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)

The apical surface of the proximal tubular epithelium is the site of both P-glycoprotein localization and postulated active secretion of organic cations in the mammalian kidney. P-glycoprotein has been shown to act as a pleiotropic drug efflux pump across the cell membrane of tumor cells expressing the multidrug resistance phenotype, whereas the renal organic anion and organic cation secretory systems serve the function of pleiotropic drug transport across the proximal tubule epithelium. Because most known substrates for P-glycoprotein are organic cations, we tested the hypothesis that the physiological function of this protein in the kidney is to mediate renal organic cation secretion. In one approach, we compared the postnatal development of organic cation transport with that of kidney mdr gene expression. Cimetidine-sensitive uptake of classical substrates for renal secretion (N-methyl nicotinamide and tetraethylammonium) into kidney slices developed gradually in neonate mice, reaching adult capacity in 4 to 6 weeks. P-glycoprotein and its mRNA, as estimated by immunohistochemical methods and RNAse protection analysis, were undetectable at birth and were expressed abruptly at the adult level between 2 and 3 weeks of age. In another approach, classical inhibitors of renal organic cation secretion (cimetidine and cyanine 863) failed to reverse resistance to adriamycin in Chinese hamster ovary and P388 cell lines, which possess the phenotypic traits of multidrug resistance. These results suggest that the cimetidine-sensitive component of organic cation secretion is mediated by a protein other than the P-glycoprotein in the mammalian kidney.
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PMID:Postnatal development of organic cation transport and mdr gene expression in mouse kidney. 135 Oct 97

The brush border membrane of the proximal tubule contains two efflux pathways for organic cations from the cell to the tubular fluid: a P-glycoprotein and an organic cation/H+ exchanger. There is evidence that they transport many of the same substrates. Their structural relatedness is unknown and is the subject of this report. The experimental approach was to identify the exchanger with photoaffinity labeling reagents. The rationale was that if the P-glycoprotein and the organic cation/H+ exchanger transport many of the same substrates, then they might be photoaffinity labeled by the same reagents. [125I]Iodoarylazidoprazosin and [3H]azidopine are two reagents, which have been used, to photoaffinity label the P-glycoprotein. We found that several polypeptides were photolabeled in a time- and concentration-dependent manner. The photoincorporation into only two of these polypeptides (41 and 28 kDa) was blocked extensively by the presence of known substrates for the exchanger. The photoaffinity labeling of only the 41-kDa polypeptide was affected by treatment with the chemical reagents, N-ethylmaleimide and dithiothreitol, which are known to affect the exchanger reaction. The findings are consistent with the interpretation that a 41-kDa polypeptide is, or is a component of, the exchanger.
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PMID:Photoaffinity labeling of the organic cation/H+ exchanger in renal brush border membrane vesicles. 135 95

We present a new transport model that may be useful for many kinds of transepithelial transport experiments. The model permits estimation of a pump Km and pump activity solely on the basis of transepithelial tracer fluxes. We apply the model to studies of a multidrug efflux pump, P-glycoprotein, which is normally located in the apical plasma membrane of certain transporting epithelia such as kidney proximal tubule cells. To determine the functional properties of this multidrug transporter in an epithelium, we studied the transepithelial transport of the chemotherapeutic drug, vinblastine, in epithelia formed by the kidney cell lines MDCK, LLC-PK1, and OK. We have previously shown that basal to apical flux of 100 nM vinblastine was about five times higher than apical to basal flux in MDCK epithelia, indicating that there is a net transepithelial transport of vinblastine across MDCK epithelia. Addition of unlabeled vinblastine reduced basal to apical flux of tracer and increased apical to basal flux of tracer in a concentration-dependent manner, a pattern expected if there is a saturable pump that extrudes vinblastine at the apical plasma membrane. The model permits estimation of a pump Km and pump activity solely on the basis of transepithelial tracer fluxes. According to the transport model the apical membrane pump has Michaelis-Menten kinetics with an apparent Km = 1.1 microM. Net basal to apical transport of vinblastine was also observed in LLC-PK1 cells and OK cells which are other kidney-derived cell lines. The order of potency of the transport is LLC-PK1 greater than MDCK greater than OK cells. The organic cation transporter is not involved in this vinblastine transport because vinblastine transport in MDCK cells was not affected by 3 mM tetramethyl- or tetraethylammonium. Inhibitors of vinblastine transport in MDCK cells was not affected by potency, were verapamil greater than vincristine greater than actinomycin D greater than daunomycin. The transport pattern we observed is that predicted to result from the function of the multidrug transporter in the apical plasma membrane.
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PMID:Transepithelial transport of vinblastine by kidney-derived cell lines. Application of a new kinetic model to estimate in situ Km of the pump. 220 28

