Background: 

Over-expression of multidrug transporters is a well-known mechanism of resistance to anticancer agents in eucaryotic cells, and to antibiotics in prokaryotic cells. In vitro, their overexpression can be obtained by prolonged exposure to drug substrates. We have examined here whether chronical exposure to fluoroquinolone antibiotics could trigger a phenotype of resistance in eucaryotic cells as well, based on our previous observation that ciprofloxacin (CIP) is subject for an active efflux by a MRP (multidrug resistance-associated protein) transporter in J774 macrophages, which decreases its accumulation and activity against intracellular bacteria, while moxifloxacin (MXF) was not affected (AAC 2005, 49:2429–2437).

Methods: 

J774 macrophages were exposed for several months to increasing concentrations in either CIP or MXF. CIP and MXF accumulation was measured by fluorimetry in the resulting cell lines as compared to wild-type macrophages. Expression levels of the Mrp were evaluated by real-time PCR and Western-blot. Transient silencing experiments with specific siRNA were made, and their impact on the corresponding protein contents, and CIP accumulation assessed.

Results: 

As compared to wild-type macrophages, which accumulate CIP about 4-fold and extrude it with a t_1/2 ~ 1.2 min, cells exposed to CIP showed a markedly decreased accumulation of CIP (< 1-fold) related to a faster efflux (t_1/2 < 0.1 min), while cells exposed to MXF showed a higher accumulation of CIP (15-fold) and a slower efflux (t_1/2 ~ 2.6 min). MXF accumulation was high (15-fold) and similar in all cell types.

Analysis at the mRNA and protein levels revealed an overexpression of Mrp2 and Mrp4 in macrophages exposed to CIP and a reduction of Mrp4 expression in macrophages exposed to MXF.

Silencing experiments showed that only Mrp4 silencing was associated with an increase in CIP accumulation (in both CIP-resistant and wild-type cells).

Conclusions: 

Exposure of J774 macrophages to the Mrp substrate CIP selects for resistant cells which over-express Mrp2 and Mrp4, but only Mrp4 is involved in CIP efflux. Exposure of the same cells to the non-substrate MXF selects for an 'anti'-resistant phenotype, with reduction in the expression of Mrp4. These data suggest that fluoroquinolones can differentially affect the expression of Mrp transporters in J774 macrophages, illustrating the complexity of the interactions between closely-related drugs and transporters.