Expression and regulation of L-cystine transporter, system X c -, in the newly developed rat retinal Müller cell line (TR-MUL)

Masatoshi Tomi, Takeshi Funaki, Hayato Abukawa, Kazunori Katayama, Tetsu Kondo, Sumio Ohtsuki, Masatsugu Ueda, Masuo Obinata, Tetsuya Terasaki, Ken Ichi Hosoya

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Abstract

The purpose of the present study was to elucidate the expression and regulation of the L-cystine transporter, system xc -, in Müller cells. In this study, newly developed conditionally immortalized rat Müller cell lines (TR-MUL) from transgenic rats harboring the temperature-sensitive SV 40 large T-antigen gene were used as an in vitro model. TR-MUL cells express large T-antigen and grow well at 33°C with a doubling time of 30 h, but do not grow at 39°C. TR-MUL cells express typical Müller cell markers such as S-100, glutamine synthetase, and EAAT1/GLAST, whereas EAAT2/GLT-1 and EAAT5 are not detected. TR-MUL cells also exhibit little or no expression of glial fibrillary acidic protein. We found that TR-MUL5 cells exhibited [14C]L-cystine uptake activity and expressed xCT and 4F2hc, which involve system xc -. The uptake of [14C]L-cystine was significantly inhibited by L-glutamic acid and L-aspartic acid, whereas L-leucine had no effect. Following diethyl maleate (DEM) treatment, the glutathione concentration in TR-MUL5 cells was reduced in the first 24 h, then gradually recovered for more than 24 h. The L-cystine uptake rate and the xCT expression level in TR-MUL5 cells were enhanced by DEM treatment. In contrast, the 4F2hc expression level was unchanged. In conclusion, TR-MUL cells have the properties of Müller cells and exhibit system x c --mediated L-cystine uptake activity. The oxidative stress conditions following DEM treatment activate L-cystine transport in TR-MUL cells due to the enhanced transcription of the xCT gene.

Original languageEnglish
Pages (from-to)208-217
Number of pages10
JournalGLIA
Volume43
Issue number3
DOIs
Publication statusPublished - 2003 Sep 1
Externally publishedYes

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Cystine
diethyl maleate
Cell Line
Viral Tumor Antigens
Transgenic Rats
Glutamate-Ammonia Ligase
Glial Fibrillary Acidic Protein
Aspartic Acid
Leucine
Genes
Glutathione
Glutamic Acid
Oxidative Stress

Keywords

  • Conditionally immortalized cell line
  • Diethyl maleate
  • Glutathione
  • Oxidative stress

ASJC Scopus subject areas

  • Immunology

Cite this

Expression and regulation of L-cystine transporter, system X c -, in the newly developed rat retinal Müller cell line (TR-MUL). / Tomi, Masatoshi; Funaki, Takeshi; Abukawa, Hayato; Katayama, Kazunori; Kondo, Tetsu; Ohtsuki, Sumio; Ueda, Masatsugu; Obinata, Masuo; Terasaki, Tetsuya; Hosoya, Ken Ichi.

In: GLIA, Vol. 43, No. 3, 01.09.2003, p. 208-217.

Research output: Contribution to journalArticle

Tomi, M, Funaki, T, Abukawa, H, Katayama, K, Kondo, T, Ohtsuki, S, Ueda, M, Obinata, M, Terasaki, T & Hosoya, KI 2003, 'Expression and regulation of L-cystine transporter, system X c -, in the newly developed rat retinal Müller cell line (TR-MUL)', GLIA, vol. 43, no. 3, pp. 208-217. https://doi.org/10.1002/glia.10253
Tomi, Masatoshi ; Funaki, Takeshi ; Abukawa, Hayato ; Katayama, Kazunori ; Kondo, Tetsu ; Ohtsuki, Sumio ; Ueda, Masatsugu ; Obinata, Masuo ; Terasaki, Tetsuya ; Hosoya, Ken Ichi. / Expression and regulation of L-cystine transporter, system X c -, in the newly developed rat retinal Müller cell line (TR-MUL). In: GLIA. 2003 ; Vol. 43, No. 3. pp. 208-217.
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AU - Funaki, Takeshi

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AU - Katayama, Kazunori

AU - Kondo, Tetsu

AU - Ohtsuki, Sumio

AU - Ueda, Masatsugu

AU - Obinata, Masuo

AU - Terasaki, Tetsuya

AU - Hosoya, Ken Ichi

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N2 - The purpose of the present study was to elucidate the expression and regulation of the L-cystine transporter, system xc -, in Müller cells. In this study, newly developed conditionally immortalized rat Müller cell lines (TR-MUL) from transgenic rats harboring the temperature-sensitive SV 40 large T-antigen gene were used as an in vitro model. TR-MUL cells express large T-antigen and grow well at 33°C with a doubling time of 30 h, but do not grow at 39°C. TR-MUL cells express typical Müller cell markers such as S-100, glutamine synthetase, and EAAT1/GLAST, whereas EAAT2/GLT-1 and EAAT5 are not detected. TR-MUL cells also exhibit little or no expression of glial fibrillary acidic protein. We found that TR-MUL5 cells exhibited [14C]L-cystine uptake activity and expressed xCT and 4F2hc, which involve system xc -. The uptake of [14C]L-cystine was significantly inhibited by L-glutamic acid and L-aspartic acid, whereas L-leucine had no effect. Following diethyl maleate (DEM) treatment, the glutathione concentration in TR-MUL5 cells was reduced in the first 24 h, then gradually recovered for more than 24 h. The L-cystine uptake rate and the xCT expression level in TR-MUL5 cells were enhanced by DEM treatment. In contrast, the 4F2hc expression level was unchanged. In conclusion, TR-MUL cells have the properties of Müller cells and exhibit system x c --mediated L-cystine uptake activity. The oxidative stress conditions following DEM treatment activate L-cystine transport in TR-MUL cells due to the enhanced transcription of the xCT gene.

AB - The purpose of the present study was to elucidate the expression and regulation of the L-cystine transporter, system xc -, in Müller cells. In this study, newly developed conditionally immortalized rat Müller cell lines (TR-MUL) from transgenic rats harboring the temperature-sensitive SV 40 large T-antigen gene were used as an in vitro model. TR-MUL cells express large T-antigen and grow well at 33°C with a doubling time of 30 h, but do not grow at 39°C. TR-MUL cells express typical Müller cell markers such as S-100, glutamine synthetase, and EAAT1/GLAST, whereas EAAT2/GLT-1 and EAAT5 are not detected. TR-MUL cells also exhibit little or no expression of glial fibrillary acidic protein. We found that TR-MUL5 cells exhibited [14C]L-cystine uptake activity and expressed xCT and 4F2hc, which involve system xc -. The uptake of [14C]L-cystine was significantly inhibited by L-glutamic acid and L-aspartic acid, whereas L-leucine had no effect. Following diethyl maleate (DEM) treatment, the glutathione concentration in TR-MUL5 cells was reduced in the first 24 h, then gradually recovered for more than 24 h. The L-cystine uptake rate and the xCT expression level in TR-MUL5 cells were enhanced by DEM treatment. In contrast, the 4F2hc expression level was unchanged. In conclusion, TR-MUL cells have the properties of Müller cells and exhibit system x c --mediated L-cystine uptake activity. The oxidative stress conditions following DEM treatment activate L-cystine transport in TR-MUL cells due to the enhanced transcription of the xCT gene.

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KW - Diethyl maleate

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KW - Oxidative stress

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