We studied transepithelial transport of 3H-labeled hydrophobic cationic drugs in epithelia formed by wild-type and by drug-resistant Madin-Darby canine kidney (MDCk) cells that had been infected with a retrovirus carrying the multidrug-resistance (MDR1) cDNA which encodes the P-glycoprotein. P-glycoprotein is an ATP consuming plasma membrane multidrug transporter responsible for the efflux of cytotoxic chemotherapeutic drugs from resistant cancer cells. Wild-type MDCK cells have small amounts of P-glycoprotein detected by immunoprecipitation. Net transepithelial transport across wild-type MDCK epithelia was demonstrated. Basal to apical flux of 100 nM vinblastine was about six times higher than apical to basal flux. Addition of unlabeled vinblastine reduced basal to apical flux of tracer and increased apical to basal flux of tracer, a pattern expected if there is a saturable pump that extrudes vinblastine at the apical plasma membrane. Daunomycin, vincristine, and actinomycin D were also actively transported and at 20 microM these agents inhibited transport of vinblastine, suggesting that wild-type MDCK cells have a common transporter for all these drugs. Vinblastine transport was also inhibited by 20 microM verapamil, which inhibits the multidrug transporter and reverses multidrug-resistance in non-polarized cells. Net transepithelial transport of all these cytotoxic drugs and of verapamil was much higher in epithelia formed by MDCK cells infected with a human MDR1 virus (MDR-MDCK) which is expressed on the apical surface of MDR-MDCK monolayers. Because the transport of these cytotoxic drugs and verapamil is increased in MDR-MDCK epithelia compared to wild-type MDCK epithelia, transport in both these cell populations can be attributed to P-glycoprotein. These results are consistent with a role for P-glycoprotein in multidrug secretory transport across the epithelium of the proximal tubule since P-glycoprotein is normally expressed on the apical membrane of proximal tubule cells.
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PMID:Transepithelial transport of drugs by the multidrug transporter in cultured Madin-Darby canine kidney cell epithelia. 257 70

We examined the distribution of RNA levels expressed by the multidrug-resistance gene (MDR1, also known as PGY1) in 42 renal cell carcinoma (RCC) samples (38 primary and four metastatic lesions). The median MDR1 RNA level for the 38 primary lesions, expressed relative to the level for KB-3-1 cells, was approximately one-half of the level in multidrug-resistant KB-8-5 cells. Elevated MDR1 RNA levels were also observed in three of the four metastatic lesions. The mean MDR1 RNA level was higher in well-differentiated RCCs than in those that were poorly differentiated, suggesting that the increased expression of the MDR1 gene in RCCs originates from the increased expression in renal proximal tubule cells. To clarify the association of the MDR1 protein product P-glycoprotein with natural resistance to doxorubicin (ADR) in RCCs, we evaluated the effects of quinidine on in vitro sensitivity to ADR in 16 RCC samples, using a [3H]thymidine incorporation assay. The enhancing effect of quinidine (7.5 micrograms/mL) on sensitivity to ADR was statistically significant only in the group with high MDR1 RNA levels. Similar enhancement by quinidine of sensitivity to ADR was also observed in the established RCC cell lines in which MDR1 RNA levels were high. These results suggest that P-glycoprotein is active in the natural resistance of RCCs to ADR.
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PMID:MDR1 RNA levels in human renal cell carcinomas: correlation with grade and prediction of reversal of doxorubicin resistance by quinidine in tumor explants. 272 49

