Contribution of mdr1b-type P-glycoprotein to okadaic acid resistance in rat pituitary GH 3 cells

Abstract

Okadaic acid as well as other, structurally different, inhibitors of serine/threonine phosphatases 1 and 2A induce apoptosis in pituitary GH3 cells. Incubation with stepwise raised concentrations of okadaic acid resulted in the isolation of cells that were increasingly less sensitive to the cytotoxic effect of this agent. After about 18 months cells were selected that survived at 300 nM okadaic acid, which is about 30 times the initially lethal concentration. This study revealed that a major pharmacokinetic mechanism underlying cell survival was the development of a P-glycoprotein-mediated multidrug resistance (MDR) phenotype. The increase in mRNA levels of the mdr1b P-glycoprotein isoform correlated with the extent of drug resistance. Functional assays revealed that increasing drug resistance was paralleled by a decreased accumulation of rhodamine 123, a fluorescent dye which is a substrate of mdr1-mediated efflux activity. Resistance could be abolished by structurally different chemosensitizers of P-glycoprotein function like verapamil and reserpine but not by the leukotriene receptor antagonist MK571 which is a modulator of the multidrug resistance-associated protein (MRP). Okadaic acid resistance included cross-resistance to other cytotoxic agents that are substrates of mdr1-type P-glycoproteins, like doxorubicin and actinomycin D, but not to non-substrates of mdr1, e.g. cytosine arabinoside. Thus, functional as well as biochemical features support the conclusion that okadaic acid is a substrate of the mdr1-mediated efflux activity in rat pituitary GH3 cells. Maintenance of resistance after withdrawal of okadaic acid as well as metaphase spreads of 100 nM okadaic acid-resistant cells suggested a stable MDR genotype without indications for the occurrence of extrachromosomal amplifications, e.g. double minute chromosomes.

Central Nervous System (CNS) Delivery of Glucocorticoids Is Fine-Tuned by Saturable Transporters at the Blood-CNS Barriers and Nonbarrier Regions

“…Together, these findings support our results and provide evidence that P-gp exerts a negligible effect in the distribution of [ 3 H]cortisol and [ 3 H]corticosterone at the BCSFB. Because P-gp has also been detected in rat and human pituitary cells (36, 37), it can be concluded that although P-gp is present and functioning in the murine choroid plexus and in the pituitary gland, P-gp does not significantly affect the accumulation of [ 3 H]cortisol or [ 3 H]corticosterone in these regions.…”

Expression Analyses of Genes Related to Multixenobiotic Resistance in Mytilus galloprovincialis after Exposure to Okadaic Acid-Producing Dinophysis acuminata

“…P-glycoprotein is a good candidate to expel okadaic acid out of the cells in bivalve mollusks because it transports a wide variety of structurally unrelated hydrophobic amphipathic compounds (such as okadaic acid) across membranes [25]. Furthermore, both functional and biochemical data support the proposal that okadaic acid is a substrate of the P-gp-mediated efflux activity in rat pituitary GH3 cells [6]. In addition, Ehlers et al [26] have shown that OA is a substrate of human P-gp and that P-gp is involved in the elimination of OA from cells using two different transwell models: (i) caco-2 cell monolayer endogenously expressing human P-gp, simulating the intestinal barrier and (ii) MDCK-II cell monolayer stably over-expressing P-gp.…”

“…Thus, OA blocks the dephosphorylation of proteins that are substrates of protein kinases, affecting many basic processes, such as the regulation of gene expression, cell-cycle control, cell adhesion, apoptosis, or cytoskeleton dynamics [ 4 , 5 ]. It has also been demonstrated that low concentrations of OA have cytotoxic and mutagenic effects on different cell lines and on different bivalve tissues [ 6 , 7 , 8 ].…”