Pgp Protein Research Articles

Effect of Pluronic P85 on ATPase Activity of Drug Efflux Transporters

Pluronic block copolymers are potent sensitizers of multi-drug resistant (MDR) cancer cells. The sensitization effect by Pluronics is a result of two processes acting in concert: i) intracellular ATP depletion, and ii) inhibition of ATPase activity of drug efflux proteins. This work characterizes effects of Pluronic P85 on ATPase activities of Pgp, MRP1, and MRP2 drug efflux transport proteins and interaction of these proteins with their substrates, vinblastine, and leucotriene C4. Using membranes overexpressing Pgp, MRP1, and MRP2, the current study evaluates effects of Pluronic P85 (P85) on the kinetic parameters (Vmax, Km, Vmax/Km) of ATP hydrolysis by these ATPases. The decreases in the maximal reaction rates (Vmax) and increases in apparent Michaelis constants (Km) for these transporters in the presence of various concentrations of P85 were observed. The mechanism of these effects may involve i) conformational changes of the transporter due to membrane fluidization and/or ii) nonspecific steric hindrance of the drug-binding sites by P85 chains embedded into cellular membranes. The extent of these alterations was increased in the row MRP1 < MRP2 << Pgp. These data suggest that there are unifying pathways for the inhibition of Pgp and MRPs by the block copolymer. However, the effect of P85 on Pgp ATPase activity is considerably greater compared with the effects on MRP1 and MRP2 ATPases. This may be a reason for greater inhibitory effects of Pluronic in Pgp- compared with MRP-overexpressing cells.

P-Glycoprotein Overexpresion and Internal Tandem Duplications of FLT3 Are Characteristic of Discrete Populations of Elderly AML Patients.

Poor outcome in elderly patients with AML has been linked to an increased incidence of p-glycoprotein (pgp) expression, poor risk cytogenetics and secondary AML. Internal tandem duplication of the FLT3 gene is prognostic for poor outcome in younger patients with AML, but has not been analysed alongside other poor risk variables in the elderly. The ongoing AML14 trial for patients of 60 years or older is conducted by the UK National Cancer Research Institute Haematological Oncology Study Group, and is sponsored by the Leukaemia Research Fund. Patients could enter randomisations to intensive (daunorubicin- and ara-C-based) or non-intensive chemotherapy regimens. Using flow cytometry, we measured functional and phenotypic p-glycoprotein as well as MRP, LRP, bcl-2 and bax in CD45-gated blasts from 274 AML 14 patients. FLT3 ITDs were measured by PCR in the same cohort.We found that pgp protein and function were associated with a lower presenting white blood count (median WBC 27.0 × 109/l in non-functional cases, 6.0 × 109/l in functional cases, p<0.0001; p=0.0002 for protein). Conversely, FLT3 ITDs were associated with a high presenting white count (wildtype median WBC 18.2 × 109/l, ITD median 58.5 × 109/l). Moreover FLT3 ITDs were strongly associated with the absence of functional p-glycoprotein (P=0.0005): 18% patients had FLT3 ITDs, 45% patients had functional pgp, but only 4% patients had both. There was also a suggestion that pgp is associated with poor risk cytogenetics (P=0.02 for pgp protein expression, P=0.01 for functional pgp).Of the 274 patients analysed, 235 had intensive chemotherapy and 39 non-intensive treatment. Of the factors investigated, only pgp was associated with lower remission rates in univariate analysis adjusted for type of chemotherapy (functional pgp, p=0.01 for trend, protein p<0.0001), largely due to increases in the incidence of resistant disease (overall p=0.0008 for functional pgp, p<0.0001 for protein). In multivariate analysis, allowing for the effects of age, cytogenetics, WBC and performance status, only pgp protein proved significant in predicting CR or resistant disease.For overall survival, none of the variables proved significant in univariate or multivariate analyses adjusted for part of trial (intensive or non-intensive). However, this finding should be construed as lack of evidence of effect, rather than lack of effect: the confidence intervals for the hazard ratio for pgp protein range from 0.92 to 1.62, so the results are still consistent with a moderate adverse effect of pgp protein. Among the cohort, 154 patients were randomised to receive the pgp inhibitor PSC-833 (n=62) or not (n=92). PSC-833 has been previously reported as conferring worse overall survival, mainly arising from early deaths. There is evidence that higher levels of pgp protein mitigate the adverse effects of PSC 833 (p=0.005 for heterogeneity) in remission rates. In conclusion, pgp over-expression was associated with a low white cell count and with increased levels of resistant disease in elderly patients with AML. We suggest that pgp positive (slow-growing) and growth-factor receptor mutated (fast-growing) AML clones form distinct subsets which may merit distinct therapeutic approaches in future trials.