The renal proximal tubule is a major site of injury in a variety of congenital/metabolic diseases including nephropathic cystinosis, the most commonly known cause of renal Fanconi's syndrome. In this lysosomal storage disease there are defects in proximal tubule function within the first few months of life. While culture of renal tubular cells from the urine of these patients is possible, development of immortalized cell lines would insure large numbers of homogeneous cells for studies of renal epithelial cell morphology and pathophysiology in this disease. To develop immortalized cells, cystinotic and normal proximal tubular cells in culture were exposed to an immortalizing vector, containing pZiptsU19 with the temperature sensitive SV40 T-antigen allele tsA58U19 and a neomycin resistance gene, and neomycin-resistant tubular cells were selected for propagation. Ten clones from cystinotic patients have been developed and characterized. All clones express T-antigen at permissive temperature (33 degrees C). Immortalized cells have an epithelial morphology and grow to form confluent monolayers; doubling times vary from 31 to 86 hours. Cystinotic clones are keratin, MDR P-glycoprotein, and alpha-95 kD brush-border associated protein positive but Tamm-Horsfall protein negative by immunocytochemistry, as are normal proximal tubule cells immortalized with this vector. This is consistent with a proximal tubule origin of the cystinotic clones. The cystine content of the cystinotic cells is 70 to 160 times that of normal renal proximal tubular cells in culture, with most of the cystine sequestered in cell lysosomes, confirming that these cell lines express the storage defect.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Renal proximal tubular epithelium from patients with nephropathic cystinosis: immortalized cell lines as in vitro model systems. 756 23

Cyclosporin A, a cyclic undecapeptide, and FK506 are efficient immunosuppressive agents. They also attract attention as effective P-glycoprotein modulators that inhibit P-glycoprotein from binding to anticancer drugs and overcome multidrug resistance. Cyclosporin A itself interacts with a common binding site of P-glycoprotein to which Vinca alkaloids and verapamil bind. We were interested to determine whether cyclosporin A and FK506 are substrates for P-glycoprotein to transport, and we studied their transcellular transport. In LLC-PK1 cells, derived from porcine kidney proximal tubule and forming a highly polarized epithelium, cyclosporin A was transported in a saturable manner. LLC-GA5-COL300, a transformant cell line derived by transfecting LLC-PK1 with human MDR1 cDNA isolated from normal adrenal gland, expresses P-glycoprotein specifically on the apical surface and shows a typical multidrug-resistant phenotype. LLC-GA5-COL300 cells showed increased transport of cyclosporin A from the basal to the apical side. Kinetic analysis showed that this transport was a typical saturable transport with the calculated apparent Michaelis constant (Kappm) and the maximum flux (Vmax) as 8.4 microM and 2.4 nmol/mg protein/h, respectively. LLC-GA5-COL300 also showed increased transport of FK506 from the basal to the apical side. These results indicate that P-glycoprotein transports the immunosuppressive agents cyclosporin A and FK506.
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PMID:Human P-glycoprotein transports cyclosporin A and FK506. 768 Oct 59

