PEPT1-Mediated Cefixime Uptake into Human Intestinal Epithelial Cells Is Increased by Ca ²⁺ Channel Blockers

Abstract

The ␤-lactam antibiotic group encompasses the penicillins, cephalosporins, carbapenems, and monobactams (13). The high intestinal absorption rates of orally active ␤-lactams are mainly explained by the fact that they serve as substrates for the intestinal peptide transporter PEPT1 (10). PEPT1 is located in the apical membranes of intestinal epithelial cells (23) and the Caco-2 human intestinal epithelial cell line (39). The peptide transporter translocates di-and tripeptides (17, 33) as well as a variety of peptidomimetic compounds, including selected immunostimulants (25, 28), angiotensin-converting-enzyme inhibitors (7, 31), and ␤-lactam antibiotics (36, 43), in a proton-coupled electrogenic mode (1).The affinities of ␤-lactams for PEPT1 show large variabilities, depending on the structure of the individual antibiotic (10, 42). Moreover, it was shown that different ␤-lactams can display completely different pH dependencies for transport into Caco-2 cells or into oocytes expressing heterologously cloned mammalian PEPT1. The optimum pH for transport of zwitterionic ␤-lactams such as cefadroxil is 6.5, whereas the dianionic cefixime is effectively transported only at pH values Ͻ6.0 (42). Since the intestinal epithelium has a fairly stable surface pH compartment (microclimate pH) with a pH that is Ͼ6.25 and Ͻ6.75 (22), interactions of ␤-lactams with PEPT1 in the normal gut and thus their in vivo absorption rates may be defined by the pH at the membrane surface. This might explain why cefixime, which is efficiently transported by PEPT1 in vitro at pH values below 6.0, shows a comparably low oral availability in vivo (3, 15). Other factors contributing to the absorption of ␤-lactams via PEPT1 may include regulation of transport capacity by protein kinase C (PKC)-and insulin-dependent pathways (9, 29). Understanding of those mechanisms responsible for alterations in the transport kinetics of PEPT1 could be important for specific enhancement of the absorption of drugs with low oral availabilities, such as cefixime, which is usually used for the treatment of respiratory tract infections (37). As it has been shown that rates of absorption of cefixime are increased by 30% when the Ca 2ϩ channel blocker nifedipine is coadministered with cefixime in humans (14), we were interested in studying the underlying mechanisms responsible for this increase. Although it had been suggested that neurohormonal regulation may be involved in mediating the effects of nifedipine on cefixime transport (18), we hypothesized that the alterations in intracellular free calcium ion (Ca 2ϩ in ) concentrations caused by nifedipine could enhance PEPT1 transport activity for cefixime.We therefore altered the Ca 2ϩ in concentration in Caco-2 cells, a human intestinal epithelial cell line that expresses PEPT1, with either Ca 2ϩ channel blockers or Ca 2ϩ ionophores and measured the effects on the apical influx of [ 14 -C]cefixime. Moreover, by use of intracellular pH (pH in ) measurements, we assessed whether alterations in Ca 2ϩ in levels secondarily ...

pCLCA1 lacks inherent chloride channel activity in an epithelial colon carcinoma cell line

“…The issue of A-23187 access to the membranes of mature Caco-2 cells has been addressed by others (21,25). Direct effects of A-23187 on Ca 2ϩ flux and indirect effects of A-23187 on signaling responses to changes in intracellular Ca 2ϩ concentration were maintained in differentiated Caco-2 cells grown on permeable supports identical to our culture conditions.…”

The proton oligopeptide cotransporter family SLC15 in physiology and pharmacology

“…As PEPT1 transport activity also depends on the transmembrane proton gradient, which is mainly generated by the apical Na/H exchanger, any manoeuvres that alter pH gradients and membrane potential secondarily affect PEPT1 activity [67]. In Caco-2 cells, PEPT1 expression is reduced by treatment with thyroid hormone [1] whereas Ca 2+ channel blockers, by decreasing intracellular calcium, increase PEPT1 activity acutely whereas elevating intracellular free calcium decreases the maximal transport activity [72]. A very interesting adaptation is observed in patients with short bowel syndrome whose colonic epithelial cells express substantial quantities of PEPT1 protein, which is normally not found in the human colon or present in only very low levels [77].…”

Effects of Capsaicin on Intestinal Cephalexin Absorption in Rats

“…It has been reported that PEPT1 activity is regulated by several intracellular factors, such as the activity of protein kinaseC, 34) cAMP level, 35) or Ca 2ϩ concentration. 24) In addition to these factors, an Na ϩ /H ϩ exchanger is also shown to be a factor determining PEPT1 activity. 36) That is, the exchanger is demonstrated to regulate the proton gradient across the cellular membrane, which PEPT1 requires as a driving force for its H ϩ /peptide co-transporting function.…”

“…23) In addition to the paracellular pathway of the intestinal drug absorption processes, capsaicin may affect the transcellular pathway of the process, since the intracellular Ca 2ϩ concentration is related to the drug transporting activity of various transporters including H ϩ /peptide co-transporter 1 (PEPT1). 24) It is therefore suggested with these findings and considerations that capsaicin has an impact on the intestinal drug absorption processes. However, little is known about the effect of capsaicin on intestinal drug absorption.…”