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Masatoshi Tomi, Ken-ichi Hosoya, Hitomi Takanaga, Sumio Ohtsuki, Tetsuya Terasaki; Induction of xCT Gene Expression and L-Cystine Transport Activity by Diethyl Maleate at the Inner Blood–Retinal Barrier. Invest. Ophthalmol. Vis. Sci. 2002;43(3):774-779.
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purpose. In this study, the expression and regulation of the
L-cystine transporter, system xc −,
at the inner blood–retinal barrier (inner BRB) was investigated using
a conditionally immortalized rat retinal capillary endothelial cell
line (TR-iBRB2) as an in vitro model.
methods. For the uptake study, TR-iBRB2 cells were cultured at 33°C in the
presence or absence of diethyl maleate (DEM), and the uptake rate of[ 14C]L-cystine was measured at 37°C. The
mRNA levels of system xc −, which consists of
xCT and 4F2hc, were determined by quantitative real-time
RT-PCR analysis with specific primers.
results. The xCT and 4F2hc mRNAs were expressed in TR-iBRB2 cells. The[ 14C]L-cystine uptake by TR-iBRB2 cells
appeared to be mediated through a saturable Na+-independent
process. The corresponding Michaelis-Menten constant was 9.18 μM. At
100 μM DEM, the xCT mRNA level and L-cystine uptake
activity in TR-iBRB2 cells were enhanced in a time-dependent manner.
Concomitantly, the glutathione concentration in TR-iBRB2 cells was
increased. In contrast, the 4F2hc mRNA level was unchanged up to 24
hours and was induced for more than 24 hours by DEM treatment. Under
both normal and DEM treatment conditions, the uptake of[ 14C]L-cystine was strongly inhibited by
L-glutamic acid, L-α-aminoadipic acid,
L-homocysteic acid, and L-quisqualic acid,
whereas L-aspartic acid and L-arginine had no
effect, which is evidence of the induction of system
conclusions. System xc −-mediated L-cystine
uptake appears to be present at the inner BRB. DEM induces
L-cystine transport through system
xc − at the inner BRB by enhanced transcription
of the xCT gene.
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