbent assay. P-gp inhibitors included verapamil (Ver), tamoxifen (Tmx), and the P-gp specific monoclonal antibody UIC2. Release of IL-2 was significantly suppressed by these inhibitors at concentrations that were also effective in blocking efflux of Rhodamine-123 from normal T lymphocytes. IL-2 mRNA expression in lymphocytes was not different between PHA control and the cultures with P-gp inhibitors. Ver and Tmx did not interfere with T-cell activation as determined by CD25 and CD69 expression. In a nonhematological model, the P-gp expressing HCT-8 adenocarcinoma cell line, exogenously added IL-2 was shown to exert an inhibitory effect on P-gp mediated Rhodamine-123 efflux. In addition, transepithelial transport of IL-2 by electrophysiologically tight and polarized HCT-8 monolayers was examined. A time-dependent flux of IL-2 across dense monolayers, which was partially inhibited by Ver, was observed. We also investigated whether or not P-gp inhibitors suppressed release of other cytokines produced by activated T cells (IL-4, IL-6, IFN-gamma). Release of IL-4 and IFN-gamma was significantly inhibited by Ver, Tmx, and UIC2; however, release of IL-6 remained unaffected. These data show P-gp mediated transmembrane flux of IL-2 in T lymphocytes and HCT-8 cells. We conclude that P-gp participates in the transport of cytokines (IL-2, IL-4, and IFN-gamma) in normal peripheral T lymphocytes.
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PMID:Involvement of P-glycoprotein in the transmembrane transport of interleukin-2 (IL-2), IL-4, and interferon-gamma in normal human T lymphocytes. 878 31

Multidrug resistance (MDR) is often related to expression of P-glycoprotein (Pgp) or Multidrug Resistance Protein (MRP). Pgp-mediated MDR can be evaluated by determining cellular retention of fluorescent substrates by flow cytometry. This study determined if agents used to evaluate Pgp function also can be used to evaluate MRP function. Cellular retention of doxorubicin (Dox), Rhodamine-123 (Rh-123), and 3,3'-diethyloxacarbocyanine iodide (DiOC2(3)) were studied in MRP-expressing cell lines (HL60/Adr and HT1080/DR4), whereas a Pgp expressing cell line (A2780/Dx5) served as a positive control. Overexpression of Pgp correlated inversely with retention of Dox, Rh-123, and DiOC2(3); however, under identical experimental conditions (1 h reincubation in drug-free medium), no retention difference of the three agents was detected between parental and MRP-expressing resistant cells. Upon extending the reincubation time to 4 h, an efflux of Rh-123 and Dox in the resistant lines became apparent and even more pronounced after 24h; however, still no efflux was detectable for DiOC2(3). Incubation of the cells with a modulator of MDR, PAK-104P, negated the observed drug efflux in Pgp and MRP expressing cells, which correlated with increased sensitivity of the MDR lines to doxorubicin. Thus both Dox and Rh-123 can be used to evaluate MRP-function, but DiOC2(3) can not.
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PMID:DiOC2(3) is not a substrate for multidrug resistance protein (MRP)-mediated drug efflux. 887 50

Clinical chemotherapy of breast carcinomas must be considered insufficient, mainly due to the appearance of drug resistance. The multidrug resistance (MDR) phenotype, either intrinsically occurring or acquired, e.g., against a panel of different antineoplastic drugs, is discussed in relation to several MDR-associated genes such as the MDR-gene mdr1 encoding the P-glycoprotein (PGP), the MRP gene (multidrug resistance protein) encoding an MDR-related protein or the LRP gene encoding the lung resistance protein. Numerous experimental and clinical approaches aiming at reversing resistance require well-characterised in vitro and in vivo models. The aim of our work was to develop multidrug resistant sublines from human xenotransplanted breast carcinomas, in addition to the broadly used line MCF-7 and its multidrug resistant subline MCF-7/AdrR. MDR was induced in vitro with increasing concentrations of Adriablastin (ADR) for several weeks, resulting in a 3.5- to 35-fold increase in IC50 values using the MTT-test. Cell lines were cross-resistant toward another MDR-related drug, vincristine, but remained sensitive to non-MDR-related compounds such as cisplatin and methotrexate. The resistance toward Adriamycin and vincristine was confirmed in vivo by a lack of tumour growth inhibition in the nude mouse system. Gene expression data for the mdr1/PGP, MRP/MRP and LRP/LRP on both the mRNA (RT-PCR) and the protein levels (immunoflow cytometry) demonstrated that induction of mdr1 gene expression was responsible for the acquired MDR phenotype. Rhodamine efflux data, indicated by PGP overexpression, underlined the development of this MDR mechanism in the newly established breast carcinoma lines MT-1/ADR, MT-3/ADR and MaTu/ADR.
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PMID:Development and characterisation of novel human multidrug resistant mammary carcinoma lines in vitro and in vivo. 931 9