Absence of Muscle Spindles in Human Facial Muscles

Background: It has not previously been clarified if human facial muscles possess either typical muscle spindles, spindle-like structures or no spindles at all. Methods: Six patients from the Institute of Anatomy were used to investigate the Mm. orbiculares oculi and orbiculares oris. The age ranged from 53 to 91 years; three were males, three females. The specimens were imbedded in paraffin and examined in serial sections. Muscles were prepared for histological (H&E and Elastica van Gieson) and immunohistochemical (protein S-100, neurofilaments, and PGP 9.5) stainings. Results: All biopsies showed skeletal muscle fibers with some discrete alterations according to age. In some regions there were many small peripheral nerve fibers in close vicinity to muscle; but spindle-like structures with intrafusal fibers could not be detected. There were no signs of a sensorimotor double innervation of striated muscle fibers. Discussion: Previous investigators came to conflicting results regarding the absence or presence of muscle spindles in human facial muscles. However, in previous studies serial sections and immunohistochemical investigations had not been performed. Our observation of lacking muscle spindles in the facial muscles has several clinical and neurophysiological implications: (1) Pathological central muscle spindle afferent processing is assumed in the pathophysiology of dystonia. Although botulinum toxin leads to a denervation of muscle spindles its effect in the treatment of facial dystonias cannot be related to that. (2) A central facial paresis shows no signs of spasticity even in the chronic stage. Spasticity results in part from abnormal spinal processing of proprioceptive input mediated by Ia afferents. Thus, the absence of muscle spindles in the facial muscles may explain the lack of spasticity in these muscles. (3) In contrast to limb muscles, electrical facial and trigeminal nerve stimulations do not evoke silent periods and facial muscles show no facilitation of magnetic evoked potentials by vibration stimuli. Both phenomena are mediated by Ia afferents and their absence can be explained by the lack of muscle spindles.

Effect of tumor necrosis factor‐α and interferon‐γ on intestinal P‐glycoprotein expression, activity, and localization in Caco‐2 cells

The P‐glycoprotein (Pgp), a drug efflux pump, is expressed in intestinal epithelial cells, where it constitutes a barrier against xenobiotics. In inflammatory bowel disease, a dysregulation in the production of tumor necrosis factor (TNF)α and interferon (IFN)γ, and an alteration of Pgp expression and activity have been reported. The aim of this study was to investigate the effects of TNFα and IFNγ on intestinal Pgp expression, activity, and localization in Caco‐2 cells grown on filters. TNFα induced both a strong time‐dependent diminution (−56%) of MDR1 mRNA (semiquantitative reverse transcription polymerase chain reaction) and a significant decrease of unidirectional transport of rhodamine 123 after 48 h of exposure at 10 ng/mL. By confocal laser scanning microscopy, the Pgp was mainly localized to the apical plasma membrane of both control and TNFα‐treated cells. By contrast, IFNγ induced up‐regulation of both mRNA MDR1 and Pgp protein expression without incidence on Pgp activity. Interestingly, a colocalization of Pgp with lateral F‐actin was observed. Associated with TNFα, IFNγ produced neither an antagonist nor synergistic effect on Pgp activity. In conclusion, our results demonstrate an inhibitory effect of TNFα and no effect of IFNγ on Pgp transport activity using rhodamine 123 as a substrate. Mechanisms of action of these cytokines remain to be studied. © 2004 Wiley‐Liss, Inc. and the American Pharmacists Association J Pharm Sci 93:1524–1536, 2004

Metabolic interaction between cyclosporine and sirolimus.

