Polyol formation and NADPH-dependent reductases in dog retinal capillary pericytes and endothelial cells

Sanai Sato, E. Filippo Secchi, Martin J. Lizak, Shigeru Fukase, Nobuo Ohta, Masatoshi Murata, Jen Yue Tsai, Peter F Kador

Research output: Contribution to journalArticle

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Abstract

PURPOSE. Dogs fed a diet containing 30% galacrose experience retinal vascular changes similar to those in human diabetic retinopathy, with selective pericyte loss as an initial lesion. In the present study the relationship among reduced nicotinamide adenine dinucleotide phosphate (NADPH)-dependent reductases, polyol formation, and flux through the polyol pathway in cultured dog retinal capillary cells were investigated. METHODS. Pericytes and endothelial cells were cultured from retina of beagle dogs. NADPH-dependent reductases were characterized by chromatofocusing after gel filtration. Sugars in cultured cells were analyzed by gas chromatography, and flux through the polyol pathway was investigated by 19F nuclear magnetic resonance (NMR) with 3-fluoro-3-deoxy-D-glucose (3FG) as a substrate. The presence of aldose reductase and sorbitol dehydrogenase in these cells was examined by northern blot analysis. RESULTS. Two distinct peaks corresponding to aldose reductase and aldehyde reductase, the latter being dominant, were observed in pericytes by chromatofocusing. Culture in medium containing either 10 mM D-galactose or 30 mM D-glucose resulted in the accumulation of sugar alcohol in pencytes that was markedly reduced by aldose reductase inhibitors. 19F NMR spectra obtained from pencytes cultured for 5 days in medium containing 2 mM 3FG displayed the marked accumulation of 3-fluoro- deoxysorbitol but not 3-fluoro-deoxyfructose. No 3FG metabolism was observed in similarly cultured endothelial cells. With northern blot analysis, aldose reductase was detected in pericytes but not in endothelial cells. Sorbitol dehydrogenase was below the detectable limit in pericytes and endothelial cells. CONCLUSIONS. Aldose, aldehyde, and glyceraldehyde reductases are present in dog retinal capillary pericytes, with aldehyde reductase being the major reductase present. Polyol accumulation easily occurs in pericytes but not in endothelial cells.

Original languageEnglish (US)
Pages (from-to)697-704
Number of pages8
JournalInvestigative Ophthalmology and Visual Science
Volume40
Issue number3
StatePublished - Mar 11 1999

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Aldehyde Reductase
Pericytes
NADP
Oxidoreductases
Endothelial Cells
Dogs
L-Iditol 2-Dehydrogenase
Northern Blotting
Cultured Cells
Magnetic Resonance Spectroscopy
Sugar Alcohols
Retinal Vessels
polyol
Diabetic Retinopathy
Galactose
Gas Chromatography
Gel Chromatography
Culture Media
Retina
Diet

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

Cite this

Polyol formation and NADPH-dependent reductases in dog retinal capillary pericytes and endothelial cells. / Sato, Sanai; Secchi, E. Filippo; Lizak, Martin J.; Fukase, Shigeru; Ohta, Nobuo; Murata, Masatoshi; Tsai, Jen Yue; Kador, Peter F.

In: Investigative Ophthalmology and Visual Science, Vol. 40, No. 3, 11.03.1999, p. 697-704.

Research output: Contribution to journalArticle

Sato, S, Secchi, EF, Lizak, MJ, Fukase, S, Ohta, N, Murata, M, Tsai, JY & Kador, PF 1999, 'Polyol formation and NADPH-dependent reductases in dog retinal capillary pericytes and endothelial cells', Investigative Ophthalmology and Visual Science, vol. 40, no. 3, pp. 697-704.
Sato, Sanai ; Secchi, E. Filippo ; Lizak, Martin J. ; Fukase, Shigeru ; Ohta, Nobuo ; Murata, Masatoshi ; Tsai, Jen Yue ; Kador, Peter F. / Polyol formation and NADPH-dependent reductases in dog retinal capillary pericytes and endothelial cells. In: Investigative Ophthalmology and Visual Science. 1999 ; Vol. 40, No. 3. pp. 697-704.
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abstract = "PURPOSE. Dogs fed a diet containing 30{\%} galacrose experience retinal vascular changes similar to those in human diabetic retinopathy, with selective pericyte loss as an initial lesion. In the present study the relationship among reduced nicotinamide adenine dinucleotide phosphate (NADPH)-dependent reductases, polyol formation, and flux through the polyol pathway in cultured dog retinal capillary cells were investigated. METHODS. Pericytes and endothelial cells were cultured from retina of beagle dogs. NADPH-dependent reductases were characterized by chromatofocusing after gel filtration. Sugars in cultured cells were analyzed by gas chromatography, and flux through the polyol pathway was investigated by 19F nuclear magnetic resonance (NMR) with 3-fluoro-3-deoxy-D-glucose (3FG) as a substrate. The presence of aldose reductase and sorbitol dehydrogenase in these cells was examined by northern blot analysis. RESULTS. Two distinct peaks corresponding to aldose reductase and aldehyde reductase, the latter being dominant, were observed in pericytes by chromatofocusing. Culture in medium containing either 10 mM D-galactose or 30 mM D-glucose resulted in the accumulation of sugar alcohol in pencytes that was markedly reduced by aldose reductase inhibitors. 19F NMR spectra obtained from pencytes cultured for 5 days in medium containing 2 mM 3FG displayed the marked accumulation of 3-fluoro- deoxysorbitol but not 3-fluoro-deoxyfructose. No 3FG metabolism was observed in similarly cultured endothelial cells. With northern blot analysis, aldose reductase was detected in pericytes but not in endothelial cells. Sorbitol dehydrogenase was below the detectable limit in pericytes and endothelial cells. CONCLUSIONS. Aldose, aldehyde, and glyceraldehyde reductases are present in dog retinal capillary pericytes, with aldehyde reductase being the major reductase present. Polyol accumulation easily occurs in pericytes but not in endothelial cells.",
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T1 - Polyol formation and NADPH-dependent reductases in dog retinal capillary pericytes and endothelial cells

