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Yoshiyuki Kubo, Akiko Obata, Shin-ichi Akanuma, Ken-ichi Hosoya; Impact of Cationic Amino Acid Transporter 1 on Blood-Retinal Barrier Transport of L-Ornithine. Invest. Ophthalmol. Vis. Sci. 2015;56(10):5925-5932. doi: 10.1167/iovs.15-17398.
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To elucidate L-ornithine transport at the blood-retinal barrier (BRB).
Integration plot and retinal uptake index (RUI) were used to investigate the in vivo [3H]L-ornithine transport across the BRB. In vitro transport studies of [3H]L-ornithine were performed with TR-iBRB2 cells and RPE-J cells, the model cells of the inner and outer BRB, respectively. Immunohistochemistry was performed on cationic amino acid transporter 1 (CAT1/SLC7A1).
The apparent influx permeability clearance of [3H]L-ornithine was found to be 18. 7 μL/(min·g retina), and the RUI of [3H]L-ornithine was reduced by L-ornithine and L-arginine, suggesting the blood-to-retina transport of L-ornithine at the BRB. [3H]L-Ornithine uptake by TR-iBRB2 cells showed a time-, temperature- and concentration-dependence with a Michaelis-Menten constant (Km) of 33.2 μM and a nonsaturable uptake rate (Kd) of 2.18 μL/(min·mg protein). The uptake was Na+-independent, and was inhibited by L-ornithine, L-arginine, and L-lysine, suggesting the involvement of CAT1 in L-ornithine transport at the inner BRB. Immunohistochemistry revealed the luminal and abluminal localization of CAT1 at the inner BRB, and at the basal localization at the outer BRB. Retinal pigment epithelium–J cells showed that the basal-to-cell (B-to-C) uptake of [3H]L-ornithine was greater than that of the apical-to-cell (A-to-C) uptake, and the B-to-C transport was inhibited by unlabeled L-ornithine, suggesting the involvement of CAT1 in the blood-to-cell transport of L-ornithine across the basal membrane at the outer BRB.
These suggest the involvement of CAT1 in L-ornithine transport at the luminal and abluminal sides of the inner BRB and the basal side of the outer BRB.
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