When the immunosuppressants cyclosporine (CsA) and sirolimus (SRL) are co-administered to transplant patients, lower doses are used than when either drug is given alone. Since both drugs share similar transport and metabolic pathways, there is the potential for an interaction leading to unpredictable effects. Furthermore, both drugs affect the activity of cytochrome P450 3A1/2 (CYP3A1/2), the rat parallel to human CYP3A4, and the multidrug transporter P-glycoprotein (Pgp). To clarify the role of metabolic enzymes and membrane transporters involved in the disposition of both drugs, we examined hepatic CYP3A1/2, Pgp, and multidrug resistance gene (mdr) mRNA during chronic therapy with CsA and SRL in salt-depleted rats. Specifically, rats were given intravenous doses of CsA 2.5 mg/kg and SRL 1 mg/kg, alone or in combination, for two weeks via constant rate intravenous infusion. CsA treatment inhibited hepatic CYP3A1/2 protein expression, catalytic activity, and mRNA levels. SRL dosing suppressed CYP3A1/2 protein expression and catalytic activity, without affecting mRNA. With combined dosing, however, there was a much greater reduction. Hepatic Pgp protein levels were elevated after treatment with either drug alone, as well as with combined dosing. Compared to controls, there were significant increases in mdr1a and mdr1b mRNA levels in all treatment groups, with the combined drugs causing the greatest increase. Both CYP3A1/2 and Pgp participate in the disposition of CsA and SRL in rats. Changes in the individual activities of CYP3A1/2 and Pgp may contribute to an interaction between CsA and SRL resulting in unanticipated effects during chronic therapy.

Increased expression of the multidrug transporter P-glycoprotein in limbic brain regions after amygdala-kindled seizures in rats

Increased expression of the multidrug transporter P-glycoprotein (Pgp; ABCB1) has previously been found in epileptogenic brain tissue from patients with pharmacoresistant temporal lobe epilepsy (TLE) as well as in the hippocampus and other limbic brain regions in the rat kainate model of TLE. Approaches to the quantification of Pgp expression have mainly been based on subjective visual estimation of the level of Pgp immunoreactivity in brain sections. In the present study, computer-assisted image analysis based on optical density (OD) measurements was used to examine immunohistochemical expression of Pgp in the kindling model of TLE. Sections from kainate-treated rats were used for comparison. Using diaminobenzidine as chromogen, Pgp was exclusively located in brain capillary endothelial cells, which was confirmed by double-labeling with an antibody against the endothelial glucose transporter (GLUT-1). After kainate-induced seizures, the intensity of endothelial Pgp staining significantly increased by 70–80% in the dentate gyrus. A significant, albeit less marked increase in Pgp expression in this area was also seen after amygdala-kindled seizures. Furthermore, Pgp was upregulated after kindling in the hilus of the dentate gyrus, the CA1 and CA3 sectors of the hippocampus, and the piriform and cerebral cortex. In kindled rats, most Pgp alterations occurred ipsilateral to the electrode in the basolateral amygdala. The data demonstrate that computer-assisted image analysis using OD is an accurate and rapid method to determine the relative amount of Pgp protein in brain sections and the effects of seizures on this multidrug transporter. The fact that Pgp overexpression in brain capillary endothelial cells occurs in two established models of difficult-to-treat TLE substantiates the notion that seizure-induced upregulation of Pgp contributes to multidrug resistance (MDR) in epilepsy.

