Human cells possess multiple nucleoside transporters (NTs) that belong to either the human equilibrative or concentrative NT (hENT: hENT1/2/3/4; hCNT: CNT1/2/3) families. In the kidney, coupling of apical hCNT3 activities to basolateral hENT1/2 activities is hypothesized to mediate renal nucleoside proximal tubular absorption while apical ENT1 may have a role in secretion. The overall aim of this research was to increase understanding of the roles of hENTs and hCNTs in renal handling of physiological nucleosides and anti-cancer nucleoside analog drugs. This was achieved by investigating the distribution of hENTs and hCNTs in human kidney tissue and the function of hENTs and hCNTs in cellular uptake and transepithelial fluxes of nucleosides in cultured human renal proximal tubule cells (hRPTCs).
Immunolocalization of hCNT3 and hENT1 in human kidney tissue revealed that hENT and hCNT3 were present in apical membranes of proximal tubules. Production and characterization of adherent hRPTC cultures demonstrated endogenous hCNT3, hENT1, and hENT2 activities. These results provided evidence for the involvement of hCNT3, hENT1, and hENT2 in renal handling of nucleosides.
Comparison of adherent hRPTC cultures derived from kidneys from different individuals demonstrated that hCNT3 activities varied between cultures. Also, the extent of cellular uptake of fludarabine, an anti-cancer nucleoside drug, and degree of cytotoxicity was reflected in the different hCNT3 activities observed between cultures. These results suggested that hCNT3 plays an important role in fludarabine renal handling and is a determinant of potential renal toxicities.
Production of polarized monolayer cultures of hRPTCs on transwell permeable inserts enabled the functional localization of hCNT3 and hENT1 to apical membranes and hENT2 to basolateral membranes. Transepithelial flux studies demonstrated that (i) apical-to-basolateral fluxes of adenosine were mediated by apical hCNT3 and basolateral hENT2, (ii) basolateral-to-apical fluxes of 2′-deoxyadenosine were mediated, in part, by apical hENT1 and basolateral hOATs, and (iii) apical-to-basolateral fluxes of fludarabine, cladribine, and clofarabine were mediated by apical hCNT3.
These studies showed that coupling of apical hCNT3 to basolateral hENT2 mediates proximal tubular nucleoside reabsorption, that coupling of basolateral human organic anion transporters (hOATs) to apical hENT1 mediates proximal tubular nucleoside secretion, and that hCNT3 is a key determinant of fludarabine proximal tubular reabsorption and cytoxicity.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:AEU.10048/433 |
| Date | 11 1900 |
| Creators | Elwi, Adam |
| Contributors | Cass, Carol (Oncology), Sawyer, Michael (Oncology), Cass, Carol (Oncology), Sawyer, Michael (Oncology), Weinfeld, Michael (Oncology), Young, James (Physiology), Cheeseman, Chris (Physiology), Gandhi, Varsha (Experimental Therapeutics, The University of Texas M.D. Anderson Cancer Center) |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | 9299343 bytes, application/pdf |
| Relation | Elwi AN, Damaraju VL, Baldwin SA, Young JD, Sawyer MB, Cass CE. Renal nucleoside transporters: physiological and clinical implications. Biochem Cell Biol. 2006; 84: 844-858., Damaraju VL, Elwi AN, Hunter C, Carpenter P, Santos C, Barron GM, Sun X, Baldwin SA, Young JD, Mackey JR, Sawyer MB, Cass CE. Localization of broadly selective equilibrative and concentrative nucleoside transporters, hENT1 and hCNT3, in human kidney. Am J Physiol Renal Physiol. 2007; 293: F200-F211., Elwi AN, Damaraju VL, Kuzma ML, Baldwin SA, Young JD, Sawyer MB, Cass CE. Human concentrative nucleoside transporter 3 is a determinant of fludarabine transportability and cytotoxicity in human renal proximal tubule cell cultures. Cancer Chemother Pharmacol. 2009; 63: 289-301., Elwi AN, Damaraju VL, Kuzma ML, Mowles DA, Baldwin SA, Young JD, Sawyer MB, Cass CE. Transepithelial fluxes of adenosine and 2'-deoxyadenosine across human renal proximal tubule cells: roles of nucleoside transporters hENT1, hENT2, and hCNT3. Am J Physiol Renal Physiol. 2009; 296: F1439-F1451. |
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