Identification of the efflux transporter of the fluoroquinolone antibiotic ciprofloxacin in murine macrophages: studies with ciprofloxacin-resistant cells.
Ciprofloxacin, the most widely used totally synthetic antibiotic, is subject to active efflux mediated by a MRP-like transporter in wild-type murine J774 macrophages. To identify the transporter among the seven potential Mrps, we used cells made resistant to ciprofloxacin obtained by long-term exposure to increasing drug concentrations (these cells show less ciprofloxacin accumulation and provide a protected niche for ciprofloxacin-sensitive intracellular Listeria monocytogenes). In the present paper, we first show that ciprofloxacin-resistant cells display a faster efflux of ciprofloxacin which is inhibited by gemfibrozil (an unspecific MRP inhibitor). Elacridar, at a concentration known to inhibit P-glycoprotein and breast cancer resistance protein (BCRP), only slightly increased ciprofloxacin accumulation, with no difference between resistant and wild-type cells. Analysis at the mRNA (real-time PCR) and protein (Western blotting) levels revealed an overexpression of Mrp2 and Mrp4. Mrp4 transcripts, however, were overwhelmingly predominant (45% [wild-type cells] to 95% [ciprofloxacin-resistant cells] of all Mrp transcripts tested [Mrp1 to Mrp7]). Silencing of Mrp2 and Mrp4 with specific small interfering RNAs showed that only Mrp4 is involved in ciprofloxacin transport in both ciprofloxacin-resistant and wild-type cells. The study therefore identifies Mrp4 as the most likely transporter of ciprofloxacin in murine macrophages but leaves open a possible common upregulation mechanism for both Mrp4 and Mrp2 upon chronic exposure of eukaryotic cells to this widely used antibiotic.
Marquez
Béatrice
B
Unité de Pharmacologie Cellulaire et Moléculaire, Louvain Drug Research Institute, UCL 7370 Avenue Mounier 73, B-1200 Brussels, Belgium.
Caceres
Nancy E
NE
Mingeot-Leclercq
Marie-Paule
MP
Tulkens
Paul M
PM
Van Bambeke
Françoise
F
D000166:Acridines / Q000494:pharmacology; D000818:Animals; D000890:Anti-Infective Agents / Q000493:pharmacokinetics*; D015153:Blotting, Western; D002460:Cell Line; D019742:Chemokines, CC / Q000032:analysis / Q000502:physiology; D002939:Ciprofloxacin / Q000493:pharmacokinetics*; D015248:Gemfibrozil / Q000494:pharmacology; D020868:Gene Silencing; D019402:Macrophage Inflammatory Proteins / Q000032:analysis / Q000502:physiology; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000032:analysis / Q000235:genetics / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D044005:Tetrahydroisoquinolines / Q000494:pharmacology
D000166:Acridines; D000890:Anti-Infective Agents; D019742:Chemokines, CC; D002939:Ciprofloxacin; D015248:Gemfibrozil; D019402:Macrophage Inflammatory Proteins; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D012333:RNA, Messenger; D044005:Tetrahydroisoquinolines; C489540:Abcc4 protein, mouse; C095218:Ccl9 protein, mouse; C083501:Elacridar; C421109:multidrug resistance-associated protein 1