Characterization of monocarboxylate transport in human kidney HK-2 cells.
The objectives of this study were to characterize the expression and function of monocarboxylate transporters (MCTs) in human kidney HK-2 cells and to compare the expression of MCTs in HK-2 cells to that found in human kidney. mRNA and protein expression of MCTs were determined by RT-PCR and Western analyses, respectively, while immunofluorescence staining was used to determine the membrane localization of MCT1. The driving force, transport kinetics, and inhibition of two MCT substrates, D-lactate and butyrate, were characterized in HK-2 cells. mRNA of MCT1, -2, -3, -4 isoforms were present in HK-2 cells and in human kidney cortex. MCT1 was present predominantly on the basal membranes of HK-2 cells. The cellular uptake of D-lactate and butyrate exhibited pH- and concentration-dependence (D-lactate, Km of 26.5 +/- 2.2 mM and Vmax of 72.0 +/- 14.5 nmol mg-1 min-1; butyrate, Km of 0.8 +/- 0.3 mM, Vmax of 29.3 +/- 2.5 nmol mg-1 min-1, and a diffusional clearance of 2.1 microL mg-1 min-1). The uptake of D-lactate and butyrate by HK-2 cells was inhibited by MCT analogues and the classical MCT inhibitors alpha-cyano-4-hydroxycinnamate, pCMB, and phloretin. The uptake of D-lactate and butyrate by HK-2 cells significantly decreased after transfection with small-interference RNA for MCT1. In summary, MCTs were present in both HK-2 cells and human kidney cortex, and HK-2 cells exhibited polarized MCT expression and pH-dependent transport of D-lactate and butyrate. Our results also support the usefulness of HK-2 cells as an in vitro model for studying monocarboxylate transport in renal proximal tubule cells.
Wang
Qi
Q
Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, University at Buffalo, State University of New York, Amherst, New York 14260, USA.
Lu
Ye
Y
Yuan
Min
M
Darling
Inger M
IM
Repasky
Elizabeth A
EA
Morris
Marilyn E
ME
D001692:Biological Transport / Q000502:physiology*; D002087:Butyrates / Q000493:pharmacokinetics; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D007672:Kidney Cortex / Q000378:metabolism / Q000502:physiology; D019344:Lactic Acid / Q000493:pharmacokinetics; D027501:Monocarboxylic Acid Transporters / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology; D009124:Muscle Proteins / Q000378:metabolism; D020033:Protein Isoforms / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D012964:Sodium / Q000494:pharmacology; D027981:Symporters / Q000037:antagonists & inhibitors / Q000378:metabolism; D000099233:Monocarboxylate Transport Protein 1
D002087:Butyrates; D019344:Lactic Acid; D027501:Monocarboxylic Acid Transporters; D009124:Muscle Proteins; D020033:Protein Isoforms; D034741:RNA, Small Interfering; D012964:Sodium; D027981:Symporters; D000099233:Monocarboxylate Transport Protein 1; C406962:SLC16A4 protein, human; C490427:SLC16A7 protein, human