Herbal Modulation of P‐Glycoprotein

P‐glycoprotein (Pgp) is a 170 kDa phosphorylated glycoprotein encoded by human MDR1 gene. It is responsible for the systemic disposition of numerous structurally and pharmacologically unrelated lipophilic and amphipathic drugs, carcinogens, toxins, and other xenobiotics in many organs, such as the intestine, liver, kidney, and brain. Like cytochrome P450s (CYP3A4), Pgp is vulnerable to inhibition, activation, or induction by herbal constituents. This was demonstrated by using an ATPase assay, purified Pgp protein or intact Pgp‐expressing cells, and proper probe substrates and inhibitors. Curcumin, ginsenosides, piperine, some catechins from green tea, and silymarin from milk thistle were found to be inhibitors of Pgp, while some catechins from green tea increased Pgp‐mediated drug transport by heterotropic allosteric mechanism, and St. John's wort induced the intestinal expression of Pgp in vitro and in vivo. Some components (e.g., bergamottin and quercetin) from grapefruit juice were reported to modulate Pgp activity. Many of these herbal constituents, in particular flavonoids, were reported to modulate Pgp by directly interacting with the vicinal ATP‐binding site, the steroid‐binding site, or the substrate‐binding site. Some herbal constituents (e.g., hyperforin and kava) were shown to activate pregnane X receptor, an orphan nuclear receptor acting as a key regulator of MDR1 and many other genes. The inhibition of Pgp by herbal constituents may provide a novel approach for reversing multidrug resistance in tumor cells, whereas the stimulation of Pgp expression or activity has implication for chemoprotective enhancement by herbal medicines. Certain natural flavonols (e.g., kaempferol, quercetin, and galangin) are potent stimulators of the Pgp‐mediated efflux of 7,12‐dimethylbenz(a)‐anthracene (a carcinogen). The modulation of Pgp activity and expression by these herb constituents may result in altered absorption and bioavailability of drugs that are Pgp substrates. This is exemplified by increased oral bioavailability of phenytoin and rifampin by piperine and decreased bioavailability of indinavir, tacrolimus, cyclosporine, digoxin, and fexofenadine by coadministered St. John's wort. However, many of these drugs are also substrates of CYP3A4. Thus, the modulation of intestinal Pgp and CYP3A4 represents an important mechanism for many clinically important herb–drug interactions. Further studies are needed to explore the relative role of Pgp and CYP3A4 modulation by herbs and the mechanism for the interplay of these two important proteins in herb–drug interactions.

Ligand Binding Determines Whether CD46 Is Internalized by Clathrin-coated Pits or Macropinocytosis

CD46 is a ubiquitous human cell surface receptor for the complement components C3b and C4b and for various pathogens, including the measles virus and human herpes virus 6. Ligand binding to CD46 affects (i) protection of autologous cells from complement attack by breakdown of complement components, (ii) intracellular signals that affect the regulation of immune cell function, (iii) antigen presentation, and (iv) down-regulation of cell surface CD46. Recent evidence indicates that CD46 signaling can link innate and acquired immune function. The molecular mechanisms for these processes and the importance of intracellular trafficking of the receptor have not yet been elucidated. We demonstrate here that, in nonlymphoid cells, CD46 is constitutively internalized via clathrin-coated pits, traffics to multivesicular bodies, and is recycled to the cell surface. However, cross-linking of CD46 at the cell surface, by either multivalent antibody or by measles virus, induces pseudopodia that engulf the ligand in a process similar to macropinocytosis, and leads to the degradation of cell surface CD46. Thus, we have elucidated two pathways for CD46 internalization, which are regulated by the valence of cross-linking of CD46 and which utilize either clathrin-coated pits or pseudopodial extension. This has important implications for CD46 signaling, antigen presentation, CD46 down-regulation, and engulfment of pathogens.

In vitro antileukemic effect of a new anthracycline analogue, MEN 11079

The biological activity of MEN 11079, a new daunorubucin analogue with a fluorine atom in C 8 of ring A, was investigated in the human leukemic cell lines K-562 and in mononuclear cells (MNCs) of 40 patients with acute myeloid leukemia (AML) and the activity compared to two well-characterized anthracyclines, idarubicin (IDA) and doxorubicin (DOXO). IDA and MEN 11079 were more active than DOXO in cytotoxicity tests (WST-1 assay). IDA and MEN 11079 ID 50 values were also significantly different from each other (K-562: P=0.038; MNCs: P=0.003). Moreover, the range was 0.002–4.300 μM for IDA and 0.002–0.670 μM for MEN 11079, in the MNCs. Therefore, the latter appeared to assure a smaller variability of response in the AML cells. Apoptosis assays (performed using Annexin-V assay and propidium iodide) and cell cycle studies demonstrated that the MEN 11079 effective concentration was 10-fold lower than the DOXO and IDA ones. MDR (Pgp and MRP1 proteins), as measured by semiquantitative RT-PCR, cytofluorimetric and functional analysis of proteins, was similarly elicited by IDA and MEN 11079. In conclusion, the response of the cells to the new anthracycline indicates that there is greater cytotoxic activity of this molecule than IDA and DOXO. Its narrower ID 50 range may allow for a more predictable response in the clinical setting.

ATP binding cassette transporters and drug resistance in breast cancer.

