title: The involvement of intracellular calcium in the MCT-mediated uptake of lactic acid by HeLa cells.
abstract: The main object of this study was to evaluate the role of intracellular free calcium ion [Ca2+](in) in monocarboxylate transporter (MCT) mediated drug uptake by HeLa cells. It was hypothesized that alterations in the [Ca2+](in) levels affect Na+-H+ exchanger (NHE) regulated pH(in) and thereby produce the proton-motivated driving force for monocarboxylate mediated substrate transport. The changes in intracellular pH (pH(in)) and MCT mediated uptake rates of L-lactic acid by HeLa cells, a human cervical adenocarcinoma cell line, were evaluated under the conditions, whose [Ca2+](in) concentrations were altered by various calcium modulators, such as EGTA-AM (a chelator), nifedipine (a Ca2+ channel antagonist) and A23187 (an ionophore). For the purpose of comparison, the L-lactic acid uptake by HeLa cells was also evaluated under various pH(in) conditions induced by dexamethasone. The effects of the extracellular sodium concentration on the L-lactic acid uptake by HeLa cells were evaluated to determine the involvement of NHE-regulated pH changes in the MCT mediated drug uptake process. The [Ca2+](in) concentrations and pH(in) in HeLa were assessed using fluorescent probes fura-2 and 2',7'-bis[2-carboxyethyl-5-carboxyfluorescein] (BCECF), respectively. The treatment of HeLa cells with A23187 at concentrations of 50 and 100 microM enhanced [Ca2+](in) by 100% and 200% of the control, respectively. EGTA/AM (50 microM) or nifedipine (100 microM) did not cause any significant changes in the [Ca2+](in) levels, whereas EGTA/AM (100 microM) and nifedipine (200 microM) reduced the [Ca2+](in) levels by 30% and 25%, respectively, as compared with the control. A23187 at a concentration of 100 microM in the incubation medium lowered pH(in) (pH 5) and subsequently the uptake rate of lactic acid by 50% (0.47 +/- 0.03 micromol/mg protein/min) of the control. In contrast, nifedipine (200 microM) and EGTA-AM (100 microM), the calcium modulators that lowered the [Ca2+](in) levels and maintained the higher pH(in) (pH > 6) of HeLa cells, enhanced the uptake rate of lactic acid by 60% and 130% of the control, respectively. The results of this study demonstrated that there was a close correlation between the [Ca2+](in) level and pH(in) and that NHEs were involved with the MCT mediated uptake process in HeLa cells. An understanding of the role of [Ca2+](in) in the MCT mediated transport process could provide an efficient strategy to improve the systemic delivery of monocarboxylate substrates through the cervical mucosa.
authors.affiliation: Division of Pharmaceutical Sciences, The University of Missouri School of Pharmacy, Kansas City, Missouri 64110-2499, USA.
authors.forename: Sravanthi
authors.initials: S
authors.lastname: Cheeti
authors.forename: Chi H
authors.initials: CH
authors.lastname: Lee
mesh_terms: D001483:Base Sequence; D001693:Biological Transport, Active / Q000187:drug effects; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002584:Cervix Uteri / Q000378:metabolism; D002614:Chelating Agents / Q000494:pharmacology; D017931:DNA Primers / Q000235:genetics; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D005260:Female; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007476:Ionophores / Q000494:pharmacology; D007700:Kinetics; D019344:Lactic Acid / Q000378:metabolism*; D027501:Monocarboxylic Acid Transporters / Q000235:genetics / Q000378:metabolism*; D009543:Nifedipine / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012964:Sodium / Q000494:pharmacology; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics / Q000378:metabolism; D027981:Symporters / Q000235:genetics / Q000378:metabolism*; D000099233:Monocarboxylate Transport Protein 1
chemical_list: D000001:Calcimycin; D002118:Calcium; D002121:Calcium Channel Blockers; D002614:Chelating Agents; D017931:DNA Primers; D004533:Egtazic Acid; D007476:Ionophores; D019344:Lactic Acid; D027501:Monocarboxylic Acid Transporters; D009543:Nifedipine; D012333:RNA, Messenger; D012964:Sodium; D017923:Sodium-Hydrogen Exchangers; D027981:Symporters; D000099233:Monocarboxylate Transport Protein 1; C046770:EGTA acetoxymethyl ester