Down-Regulation of Organic Anion Transporter Expression in Human Hepatocytes Exposed to the Proinflammatory Cytokine Interleukin 1β

The short-term regulation of multidrug resistance-associated protein 3 (Mrp3/MRP3) by cAMP and PKC was investigated in sandwich-cultured rat and human hepatocytes and isolated perfused rat livers. The modulator glucagon (500 nM) and the phorbol ester PMA (0.1 μM) were utilized to increase intracellular cAMP and PKC levels, respectively. In glucagon-treated rat hepatocytes, efflux of the Mrp3 substrate 5-(6)-carboxy-2′,7′-dichlorofluorescein (CDF) increased ∼1.5-fold, even in hepatocytes treated with the organic anion transporter (Oatp) inhibitor sulfobromophthalein (BSP). Confocal microscopy revealed more concentrated Mrp3 fluorescence in the basolateral membrane (less diffuse staining pattern) with glucagon treatment. PMA had no effect on Mrp3 activity or localization in sandwich-cultured rat hepatocytes. Glucagon and PMA treatment in isolated perfused rat livers resulted in a threefold increase (14 ± 4.6 μl·min−1·g liver−1) and a fourfold decrease (1.3 ± 0.3 μl·min−1·g liver−1) in CDF basolateral clearance compared with control livers (4.7 ± 2.3 μl·min−1·g liver−1), whereas CDF biliary clearance was not statistically different. In sandwich-cultured human hepatocytes, glucagon treatment resulted in a 1.3-fold increase in CDF efflux and a concomitant increase in MRP3 fluorescence in the basolateral membrane. In summary, cAMP and PKC appear to be involved in the short-term regulation of Mrp3/MRP3, as demonstrated by alterations in activity and localization in rat and human hepatocytes.

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Combination of Tenofovir and Emtricitabine plus Efavirenz: In Vitro Modulation of ABC Transporter and Intracellular Drug Accumulation

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00733-08 ◽

2008 ◽

Vol 53 (3) ◽

pp. 896-902 ◽

Cited By ~ 39

Author(s):

Laurence Bousquet ◽

Alain Pruvost ◽

Anne-Cécile Guyot ◽

Robert Farinotti ◽

Aloïse Mabondzo

Keyword(s):

Mrna Expression ◽

Organic Anion ◽

Antiretroviral Drugs ◽

Organic Anion Transporter ◽

Target Cells ◽

Efflux Transporter ◽

Anion Transporter ◽

Drug Concentrations ◽

Transporter Expression

ABSTRACT Efflux proteins have been shown to greatly affect the uptake of antiretroviral drugs by cells and to hamper their access to the human immunodeficiency virus type 1 replication site. This study evaluated the factors that may lead to drug-drug interactions between emtricitabine (FTC), tenofovir (TFV), and efavirenz (EFV), including the modulation of efflux transporter expression and function. Peripheral blood mononuclear cells from healthy volunteers were used to determine whether or not an interaction between antiretroviral drugs and target cells occurred in any combination of FTC, TFV, EFV, FTC-TFV, TFV-EFV, or FTC-TFV-EFV. Following 20 h of treatment, intracellular drug concentrations were measured by liquid chromatography-tandem mass spectrometry. Efflux transporter functionality and inhibitor drug properties were assessed by measuring fluorescent dye efflux. ABCB1 (P-glycoprotein), ABCC 1 to 6 (multidrug resistance-associated protein), and OAT (organic anion transporter) expression in response to the treatments was quantified by semiquantitative real-time PCR. Cells treated with a double combination (FTC-TFV or TFV-EFV) or the triple combination (FTC-TFV-EFV) produced higher FTC and TFV intracellular concentrations than cells treated with FTC or TFV alone. However, no change in the EFV intracellular concentration was observed. FTC tended to induce abcc5 mRNA expression and EFV tended to induce abcc1 and abcc6 mRNA expression, whereas TFV tended to reduce mdr1, abcc1, abcc5, and abcc6 mRNA expression. Under these conditions, a decrease in the functionality of ABCC was observed, and this decrease was associated with the direct inhibitory actions of these drugs. This in vitro study reveals a benefit of the combination FTC-TFV-EFV in terms of the intracellular FTC and TFV concentrations and highlights the pharmacological mechanisms that lead to this effect.

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