AU - Sato, Sanai

AU - Secchi, E. Filippo

AU - Lizak, Martin J.

AU - Fukase, Shigeru

AU - Ohta, Nobuo

AU - Murata, Masatoshi

AU - Tsai, Jen Yue

AU - Kador, Peter F

PY - 1999/3/11

Y1 - 1999/3/11

N2 - PURPOSE. Dogs fed a diet containing 30% galacrose experience retinal vascular changes similar to those in human diabetic retinopathy, with selective pericyte loss as an initial lesion. In the present study the relationship among reduced nicotinamide adenine dinucleotide phosphate (NADPH)-dependent reductases, polyol formation, and flux through the polyol pathway in cultured dog retinal capillary cells were investigated. METHODS. Pericytes and endothelial cells were cultured from retina of beagle dogs. NADPH-dependent reductases were characterized by chromatofocusing after gel filtration. Sugars in cultured cells were analyzed by gas chromatography, and flux through the polyol pathway was investigated by 19F nuclear magnetic resonance (NMR) with 3-fluoro-3-deoxy-D-glucose (3FG) as a substrate. The presence of aldose reductase and sorbitol dehydrogenase in these cells was examined by northern blot analysis. RESULTS. Two distinct peaks corresponding to aldose reductase and aldehyde reductase, the latter being dominant, were observed in pericytes by chromatofocusing. Culture in medium containing either 10 mM D-galactose or 30 mM D-glucose resulted in the accumulation of sugar alcohol in pencytes that was markedly reduced by aldose reductase inhibitors. 19F NMR spectra obtained from pencytes cultured for 5 days in medium containing 2 mM 3FG displayed the marked accumulation of 3-fluoro- deoxysorbitol but not 3-fluoro-deoxyfructose. No 3FG metabolism was observed in similarly cultured endothelial cells. With northern blot analysis, aldose reductase was detected in pericytes but not in endothelial cells. Sorbitol dehydrogenase was below the detectable limit in pericytes and endothelial cells. CONCLUSIONS. Aldose, aldehyde, and glyceraldehyde reductases are present in dog retinal capillary pericytes, with aldehyde reductase being the major reductase present. Polyol accumulation easily occurs in pericytes but not in endothelial cells.

AB - PURPOSE. Dogs fed a diet containing 30% galacrose experience retinal vascular changes similar to those in human diabetic retinopathy, with selective pericyte loss as an initial lesion. In the present study the relationship among reduced nicotinamide adenine dinucleotide phosphate (NADPH)-dependent reductases, polyol formation, and flux through the polyol pathway in cultured dog retinal capillary cells were investigated. METHODS. Pericytes and endothelial cells were cultured from retina of beagle dogs. NADPH-dependent reductases were characterized by chromatofocusing after gel filtration. Sugars in cultured cells were analyzed by gas chromatography, and flux through the polyol pathway was investigated by 19F nuclear magnetic resonance (NMR) with 3-fluoro-3-deoxy-D-glucose (3FG) as a substrate. The presence of aldose reductase and sorbitol dehydrogenase in these cells was examined by northern blot analysis. RESULTS. Two distinct peaks corresponding to aldose reductase and aldehyde reductase, the latter being dominant, were observed in pericytes by chromatofocusing. Culture in medium containing either 10 mM D-galactose or 30 mM D-glucose resulted in the accumulation of sugar alcohol in pencytes that was markedly reduced by aldose reductase inhibitors. 19F NMR spectra obtained from pencytes cultured for 5 days in medium containing 2 mM 3FG displayed the marked accumulation of 3-fluoro- deoxysorbitol but not 3-fluoro-deoxyfructose. No 3FG metabolism was observed in similarly cultured endothelial cells. With northern blot analysis, aldose reductase was detected in pericytes but not in endothelial cells. Sorbitol dehydrogenase was below the detectable limit in pericytes and endothelial cells. CONCLUSIONS. Aldose, aldehyde, and glyceraldehyde reductases are present in dog retinal capillary pericytes, with aldehyde reductase being the major reductase present. Polyol accumulation easily occurs in pericytes but not in endothelial cells.

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