Dehydroascorbic acid uptake and intracellular ascorbic acid accumulation in cultured Müller glial cells (TR-MUL)

Ken ichi Hosoya, Genki Nakamura, Shin ichi Akanuma, Masatoshi Tomi, Masanori Tachikawa

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

Vitamin C is mainly transported across the inner blood-retinal barrier (inner BRB) as dehydroascorbic acid (DHA) via a facilitative glucose transporter (GLUT) 1, and accumulates as ascorbic acid (AA) in the retina. Müller cells, huge glial cells, exhibit passive structural and metabolic functions for retinal neurons and the inner BRB. We characterized DHA transport and its corresponding transporter in a rat Müller cell line (TR-MUL5 cells). [14C]DHA uptake by TR-MUL5 cells took place in a time-dependent and Na+-independent manner. [14C]DHA uptake was inhibited by substrates and inhibitors of GLUTs, suggesting that Müller cells take up DHA via GLUTs. HPLC analysis revealed that most of the DHA taken up by TR-MUL5 cells was converted to AA and accumulated as AA in TR-MUL5 cells. [14C]DHA uptake by TR-MUL5 cells took place in a concentration-dependent manner with a Michaelis-Menten constant of 198 μM and was inhibited by cytochalasin B in a concentration-dependent manner with a 50% inhibition concentration of 0.283 μM. Although GLUT1, 3, and 4 mRNA are expressed in TR-MUL5 cells, quantitative real-time PCR revealed that GLUT1 mRNA expression was 5.85- and 116-fold greater than that of GLUT3 and 4, respectively. Western blot analysis supports the expression of GLUT1 protein with 45 kDa in TR-MUL5 cells. In conclusion, DHA is taken up by facilitative glucose transporters, most likely GLUT1, and converted to AA in TR-MUL5 cells.

Original languageEnglish
Pages (from-to)1351-1357
Number of pages7
JournalNeurochemistry International
Volume52
Issue number7
DOIs
Publication statusPublished - 2008 Jun
Externally publishedYes

Fingerprint

Dehydroascorbic Acid
Neuroglia
Ascorbic Acid
Blood-Retinal Barrier
Facilitative Glucose Transport Proteins
Glucose Transporter Type 1
Retinal Neurons
Messenger RNA
Cytochalasin B
Retina
Real-Time Polymerase Chain Reaction
Western Blotting
High Pressure Liquid Chromatography

Keywords

  • Dehydroascorbic acid
  • GLUT1
  • Inner blood-retinal barrier
  • Müller cells
  • Vitamin C

ASJC Scopus subject areas

  • Cell Biology
  • Molecular Biology
  • Cellular and Molecular Neuroscience

Cite this

Dehydroascorbic acid uptake and intracellular ascorbic acid accumulation in cultured Müller glial cells (TR-MUL). / Hosoya, Ken ichi; Nakamura, Genki; Akanuma, Shin ichi; Tomi, Masatoshi; Tachikawa, Masanori.

In: Neurochemistry International, Vol. 52, No. 7, 06.2008, p. 1351-1357.

Research output: Contribution to journalArticle

Hosoya, Ken ichi ; Nakamura, Genki ; Akanuma, Shin ichi ; Tomi, Masatoshi ; Tachikawa, Masanori. / Dehydroascorbic acid uptake and intracellular ascorbic acid accumulation in cultured Müller glial cells (TR-MUL). In: Neurochemistry International. 2008 ; Vol. 52, No. 7. pp. 1351-1357.
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AB - Vitamin C is mainly transported across the inner blood-retinal barrier (inner BRB) as dehydroascorbic acid (DHA) via a facilitative glucose transporter (GLUT) 1, and accumulates as ascorbic acid (AA) in the retina. Müller cells, huge glial cells, exhibit passive structural and metabolic functions for retinal neurons and the inner BRB. We characterized DHA transport and its corresponding transporter in a rat Müller cell line (TR-MUL5 cells). [14C]DHA uptake by TR-MUL5 cells took place in a time-dependent and Na+-independent manner. [14C]DHA uptake was inhibited by substrates and inhibitors of GLUTs, suggesting that Müller cells take up DHA via GLUTs. HPLC analysis revealed that most of the DHA taken up by TR-MUL5 cells was converted to AA and accumulated as AA in TR-MUL5 cells. [14C]DHA uptake by TR-MUL5 cells took place in a concentration-dependent manner with a Michaelis-Menten constant of 198 μM and was inhibited by cytochalasin B in a concentration-dependent manner with a 50% inhibition concentration of 0.283 μM. Although GLUT1, 3, and 4 mRNA are expressed in TR-MUL5 cells, quantitative real-time PCR revealed that GLUT1 mRNA expression was 5.85- and 116-fold greater than that of GLUT3 and 4, respectively. Western blot analysis supports the expression of GLUT1 protein with 45 kDa in TR-MUL5 cells. In conclusion, DHA is taken up by facilitative glucose transporters, most likely GLUT1, and converted to AA in TR-MUL5 cells.

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