Resistance to chemotherapy is a critical issue in the management of breast cancer patients. The nature of clinical drug resistance is likely to be multifactorial. However, in the last decade considerable attention has been dedicated to the role played by membrane transporter proteins belonging to the ATP binding cassette protein superfamily, and in particular by the MDR1 product P-glycoprotein (Pgp) and the multidrug resistance protein (MRP1). Heterogeneity of results is a common feature of studies evaluating the expression and prognostic role of these proteins, due to both methodological and biological factors. Nonetheless, Pgp and MRP1 are detected in a significant proportion of untreated breast cancers (on average 40 and 50% respectively, by immunohistochemistry), without a clear and consistent association with cancer stage. Exposure to chemotherapy increases the expression of both proteins. In vitro studies on primary cultures of breast cancer cells obtained at surgery consistently show an association between Pgp (protein) or MDR1 (mRNA) expression and resistance to chemotherapy. However, the correlation with clinical drug resistance is not as well defined. A stronger association of Pgp/MDR1 with response rates has been observed when expression or an increase in expression are detected immediately following chemotherapy. Correlations with prognosis appear more evident in studies using immunohistochemistry, in adjuvant and neoadjuvant settings. Evidence of clinical reversal of drug resistance by verapamil suggests a functional role of Pgp in drug resistance, although the significance of the evidence is generally weakened by poor trial designs. Future studies should take into account the multifactorial nature of drug resistance in breast cancer and use standardized approaches with adequate controls. Expression studies should be complemented by well-designed trials of drug-resistance reversal using target-specific chemosensitizing agents, and relating the results to the levels of expression of the target proteins.

Differential modulation of cytochrome P-450 1A and P-glycoprotein expression by aryl hydrocarbon receptor agonists and thyroid hormone in Xenopus laevis liver and intestine

Several defence mechanisms, such as cytochrome P450 1A (CYP1A) enzymes and P-glycoprotein (Pgp), may influence the intracellular concentration and consequently the toxicity of xenobiotics. The parallel expression of CYP1A and Pgp has been investigated in mammals and, to a lesser extent in fish, in search for evidence of co-ordinated responses to xenobiotic exposure. The aryl hydrocarbon receptor (AHR) agonists are well known CYP1A inducers but some of them resulted not to have a uniquely defined action on Pgp levels in mammalian and fish species. To the best of our knowledge, no detailed studies have been carried out so far on amphibians Xenopus laevis. For this reason, in this work, the time dependent responses of the hepatic CYP1A and Pgp, to the prototypical CYP1A inducers, benzo(a)pyrene (B(a)P), 3-methylcholanthrene (3MC) and 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) in X. laevis have been assessed at the protein level and compared. The responsiveness of Xenopus intestinal Pgp to these compounds has also been analysed, as the epithelial cells lining the lumen of intestine represent another preferential site of Pgp expression. In addition, since the thyroid hormone has been demonstrated to down regulate the mdr gene during Xenopus development and in primary culture of Xenopus intestinal epithelial cells, the effects of 3,3′,5-triiodo- l-thyronine (T 3) on CYP1A and Pgp protein levels have been investigated in adult organisms. Western blot evidenced that a single injection of B(a)P (100 mg/kg), 3MC (20 mg/kg), and TCDD (3 μg/kg) elicited a statistically significant induction of hepatic CYP1A at all time points considered (72, 120 and 168 h) which decreased in time. The same trend of liver CYP1A induction was observed in T 3 treated Xenopus (15 μg/kg). Unlike CYP1A induction, the modulation of hepatic and intestinal Pgp expression exhibits an heterogeneous pattern. The basal levels of hepatic and intestinal Pgp were not statistically significant affected by treatments. In particular, the hepatic Pgp levels seem not to be induced by TCDD and T 3 at all times considered in comparison to control. For the first time the modulation of CYP1A and Pgp levels by B(a)P, 3MC and in particular by TCDD and T 3 in Xenopus has been demonstrated and the results herewith indicate that the two target defence mechanisms respond to AHR agonists in a dissimilar way in terms of proteins induction in Xenopus. Moreover, these data suggest additional experiments in order to clarify the complex mechanism, which adjusts the parallel expression of CYP1A and Pgp in Xenopus.

Subcellular detection and localization of the drug transporter P-glycoprotein in cultured tumor cells.

