Identification of transport pathways for citric acid cycle intermediates in the human colon carcinoma cell line, Caco-2.
Citric acid cycle intermediates are absorbed from the gastrointestinal tract through carrier-mediated mechanisms, although the transport pathways have not been clearly identified. This study examines the transport of citric acid cycle intermediates in the Caco-2 human colon carcinoma cell line, often used as a model of small intestine. Inulin was used as an extracellular volume marker instead of mannitol since the apparent volume measured with mannitol changed with time. The results show that Caco-2 cells contain at least three distinct transporters, including the Na+-dependent di- and tricarboxylate transporters, NaDC1 and NaCT, and one or more sodium-independent pathways, possibly involving organic anion transporters. Succinate transport is mediated mostly by Na+-dependent pathways, predominantly by NaDC1, but with some contribution by NaCT. RT-PCR and functional characteristics verified the expression of these transporters in Caco-2 cells. In contrast, citrate transport in Caco-2 cells occurs by a combination of Na+-independent pathways, possibly mediated by an organic anion transporter, and Na+-dependent mechanisms. The non-metabolizable dicarboxylate, methylsuccinate, is also transported by a combination of Na+-dependent and -independent pathways. In conclusion, we find that multiple pathways are involved in the transport of di- and tricarboxylates by Caco-2 cells. Since many of these pathways are not found in human intestine, this model may be best suited for studying Na+-dependent transport of succinate by NaDC1.
Weerachayaphorn
Jittima
J
Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, TX 77555-0645, USA.
Pajor
Ana M
AM
D001692:Biological Transport / Q000187:drug effects; D018938:Caco-2 Cells; D002951:Citrates / Q000378:metabolism; D002952:Citric Acid Cycle / Q000187:drug effects; D003110:Colonic Neoplasms / Q000378:metabolism*; D029382:Dicarboxylic Acid Transporters / Q000235:genetics / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D005110:Extracellular Space / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism; D007700:Kinetics; D008954:Models, Biological; D027361:Organic Anion Transporters / Q000378:metabolism; D029363:Organic Anion Transporters, Sodium-Dependent / Q000235:genetics / Q000378:metabolism; D010969:Plastics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012964:Sodium / Q000494:pharmacology; D013379:Substrate Specificity / Q000187:drug effects; D013386:Succinates / Q000378:metabolism; D027981:Symporters / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D002951:Citrates; D029382:Dicarboxylic Acid Transporters; D027361:Organic Anion Transporters; D029363:Organic Anion Transporters, Sodium-Dependent; D010969:Plastics; D012333:RNA, Messenger; C092524:SLC13A2 protein, human; C470492:SLC13A5 protein, human; D013386:Succinates; D027981:Symporters; D012964:Sodium