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P-gp Activity (p-gp + activity)
Selected AbstractsJC-1, a sensitive probe for a simultaneous detection of P-glycoprotein activity and apoptosis in leukemic cellsCYTOMETRY, Issue 3 2006Driss Chaoui Abstract Background JC-1 probe has been successfully used for the analysis of either apoptosis or P-glycoprotein (P-gp) activity. Therefore, we wanted to see if JC-1 could also simultaneously assess both, P-gp activity and apoptosis, in acute myeloid leukemia (AML) cells. Methods P-gp activity was measured using JC-1 and compared to the results of the Rhodamine 123 (Rh 123) assay in P-gp negative and P-gp positive cell lines, and 12 AML samples. For apoptosis, spontaneous apoptosis, as well as, apoptosis induced by Cytosine Arabinosine and Homoharringtonine were analyzed. Both mitochondrial red fluorescence and cytoplasmic green fluorescence of JC-1 with and without a P-gp inhibitor (Cyclosporine A : CsA) were used for the identification of apoptotic cells, and this was compared to Annexin V/PI staining. Results (1) We found a good correlation between JC-1 and Rh 123 in viable cells. Even in a small population of viable cells, P-gp positive cells emitting low red fluorescence, gained on red fluorescence after P-gp inhibition with CsA permitting an evaluation of P-gp activity. (2) We found a good correlation between the Annexin V/PI staining and JC-1 (P < 0.0001) in the assessment of apoptotic cells. Most importantly, the apoptotic cells could be distinguished by the loss of red fluorescence and the increase of green fluorescence without any change after P-gp inhibition with CsA. Conclusions JC-1 can simultaneously evaluate two important parameters involved in drug resistance in AML cells, P-gp activity and apoptosis. © 2006 International Society for Analytical Cytology [source] Grapefruit juice,drug interactions: Grapefruit juice and its components inhibit P-glycoprotein (ABCB1) mediated transport of talinolol in Caco-2 cellsJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 10 2007Whocely Victor de Castro Abstract To investigate the potential interaction between selected ingredients of grapefruit juice and, the transport of talinolol, a P-gp substrate, across Caco-2 cells monolayers was determined in the absence and presence of distinct concentrations of grapefruit juice, bergamottin, 6,,7,-dihydroxybergamottin, 6,,7,-epoxybergamottin, naringin, and naringenin. Talinolol permeability was selectively inhibited by grapefruit juice and its components. The furano coumarin, 6,,7,-epoxybergamottin, was the most potent inhibitor (IC50,=,0.7 µM), followed by 6,,7,-dihydroxybergamottin (IC50,=,34 µM) and bergamottin that did not show any inhibition at concentrations up to 10 µM. The flavonoid aglycone naringenin was around 10-fold more potent than its glycoside naringin with IC50 values of 236 and 2409 µM, respectively. The flavonoids and furanocoumarins tested in this study are in the same range of concentration they are present in the juice contributing, therefore, for the overall inhibitory effect of GFJ on P-gp activity. The in vitro data suggest that compounds present in grapefruit juice are able to inhibit the P-gp activity modifying the disposition of drugs that are P-gp substrates such as talinolol. © 2007 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 96: 2808,2817, 2007 [source] Effect of interferons on P-glycoprotein-mediated rhodamine-123 efflux in cultured rat hepatocytesJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 10 2002Yukiko Akazawa Abstract The effect of interferon (IFN)-, and IFN-, on P-glycoprotein (P-gp)-mediated efflux of rhodamin-123(Rho-123), a typical substrate of P-gp, was studied in rat hepatocytes in primary culture. After treatment with IFN-,, IFN-,, or both for 3 days, steady-state levels of Rho-123, incorporated into the hepatocytes, were measured to evaluate the P-gp activity. Whereas IFN-, did not affect the intracellular level of Rho-123, IFN-, treatment caused a significant increase of the level, suggesting that IFN-, treatment suppresses the expression of P-gp or its activity. A combination of the two types of IFN exhibited a similar effect to that of IFN-, alone. The effect of IFN-, was still observed in the presence of H2O2, which enhances the expression and activity of P-gp. Immunoblot analysis using a monoclonal antibody C219 revealed, however, that P-gp expression was increased after treatment with IFN-,, but only slightly by IFN-, treatment. These results suggest that the enhanced Rho-123 uptake of rat primary hepatocytes induced by IFN-, does not result from reduced expression of P-gp but, rather, from impaired maturation or dysfunction of the efflux transporter. © 2002 Wiley-Liss Inc. and the American Pharmaceutical Association J Pharm Sci 91:2110,2115, 2002 [source] YB-1 is upregulated during prostate cancer tumor progression and increases P-glycoprotein activityTHE PROSTATE, Issue 3 2004Pepita Giménez-Bonafé Abstract