In vitro studies on the cellular location of P-glycoprotein (Pgp) are reported with the aim to clarify the relationship between its intracellular expression and the multidrug resistance (MDR) level of tumor cells. Pgp was found abnormally expressed on the plasma membrane of tumor cells with "classical" MDR phenotype. However, Pgp was also often detected on the nuclear envelope and on the membrane of cytoplasmic organelles. The hypothesis that this drug pump maintains a transport function when located in these compartments, is still under debating. Our results, together with those obtained by other researchers, demonstrate that cytoplasmic Pgp regulates the intracellular traffic of drugs so that they are no more able to reach their cellular targets. In particular, we revealed that in MDR breast cancer cells (MCF-7) a significant level of Pgp was expressed in the Golgi apparatus. A similar result was found in human melanoma cell lines, which never undergone cytotoxic drug treatment and did not express the transporter molecule on the plasma membrane. A strict relationship between intracellular Pgp and intrinsic resistance was demonstrated in a human colon carcinoma (LoVo) clone, which did not express the drug transporter on the plasma membrane. Finally, a structural and functional association between Pgp and ERM proteins has been discovered in drug-resistant human T- lymphobastoid cells (CEM-VBL 100). Our findings strongly suggest a pivotal role of the intracytoplasmic Pgp in the transport of drugs into cytoplasmic vesicles, thus actively contributing to their sequestration and transport outwards the cells. Thus, intracellular Pgp seems to represent a complementary protective mechanism of tumor cells against cytotoxic agents.

The Medicinal Chemistry of Multidrug Resistance (MDR) Reversing Drugs.

AbstractFor Abstract see ChemInform Abstract in Full Text.

Effect of interleukin-2 on intestinal P-glycoprotein expression and functionality in mice.

P-glycoprotein (Pgp), an active drug transporter expressed in enterocytes, can reduce intestinal absorption of drugs. Until now, interleukin-2 (IL2) has been reported as a Pgp modulator only in vitro. The present study examines the effects in vivo of IL2 after chronic treatment on intestinal Pgp protein expression and activity. This work also describes the effects of IL2 on the oral bioavailability of a Pgp substrate (digoxin) and of a Pgp/CYP3A cosubstrate (saquinavir). Human recombinant interleukin-2 (rIL2), administered to mice at 9 million international units/kg by intraperitoneal route twice daily for 4 days, led to a decrease in intestinal Pgp protein expression evaluated by Western blot with C219 antibody. In an in vitro everted gut sac model, rIL2 pretreatment decreased the Pgp-mediated transport of rhodamine 123 across mouse intestine by 37%. Moreover, rIL2 pretreatment markedly raised the area under the curve of orally administered digoxin from 3.5 +/- 0.5 to 9.7 +/- 1.5 mg min l(-1) as a consequence of the reduction in intestinal Pgp activity. rIL2 treatment increased saquinavir bioavailability from 2.5 to 4.5%, showing that first-pass metabolism is not affected and that Pgp by itself has only a moderate effect on saquinavir oral bioavailability. In conclusion, rIL2 pretreatment reduces intestinal Pgp protein expression and activity in mice. However, the effect of such a treatment on drug bioavailability depends on the extent of their metabolism by CYP3A.

P-Glycoprotein Expression in Mouse Brain Increases with Maturation

The mdr1a isoform of P-glycoprotein (Pgp) is an integral plasma membrane efflux pump expressed in adult brain capillary endothelial cells and astrocytes of the blood-brain barrier. We determined the developmental pattern of Pgp expression in brain tissue at embryonic day 16 (E16), day of life 0 (D0), day of life 7 (D7), day of life 21 (D21), and adults (Ad). The relative expression of Pgp mRNA and protein was indexed as a percent (mean ± SEM) of D0 levels. Pgp mRNA levels increased significantly (p < 0.01) with maturation (E16: 75 ± 8%; D21: 303 ± 37%, and Ad: 1,160 ± 120%). Similarly, Pgp protein expression was observed at E16 and increased significantly (p < 0.01) during development (E16: 52 ± 8%; D7: 187 ± 23%; D21: 440 ± 48%, and Ad: 441 ± 56%). This developmental pattern of enhanced blood-brain barrier Pgp expression with maturation was confirmed by immunohistochemistry. We conclude that (i) Pgp expression in mouse brain is limited during late embryogenesis and the newborn period; (ii) Pgp expression increases markedly with postnatal maturation, and (iii) by D21 brain Pgp protein expression approximates adult levels.

