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Zn disrupts Cd‐induced stimulation of apical choline uptake in cultured choroid plexus independent of GSH availability (685.4)
Author(s) -
Villalobos A.,
Young R,
Francis Stuart S
Publication year - 2014
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.28.1_supplement.685.4
Subject(s) - gclm , gclc , glutathione , oxidative stress , metallothionein , chemistry , choroid plexus , stimulation , buthionine sulfoximine , medicine , microbiology and biotechnology , endocrinology , biochemistry , biology , enzyme , gene , central nervous system
In cultured neonatal rat choroid plexus we showed cadmium (Cd) stimulated apical choline uptake and induced oxidative stress and glutathione (GSH) synthesis. Zinc (Zn) supplementation disrupted Cd‐induced oxidative stress and stimulation of choline uptake. Our objectives are to further elucidate GSH regulation by Cd and Zn and determine whether GSH is critical to disruption of Cd‐induced stimulation of choline uptake by Zn. Cultured choroid plexus cells were supplemented (48 h) with 25 μM ZnCl2 in complete medium before 12‐h treatment with 0 or 500 nM CdCl2±10 μM ZnCl2 without serum. By immunoblot analysis and qRT‐PCR, we assessed induction of HO‐1, HSP70, Cu/Zn‐SOD (SOD1), metallothionein‐1 (MT‐1) and both modifier (GCLM) and catalytic subunits (GCLC) of glutamate‐cysteine ligase, the rate‐limiting enzyme in GSH synthesis. Cd induced GCLM, GCLC, MT‐1, HO‐1, and SOD1 mRNA levels by 6‐, 3‐, 16‐, 12‐ and 3‐fold; Zn supplementation attenuated induction. Cd alone increased GSH and GSSG by 2‐ and 30‐fold; Zn supplementation abated increases in each. To determine whether the effects of Zn were dependent on GSH availability, we treated cells with 100 μM buthionine sulfoximine (BSO) to inhibit GCL during Zn supplementation and subsequent 12‐h treatment with 250 nM CdCl2±10 μM ZnCl2. BSO alone or with Zn±Cd reduced GSH by 90% and increased GSSG by 15‐fold above control. In Cd‐treated cells, BSO elicited a compensatory induction of HO‐1, HSP70, SOD1, GCLM, GCLC and MT‐1 and further stimulated apical choline uptake. Zn supplementation attenuated induction of cytoprotective genes and proteins and stimulation of apical choline uptake in cells treated with Cd±BSO. Thus, Zn abates Cd‐induced oxidative stress and GSH regulation, but disrupts stimulation of choline uptake independent of GSH availability. Grant Funding Source : NSF #IOS‐1052654