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Antimicrobial Agents and Chemotherapy, May 2002, p. 1375-1380, Vol. 46, No. 5
0066-4804/02/$04.00+0     DOI: 10.1128/AAC.46.5.1375-1380.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

PEPT1-Mediated Cefixime Uptake into Human Intestinal Epithelial Cells Is Increased by Ca²⁺ Channel Blockers

Uwe Wenzel, Sabine Kuntz, Simone Diestel, and Hannelore Daniel^*

Department of Food and Nutrition, Molecular Nutrition Unit, Technical University of Munich, D-85350 Freising-Weihenstephan, Germany

Received 24 May 2001/ Returned for modification 23 December 2001/ Accepted 31 January 2002

Ca²⁺ channel blockers like nifedipine have been shown to increase the oral bioavailability of ß-lactam antibiotics, such as cefixime, in humans. The molecular mode of action of Ca²⁺ channel blockers on ß-lactam absorption, however, has not yet been defined. Using the Caco-2 human intestinal epithelial cell line, we assessed whether alterations in intracellular free Ca²⁺ ion (Ca²⁺_in) concentrations by Ca²⁺ channel blockers or by Ca²⁺ ionophores affect [¹⁴C]cefixime absorption. Reduction of Ca²⁺_in levels by Ca²⁺ channel blockers (nifedipine, verapamil, diltiazem, or bepridil) at concentrations of 100 µM led to 35 to 50% increases in the cellular uptake of 1 mM [¹⁴C]cefixime. Increases in Ca²⁺_in levels by Ca²⁺ ionophores, on the other hand, led to 40% reductions in [¹⁴C]cefixime absorption. Nifedipine increased the V_max of cefixime transport by 67%, whereas the K_m of cefixime transport remained unaffected. By measuring the pH in Caco-2 cells loaded with the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5-(6)-carboxyfluorescein, we show that cefixime transport mediated by the intestinal H⁺-coupled peptide transporter PEPT1 leads to intracellular acidification. This acid load was reduced by nifedipine, although the Ca²⁺ channel blocker increased the level of H⁺ and cefixime cotransport. Increases in Ca²⁺_in levels by ionomycin enhanced the decline in intracellular pH induced by cefixime alone, although ionomycin reduced the level of H⁺ and cefixime cotransport. In conclusion, our studies demonstrate that alterations of Ca²⁺_in levels, e.g., by Ca²⁺ channel blockers, affect pH regulatory systems, such as apical Na⁺ and H⁺ exchange, and thereby alter the H⁺ gradient that serves as the driving force for uptake of ß-lactams into intestinal epithelial cells.

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* Corresponding author. Mailing address: Department of Food and Nutrition, Hochfeldweg 2, D-85350 Freising-Weihenstephan, Germany. Phone: 49 8161/71-3400. Fax: 49 8161/71-3999. E-mail: daniel{at}wzw.tum.de.

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Antimicrobial Agents and Chemotherapy, May 2002, p. 1375-1380, Vol. 46, No. 5
0066-4804/02/$04.00+0     DOI: 10.1128/AAC.46.5.1375-1380.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

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