Expression of P-glycoprotein in the chicken

The multidrug resistance gene product, P-glycoprotein, may act as a defense mechanism against natural and man-made environmental toxins. Like mammals, chickens show high levels of P-glycoprotein expression in the liver, small intestine, and kidney. Expression of P-glycoprotein rapidly increased with age in the liver and kidney reaching a plateau by 2 and 4 days of age, respectively; however, expression of P-glycoprotein in the duodenum did not significantly change with age. Addition of dietary antibiotics (monensin, bacitracin), as models for dietary toxins, altered P-glycoprotein expression. Monensin increased P-glycoprotein expression in the liver and duodenum. Bacitracin reduced P-glycoprotein expression by 45% in the liver, but did not alter expression in the duodenum. Intraperitoneal injection of E. coli lipopolysaccharide, a model for acute inflammation, rapidly increased expression of Pgp protein in the liver (∼2-fold). Expression then declines to pre-induction levels by 24 h. Similar responses were observed in the spleen and kidney but not the duodenum. These results confirm the presence of an avian P-glycoprotein homologue and suggest that dietary constituents regulate the expression of P-glycoprotein. Changes in P-glycoprotein expression may represent an important physiological response to foods containing toxins and an important component of the acute phase immune response.

Ontogeny of P-Glycoprotein in Mouse Intestine, Liver, and Kidney

BackgroundP-glycoprotein (Pgp) is an ATP-dependent, integral plasma-membrane efflux pump that is constitutively expressed on (i) adult apical brush-border epithelial cells of the intestine, (ii) the bile canalicular face of hepatocytes,...

Nerve Cells in Transplanted Pancreatic Islets: No Effects of Cyclosporin or Tacrolimus on Immediate Neuronal Survival

Previous experiments have demonstrated that neuronal cells within pancreatic islets survive the isolation procedure and constitute an integral part of transplanted pancreatic islets. The aim of the present study was to investigate to what extent immunosuppressive drugs affects the acute survival of intra-islet neurons after pancreatic islet transplantation. For this purpose, C57BL/6 mice were syngeneically transplanted with 250 islets under the renal capsule. The animals were treated for 7 consecutive days with subcutaneous injections of cyclosporin, tacrolimus or vehicle. After this, the animals were killed and the grafts were removed, fixed and stained for the presence of the neuron-specific protein PGP 9.5. The number of nerves were then morphologically quantitated. No differences between the experimental groups were seen, and the number of nervous elements were approximately 5 per mm2 in all animals. It is concluded that immunosuppressive treatment does not affect the acute survival of graft neurons after experimental islet transplantation.

Identification of homologs of the mammalian P-glycoprotein in the mussel, Perna perna

The ATP-binding cassette superfamily of proteins includes various ATP-driven transporters that are structurally characterized by a conserved ATP-binding domain. This family of proteins is able to transport drugs, inorganic ions, amino acids, proteins and sugars, and includes P-glycoprotein (Pgp), which is responsible for multidrug resistance in certain mammalian tumour cells. We became interested in the possible presence of Pgp homologs in Perna perna since this organism is able to survive in contaminated sites suggesting the existence of a multixenobiotic resistance mechanism. Polymerase chain reaction amplification with degenerate primers homologous to conserved regions of the Pgp protein was used to detect the presence of Pgp-like genes in genomic DNA from mussels. Amplified fragments were obtained and sequenced. Also, immunodetection of Pgp homologs was performed with the C219 monoclonal antibody. Our results confirm the existence of a gene coding for a Pgp homolog as well as the expression of a putative Pgp protein in P. perna. The mechanisms underlying the regulation of the expression of this gene are under investigation.

Increased expression of multidrug resistance related proteins Pgp, MRP1, and LRP/MVP occurs early in colorectal carcinogenesis.

AIM: To analyse the expression of multidrug resistance (MDR) related proteins at different steps in colorectal carcinogenesis. METHODS: The presence of three MDR related proteins (Pgp, MRP1, and LRP/MVP) was...