Mechanism of multixenobiotic resistance (MXR), identical to multidrug resistance (MDR) in tumor cells, has been found in aquatic invertebrates. The presence of this ATP-dependent membrane P-glycoprotein (Pgp) pump was confirmed by biochemical ('binding'), molecular (immunohistochemical, Western, Northern), physiological (verapamil-sensitivity) and toxicological (modulation of toxicity) methods. The inducibility of MXR in the presence of xenobiotics and its wide taxonomic distribution suggests its role as a general biological defense mechanism that rescues organisms by pumping potentially toxic xenobiotics out of the cells. Some xenobiotics, the chemosensitizers, can inhibit this defense mechanism. The presence of these MXR-inhibitors has important implications on environmental parameters like exposure, uptake, internal dose, bioaccumulation, response, synergism and toxicity. Such MXR-inhibitors, for example, enhance the accumulation of carcinogenic aromatic amines in mussel, with subsequent enhancement in production of their mutagenic metabolites, in induction of single strand breaks in DNA, and in induction of DNA-adducts. The property to inhibit defense mechanism of organisms classifies MXR-inhibitors among top-hazardous environmental chemicals. Therefore, we measured the concentration of chemosensitizers in water concentrates or sediment extracts as their potential to modulate the accumulation of fluorescent dyes in a cell-culture of NIH 3T3 mouse fibroblasts stable transfected with human MDR1 gene, or as the potential of native waters to decrease the efflux-rate of Rhodamine B from gills of mussels. We found significantly higher concentrations of MXR-inhibitors in samples from polluted marine sites or from polluted rivers than in samples from corresponding unpolluted sites. These concentrations were able to enhance the accumulation of fluorescent dyes or carcinogenic aromatic amines in clams, mussels, snails and sponges exposed to these xenobiotics, demonstrating the ecotoxicological relevance of MXR-inhibitors present in polluted waters.
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PMID:The chemosensitizers of multixenobiotic resistance mechanism in aquatic invertebrates: a new class of pollutants. 963 88

Curcumin is a natural phenolic compound found in the rhizomes of Curcuma longa and endowed with beneficial biological activities including antioxidant, anticarcinogenic and hepatoprotective effects. In this study curcumin was tested for its potential ability to interact in vitro with hepatic P-glycoprotein (Pgp), in a model system represented by primary cultures of rat hepatocytes, in which spontaneous overexpression of multidrug resistance (mdr) genes occurs. In both freshly-plated hepatocytes, containing low levels of Pgp, and 72 hour-cultured hepatocytes, containing high levels of Pgp, the Rhodamine-123 (R-123) efflux, which represents a specific functional test for Pgp-mediated transport, was inhibited by curcumin in a dose-dependent manner. Western blot analysis showed that 25microM curcumin, when included in the culture medium throughout the experimental observation (72 hours), was able to significantly lower the increase of mAb C219-immunoreactive protein spontaneously occurring in the cells during culture. Curcumin, at doses ranging from 50 to 150microM was cytotoxic for freshly-plated hepatocytes, as shown by the strong decrease in the cell ability to exclude trypan blue 24 hours later, but it was significantly less cytotoxic when added to 24 or 48 hour-cultured cells. The resistance to curcumin, progressively acquired by cells during culture, was significantly reduced by high concentrations of dexamethasone (DEX) or dimethyl-sulfoxide (DMSO), culture conditions known to inhibit the spontaneous overexpression of Pgp. In addition, in a concentration-dependent manner, verapamil reverted curcumin resistance in Pgp overexpressing hepatocytes. In photoaffinity labeling studies, curcumin competed with azidopine for binding to Pgp, suggesting a direct interaction with glycoprotein. These results suggest that curcumin is able to modulate in vitro both expression and function of hepatic Pgp and support the hypothesis that curcumin, a chemopreventive phytochemical, could reveal itself also as a compound endowed with chemosensitizing properties on mdr phenotype.
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PMID:Effects of curcumin on P-glycoprotein in primary cultures of rat hepatocytes. 965 Nov 24