BACKGROUND Currently, the main obstacle to curing advanced prostate cancer is development of androgen independence (AI), where malignant cells acquire the ability to survive in the absence of androgens. Our initial experimental approach used cDNA microarrays to characterize changes in gene expression in the LNCaP human prostate tumor model during progression to AI. The transcription factor Y-box binding protein (YB-1) was shown to be one of the genes upregulated. We focused on increased YB-1 expression during progression in clinical specimens, and further examined one of its downstream targets, P-glycoprotein (P-gp). METHODS Northern blot analysis was performed on LNCaP tumor series, as well as immunohistochemical analyses of human prostate cancer tissue samples. YB-1 was transiently transfected and transport analysis were performed to analyze P-gp efflux activity. RESULTS YB-1 expression is markedly increased during benign to malignant transformation and further following androgen ablation. In addition, increased YB-1 expression after castration in the LNCaP model is linked to upregulation of P-gp. We demonstrate that YB-1 upregulates P-gp activity resulting in a 40% intracellular decrease in the P-gp substrate vinblastine. We have also found that P-gp increases the efflux of the endogenous androgen, dihydrotestosterone (DHT), from prostate cells and leads to decreased androgen regulated gene expression. CONCLUSIONS We hypothesize that early in prostate cancer progression, increased expression of YB-1 may increase P-gp activity which may in turn lower androgen levels in the prostate tumor cells. Suppression of androgen levels may activate cell survival pathways and lead to an adaptive survival advantage of androgen independent prostate cancer cells following androgen ablation therapy. © 2004 Wiley-Liss, Inc. [source] Expression levels of human P-glycoprotein in In Vitro cell lines: correlation between mRNA and protein levels for P-glycoprotein expressed in cellsBIOPHARMACEUTICS AND DRUG DISPOSITION, Issue 3 2009Yoshiyuki Shirasaka Abstract The purpose of this study was to investigate the correlation between mRNA and protein levels for P-glycoprotein (P-gp) expressed in various cell lines to validate the estimation of P-gp activity from its mRNA levels. P-gp expression levels in various cell monolayers, normal, P-gp-induced, P-gp-highly induced, (multidrug resistance, MDR) MDR1-knockdown (A2-2) and MDR1-knockdown (B2-2) Caco-2 cells and MDCKII/MDR1 cells, were quantified by real-time quantitative polymerase chain reaction (PCR) and western blot analysis. Both mRNA and protein levels of P-gp were lowest in the MDR1-knockdown (B2-2) Caco-2 cells, followed by the MDR1-knockdown (A2-2) Caco-2, normal Caco-2, P-gp-induced Caco-2 and P-gp-highly induced Caco-2 cells, and highest in the MDCKII/MDR1 cells. Except for the MDCKII/MDR1 cells, the protein levels of P-gp in all Caco-2 cell lines showed a linear correlation with its mRNA levels; however, although the MDR1 mRNA level in MDCKII/MDR1 cells was much higher than in the P-gp-highly induced Caco-2 cells, the protein levels were almost the same in both cells. From these findings, it was suggested that P-gp activity in MDCKII/MDR1 cells could not be estimated from its mRNA levels. Copyright © 2009 John Wiley & Sons, Ltd. [source] Up-regulation of CD147 and matrix metalloproteinase-2, -9 induced by P-glycoprotein substrates in multidrug resistant breast cancer cellsCANCER SCIENCE, Issue 11 2007Qing-Quan Li Treatment of animals bearing multidrug resistant (MDR) tumor cells with P-glycoprotein (P-gp) substrates could worsen host survival. It is assumed that this is due to increased tumor metastasis. To clarify the mechanism(s) underlying this observation, the MDR human breast cancer cell line, MCF-7/AdrR, and its sensitive parental line, MCF-7, was treated with various concentrations of P-gp substrate drugs (vincristine, paclitoxel, adriamycin) and a P-gp non-substrate drug (bleomycin) in serum-free media. Increased production of CD147, and matrix metalloproteinases (MMP)-2, -9 was observed only in MDR cancer cells exposed to P-gp substrates, as determined using real-time polymerase chain reaction, western blotting and zymography. Correspondingly, P-gp substrates significantly enhanced the in vitro invasion abilities of MCF-7/Adr cells. It was also found that the drug-induced promotion of CD147, and MMP-2, -9 was consistent with increased expression of epidermal growth factor receptor (EGFR) and that inhibition of either EGFR or P-gp activity could significantly interrupt the downstream effects, and so inhibit in vitro invasion abilities motivated by P-gp substrates. These results imply that treatment of MDR tumors with P-gp substrates could adversely affect therapeutic outcomes through modulating the production of CD147, MMP-2, -9, and EGFR, and suggest that this effect may be initiated by the transporter function of P-gp. (Cancer Sci 2007; 98: 1767,1774) [source] | ||||||||||