In:
American Journal of Physiology-Renal Physiology, American Physiological Society, Vol. 293, No. 3 ( 2007-09), p. F705-F712
Abstract:
Chronic exposure to Cd 2+ causes renal proximal tubular (PT) damage. Cd 2+ reaches the PT mainly as cadmium-metallothionein 1 (CdMT-1) complexes that are filtered at the glomerulus and then internalized in part via endocytosis mediated by megalin and cubulin. Subsequently, Cd 2+ is thought to be released in the cytosol to activate cell death pathways. The proton-coupled divalent metal transporter DMT1 also transports Cd 2+ and is expressed exclusively in endosomes/lysosomes in rat PT cells. Using vector-based RNA interference with short-hairpin small-interfering RNAs (shRNAs) to downregulate DMT1 in the rat renal PT cell line WKPT-0293 Cl.2, we tested the hypothesis that endosomal/lysosomal DMT1 is involved in CdMT-1 nephrotoxicity. One out of 5 shRNAs tested (sh3) significantly reduced expression of DMT1 protein detected by immunoblotting and DMT1 mRNA as determined by RT-PCR by 45.1 ± 9.6 and 36.9 ± 14.4% ( n = 5–6), respectively. Similarly, sh3 reduced perinuclear DMT1 immunostaining in transfected cells. Consistent with the assumed role of DMT1 in CdMT-1 cytotoxicity, sh3, but not the empty vector or sh5, significantly attenuated cell death induced by a 24-h exposure to 14.3 μM CdMT-1 by 35.6 ± 4.2% ( n = 13). In contrast, neither fluorescently labeled metallothionein-1 (MT-1) uptake nor free Cd 2+ toxicity was altered by the effective DMT1 shRNA (sh3), indicating that cellular uptake of metal-MT-1 complexes and Cd 2+ -induced cell death signaling are not affected by DMT1 knockdown. Thus we conclude that endosomal/lysosomal DMT1 plays a role in renal PT CdMT-1 toxicity.
Type of Medium:
Online Resource
ISSN:
1931-857X
,
1522-1466
DOI:
10.1152/ajprenal.00198.2007
Language:
English
Publisher:
American Physiological Society
Publication Date:
2007
detail.hit.zdb_id:
1477287-5
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