Vitamin D-sensitive and quinacrine-sensitive zinc transport in human intestinal cell line Caco-2.
We studied the mechanism of transepithelial zinc (Zn) transport using monolayers of Caco-2 cells grown on permeable filter supports. 65Zn transport could be fitted to a modified Michaelis-Menten equation, which includes a nonsaturable [linear diffusion constant of nonsaturable component (Kd) = 0.08%.cm-2.90 min-1] and a saturable component [upper well Zn concentration at half Jmax (Kt) = 226 microM and maximal rate of saturable Zn transport (Jmax) = 1.06 nmol.cm-2.90 min-1]. Caco-2 cells contained metal-inducible metallothionein (MT) protein and mRNA as well as mRNA for cysteine-rich intestinal protein (CRIP). Cells pretreated with 10 nM 1 alpha,25-dihydroxyvitamin D3 [1 alpha,25(OH)2D3] for 3 days transported more Zn (159%) than controls (0.48 +/- 0.02 nmol.cm-2.90 min-1) when each was incubated with 100 microM Zn for 90 min. This effect was significant after 24 h of 1 alpha,25-(OH)2D3 pretreatment and continued to increase up to 72 h, with concomitant increases in MT mRNA levels being observed (4-fold by 24 h, 10-fold by 72 h). MT protein levels were only modestly elevated by 72 h 1 alpha,25(OH)2D3 treatment (from 0.32 +/- 0.04 to 0.45 +/- 0.03 nmol MT/mg protein). CRIP mRNA levels were reduced by 1 alpha,25(OH)2D3 treatment. The lysosome-disrupting agent quinacrine (0.5 mM) inhibited basal Zn transport by 68%, suggesting the possible presence of a lysosome-mediated component for transepithelial Zn transport in Caco-2 cells. 1 alpha,25(OH)2D3-stimulated Zn transport was not affected by quinacrine, suggesting that 1 alpha,25(OH)2D3-induced Zn transport is distinct from the putative lysosome-mediated Zn transport pathway.
Fleet
J C
JC
Mineral Bioavailability Laboratory, US Department of Agriculture Human Nutrition Research Center on Aging, Tufts University, Boston, Massachusetts 02111.
Turnbull
A J
AJ
Bourcier
M
M
Wood
R J
RJ
D001692:Biological Transport / Q000187:drug effects; D002117:Calcitriol / Q000494:pharmacology; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007422:Intestines / Q000166:cytology; D060588:LIM Domain Proteins; D008668:Metallothionein / Q000235:genetics / Q000378:metabolism; D011506:Proteins; D011796:Quinacrine / Q000494:pharmacology*; D012333:RNA, Messenger / Q000378:metabolism; D014807:Vitamin D / Q000494:pharmacology*; D015032:Zinc / Q000493:pharmacokinetics*
C486181:CRIP1 protein, human; D002352:Carrier Proteins; D060588:LIM Domain Proteins; D011506:Proteins; D012333:RNA, Messenger; D014807:Vitamin D; D008668:Metallothionein; D002117:Calcitriol; D011796:Quinacrine; D015032:Zinc