Primary monolayer cultures of winter flounder renal proximal tubule epithelium mounted in Ussing chambers were used to characterize transepithelial transport of daunomycin (Dau). Control tissues performed active net secretion of Dau (0.064 +/- 0.027 nmol.cm-2.h-1). Mild heat shock (5 degrees C elevation for 6-8 h followed by return to normal temperature) almost doubled Dau secretion (0.114 +/- 0.026 nmol.cm-2.h-1). This response was inhibited approximately 40% by addition of the protein synthesis inhibitor, cycloheximide. Dau secretion was inhibited by verapamil, vinblastine, cyclosporin A, and to a lesser degree by the organic cation, tetraethylammonium. In addition, tetraethylammonium secretion was inhibited by vinblastine. Dau secretion was not inhibited by the organic anion, p-aminohippurate, and p-aminohippurate secretion was not inhibited by vinblastine. The transepithelial reabsorptive flux of Dau and the electrical characteristics of the tissues, including rheogenic glucose transport, were unaffected by any of the above treatments. Reaction of tissues with a monoclonal antibody to P-glycoprotein (C219) revealed the presence of this transporter on only apical microvilli. The data indicate that flounder possess an active mechanism for the renal excretion of Dau that is stimulated by mild heat shock. This mechanism is distinct from organic anion, but not organic cation, transport and has characteristics consistent with transport by an apical P-glycoprotein.
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PMID:Heat-shock-stimulated transepithelial daunomycin secretion by flounder renal proximal tubule primary cultures. 784 Feb 40

The transport of a fluorescent cyclosporin analogue was measured in killifish (Fundulus heteroclitus) proximal tubules by means of epifluorescence microscopy and digital image analysis. Renal cells rapidly accumulated the cyclosporin analogue from the medium and attained steady state within 60 min; luminal fluorescence increased over the first 60-90 min. At steady state, luminal fluorescence intensity was two to three times higher than cellular. Cellular fluorescence intensity was a linear function of medium substrate concentration and was not affected by any treatment used. In contrast, luminal fluorescence exhibited a saturable component as the medium concentration of the cyclosporin was increased. Secretion into the lumen was blocked by metabolic inhibitors, vanadate, other cyclosporins, such as cyclosporin A and cyclosporin G, and substrates for P-glycoprotein (verapamil, vinblastine, and quinine) but not by substrates for the renal organic anion or organic cation transport systems, such as p-aminohippurate or tetraethylammonium. The data are consistent with the fluorescent cyclosporin analogue entering proximal tubule cells by simple diffusion and then being pumped into the tubular lumen by P-glycoprotein.
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PMID:P-glycoprotein-mediated secretion of a fluorescent cyclosporin analogue by teleost renal proximal tubules. 784 Feb 47

The MDR1 gene product, P-glycoprotein, has been localized to the apical surface of the renal proximal tubule, but its functional role in the kidney is unknown. We studied renal luminal and antiluminal uptake of three known substrates of P-glycoprotein: vinblastine, vincristine and colchicine, by using the single pass multiple indicator dilution method under control conditions and in the presence of increasing concentrations of cyclosporin A, a potent inhibitor of P-glycoprotein. A bolus of [125I]albumin (plasma reference), L-[14C]glucose (extracellular and glomerular reference) and tracer 3H-substrate was injected into the left renal artery of anesthetized dogs and timed serial samples were collected from the left renal vein and left and right ureters. In a single pass, approximately 38, 13 and 8% of [3H]vinblastine, [3H] vincristine and [3H]colchicine, respectively, was extracted from the postglomerular circulation. Drug binding to plasma proteins was determined to be 81% for [3H]vinblastine, 71% for [3H] vincristine and 23% for [3H]colchicine. Despite the high degree of drug protein binding, the urine recoveries of [3H]vinblastine, [3H]vincristine and [3H]colchicine relative to L-[14C]glucose were 0.75 +/- 0.06, 0.69 +/- 0.06 and 0.94 +/- 0.02, confirming net secretion of each of these substrates. Infusion of cyclosporin A (0.1-5 microM) significantly decreased the urine recovery of [3H] vinblastine and [3H]vincristine relative to L-[14C]glucose in a dose-dependent manner. The renal excretion of [3H]colchicine was not affected by cyclosporin A at the concentrations tested (1-2 microM). The evidence suggests that net secretion of [3H]vinblastine and [3H]vincristine occurs across the luminal membrane of the renal cell.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Renal secretion of vinblastine, vincristine and colchicine in vivo. 790 30


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