We have cloned from a chicken intestinal cDNA library Cmdr1, the first avian P-glycoprotein. Cmdr1 is 67% and 69% identical to proteins encoded by the human MDR1 and MDR2 genes, respectively. Functional expression of Cmdr1 in both mouse NIH 3T3 and yeast cells demonstrated that Cmdr1 represents the avian ortholog of human Mdr1, since it confers resistance to several anticancer drugs and the fluorescent dye rhodamine 6G. Northern and immunoblot analysis showed that CMDR1 is highly expressed throughout the intestine and in the liver, and to a considerable extent in kidney, brain, lung, heart, eye and follicles. In situ hybridization revealed a cell type-specific expression of CMDR1 in the intestinal epithelium, with high levels in the villi of the small and large intestine as well as crypt cells. These data suggest that Cmdr1 could play a role in intestinal detoxification. Most interestingly, immunoblotting showed that Cmdr1 is also expressed in ovarian tissues, particularly in theca cells, the major site for ovarian estrogen production in birds. Indeed, competition experiments indicated that Cmdr1 interacts with estradiol, since rhodamine 6G efflux was efficiently blocked by estradiol in NIH 3T3 cells expressing Cmdr1. Rhodamine efflux was also blocked by PSC-833, a specific inhibitor of steroid-transporting P-glycoproteins from mammalian cells. We propose that Cmdr1 in ovarian cells could be involved in the cell type-specific transport or release of estrogen that is essential for avian follicular development.
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PMID:Cmdr1, a chicken P-glycoprotein, confers multidrug resistance and interacts with estradiol. 1019 30

Much remains to be learned about drug resistance in the biology of RCC and its metastases. We measured MDR-1/P-glycoprotein expression in 19 tumor samples from patients with metastatic RCC by RNase protection and quantitative PCR assays. The median level of the 16 tumor metastases was 4.9 (range: 0.10 to 156.2) relative to the level of 10 assigned to a reference cell line, SW620, which has been characterized as expressing a minimum level of MDR-1. Since these levels were lower than expected for RCC, we asked whether the metastases possessed a phenotype different from primary RCC and examined MDR-1 expression in 5 paired cell lines derived from primary and metastatic RCC. In 8/10 lines, MDR-1 expression was >10. Relative to the level in the primary line, MDR-1 expression was decreased (3 to 50-fold) in 3 metastatic lines, was increased in 1, and unchanged in 1. MRP mRNA expression was lower in the metastatic lines while EGFR expression was variable. IC50 values for 6 compounds (including 4 standard agents and one new Phase 1 agent) were determined for the paired lines. Rhodamine and calcein efflux assays were performed as measures of P-glycoprotein and MRP function. Rhodamine efflux correlated with MDR-1 mRNA expression (r = 0.87) and with the IC50s (r = 0.60) for paclitaxel in the paired cell lines. In contrast, calcein efflux did not correlate with MRP expression. Lastly, MDR-1 expression correlated with cytokeratin 8 (CK8) protein levels, a measure of cellular differentiation. In sum, these data suggest renal cell carcinoma (RCC) metastases have altered MDR-1 expression potentially due to altered differentiation relative to the primary tumor. Thus, the drug resistance phenotype of primary RCC tumors may not reflect that of their metastases.
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PMID:Intrinsic drug resistance in primary and metastatic renal cell carcinoma. 1037 90

The clinical benefit of Cyclosporine A (CsA) in IBD is controversial. Drugs including CsA are substrates of the P-glycoprotein-170 (Pgp-170) cell surface pump. Although the mechanism of action of CsA is complex, we determined that Pgp-170 might be expressed differentially between these two diseases. Intra-epithelial, lamina propria and peripheral blood lymphocytes were stained with the antibody MRK-16, which recognizes the surface antigen Pgp-170. Functional activity was assayed using a Rhodamine dye efflux assay (Rh123). RT-PCR was used to detect Pgp-170 mRNA. Overall, less P-gp-170 surface expression was found on UC CD3+ intestinal lymphocytes compared to controls and Crohn's disease. A decrease in Pgp-170 activity was also measured using the Rh123 functional assay. Fewer CD8+ intraepithelial lymphocytes (IEL) (which intrinsically have more Pgp-170 function) were also found in UC. Furthermore, UC IEL P-gp expression was under the limit of detection by RT-PCR. Overall, greater and differential expression, function and mRNA for Pgp-170 was found in Crohn's disease compared to the UC and normal tissues analyzed.
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PMID:Differences in P-glycoprotein-170 expression and activity between Crohn's disease and ulcerative colitis. 1043 13


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