Transporters of glycine, cystine, glutamate and glutamine in canine lens epithelial cells

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Abstract

Purpose

The aim of this study is to evaluate GSH synthesis-related amino acid (glycine, cystine, glutamate and glutamine) transport activity in a canine lens epithelial cell line (cdLEC) and define the transporter.

Methods

The primary lens epithelial cells of mature cataract dog were transfected with the expression plasmid DNA of large T antigen from replication origin-defective simian virus 40 (SV40), then cloned using glass a cylinder. The transport activities of four amino acids in lens epithelial cells were determined. The cDNA sequences of transporter contributing GSH synthesis were also determined.

Results

Na-dependent glycine and glutamate transport activity was 2.70 + 0.26 and 4.40 + 1.10 nmol/mg protein min, respectively. The cystine transport was Na-independent and the transport activity was 0.626 + 0.17 nmol/mg protein min. The cDNA sequence of the glycine transporter 1(GlyT1), cystine/glutamate exchanger (xCT,), Na-dependent neutral amino acid transporter 2(ASCT2), and glutamate transporter (EAAT) were determined. Western blot analysis confirmed their expression in membrane protein sample of cdLEC.

Conclusion

Lens epithelial cells possess GlyT1, xCT, ASCT2 and EAAT1-5.

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