The influence of inhibitors of P-glycoprotein (verapamil [VE], cyclosporine [CY], and GF120918 [GF]) on the cell handling of macrolides (erythromycin [ERY], clarithromycin [CLR], roxithromycin [ROX], azithromycin [AZM], and telithromycin [TEL]) was examined in J774 murine macrophages. The net influx rates of AZM and TEL were increased from 2-to 3.5-fold in the presence of these inhibitors, but their efflux was slowed only marginally. At 3 h, the inhibitors increased the levels of AZM, ERY, and TEL accumulation approximately three-to fourfold (the effect of VE, however, was lower) but did not influence CLR accumulation (
1). P-glycoprotein was detected by immunostaining at the cell surface as well as in intracellular vacuoles (endosomes and lysosomes). The data suggest that the influx of AZM, ERY, TEL, and ROX is adversely influenced by the activity of P-glycoprotein in J774 macrophages, resulting in suboptimal drug accumulation.Active drug transporters have been described in both procaryotic and eucaryotic cells. Originally described as conferring resistance to anticancer agents in cancer cells, antibiotics in bacteria, or antifungal agents in fungi, these proteins appear today to be part of a very general mechanism that cells have developed to protect themselves from invasion by diffusible, foreign molecules (for a review, see reference 37). In this context, the occurrence of antibiotic transporters in eucaryotic cells has become a common observation (7, 33). More specifically, P-glycoprotein (also referred to as MDR1) and MRP, which are expressed in most cell types and which transport a large variety of drugs, have received much attention. These two types of transporters belong to the superfamily of ATP binding cassette transporters and use ATP hydrolysis as an energy source (28). They play a key role in drug disposition by modulating drug transport through epithelia and other biological barriers to an extent that was completely unsuspected only a few years ago (1).Focusing on macrolides, erythromycin has been shown to be transported by P-glycoprotein in Caco-2 intestinal cells (29, 34). In parallel, erythromycin and azithromycin are capable of inhibiting the transport of various substrates of the P-glycoprotein in epithelial cells in vitro as well as in vivo (9,12,13,23,30,31,39). Yet, little is known about the role of efflux transporters in the handling of macrolides by macrophages, in which these drugs are known to accumulate in large amounts (2,3,20,24).In the present study, we have examined directly in macrophages the potential influence of P-glycoprotein and MRP on the accumulation and efflux of five macrolides of clinical interest. We used both broad-spectrum, nonspecific inhibitors of P-glycoprotein (verapamil and cyclosporine) and MRP (probenecid and gemfibrozil) and the specific P-glycoprotein modulator GF120918 (11, 15). We selected the murine J774 murine macrophage line since much is already known about the dispositions of macrolides in these cells (2,3,36).
MATERIALS AND METHODS
Cells.We used J77...