Role of aldehyde dehydrogenase isozymes in the defense of rat lens and human lens epithelial cells against oxidative stress

Sanjeev Cboudhary, Tianlin Xiao, Leoncio A. Vergara, Sanjay Srivastava, David Nees, Joram Piatigorsky, Naseem Ansari

Research output: Contribution to journalArticle

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Abstract

PURPOSE. 4-Hydroxynonenal (HNE), a metastable lipid peroxidation product, is highly toxic to various cell types if not detoxified. Because of its constant exposure to light, the ocular lens continuously generates reactive oxygen species which, under conditions of oxidative stress, may lead to excessive lipid peroxidation and consequent formation of lipid-derived aldehydes (LDAs) such as HNE. The contribution of various isozymes of aldehyde dehydrogenase (ALDH) to the oxidation of LDAs has never been systematically investigated in the lens. The present study was undertaken to ascertain the role of ALDH1A1 and -3A1 in HNE metabolism and HNE-induced toxicity in cultured human lens epithelial cells (HLECs) and in rat and mouse lenses. METHODS. The metabolism of 3H-HNE was studied in ALDH3A1-knockout mouse lens and in HLECs transfected with ALDH1A1- or -3A1-specific antisense RNA and short interfering (Si)RNA. Appropriate controls were used, including wild-type mouse lens, scrambled oligonucleotides, and a transfection reagent. Transfected HLECs were exposed to oxidative stress (Fenton reaction) or HNE (30 μM) for 3 hours. Toxicity parameters, such as cell viability, apoptosis, and protein-HNE adducts and oxidation of exogenously added 3H-HNE were measured. Rat lenses were transfected with the SiRNA specific to ALDH1A1, and oxidation of 3H-HNE and the susceptibility of the transfected lenses to oxidation-induced opacification were measured. RESULTS. Rat lenses transfected with ALDH1A1-specific SiRNA, or cultured in the presence of the ALDH inhibitor cyanamide/disulfiram and subjected to oxidative stress displayed accelerated loss of transparency and a diminished capacity to oxidize HNE. Similarly, inhibition of ALDH1A1 in HLECs by ALDH1A1-specific antisense RNA or SiRNA was associated with decreased oxidation of 3H-HNE and increased susceptibility of the cells to oxidative damage, including apoptosis. Furthermore, 3H-HNE metabolism and HNE-induced toxicity were not affected in ALDH3A1-specific SiRNA- or antisense RNA-treated rat lenses, HLECs, or ALDH3A1-null mouse lenses. CONCLUSIONS. The results suggest that, under oxidative stress, HNE produced in the lens epithelium can cause toxicity and thus contribute to oxidation-induced cataractogenesis. Furthermore, the studies indicate that ALDH1A1 is a critical isozyme for maintaining clarity in human, rat, and mouse lenses.

Original languageEnglish (US)
Pages (from-to)259-267
Number of pages9
JournalInvestigative Ophthalmology and Visual Science
Volume46
Issue number1
DOIs
StatePublished - Jan 2005

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Aldehyde Dehydrogenase
Lenses
Isoenzymes
Oxidative Stress
Epithelial Cells
Antisense RNA
Aldehydes
Lipid Peroxidation
Cyanamide
Apoptosis
Lipids
Disulfiram
Crystalline Lens
Poisons

ASJC Scopus subject areas

  • Ophthalmology

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Role of aldehyde dehydrogenase isozymes in the defense of rat lens and human lens epithelial cells against oxidative stress. / Cboudhary, Sanjeev; Xiao, Tianlin; Vergara, Leoncio A.; Srivastava, Sanjay; Nees, David; Piatigorsky, Joram; Ansari, Naseem.

In: Investigative Ophthalmology and Visual Science, Vol. 46, No. 1, 01.2005, p. 259-267.

Research output: Contribution to journalArticle

Cboudhary, Sanjeev ; Xiao, Tianlin ; Vergara, Leoncio A. ; Srivastava, Sanjay ; Nees, David ; Piatigorsky, Joram ; Ansari, Naseem. / Role of aldehyde dehydrogenase isozymes in the defense of rat lens and human lens epithelial cells against oxidative stress. In: Investigative Ophthalmology and Visual Science. 2005 ; Vol. 46, No. 1. pp. 259-267.
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AU - Xiao, Tianlin

AU - Vergara, Leoncio A.

AU - Srivastava, Sanjay

AU - Nees, David

AU - Piatigorsky, Joram

AU - Ansari, Naseem

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N2 - PURPOSE. 4-Hydroxynonenal (HNE), a metastable lipid peroxidation product, is highly toxic to various cell types if not detoxified. Because of its constant exposure to light, the ocular lens continuously generates reactive oxygen species which, under conditions of oxidative stress, may lead to excessive lipid peroxidation and consequent formation of lipid-derived aldehydes (LDAs) such as HNE. The contribution of various isozymes of aldehyde dehydrogenase (ALDH) to the oxidation of LDAs has never been systematically investigated in the lens. The present study was undertaken to ascertain the role of ALDH1A1 and -3A1 in HNE metabolism and HNE-induced toxicity in cultured human lens epithelial cells (HLECs) and in rat and mouse lenses. METHODS. The metabolism of 3H-HNE was studied in ALDH3A1-knockout mouse lens and in HLECs transfected with ALDH1A1- or -3A1-specific antisense RNA and short interfering (Si)RNA. Appropriate controls were used, including wild-type mouse lens, scrambled oligonucleotides, and a transfection reagent. Transfected HLECs were exposed to oxidative stress (Fenton reaction) or HNE (30 μM) for 3 hours. Toxicity parameters, such as cell viability, apoptosis, and protein-HNE adducts and oxidation of exogenously added 3H-HNE were measured. Rat lenses were transfected with the SiRNA specific to ALDH1A1, and oxidation of 3H-HNE and the susceptibility of the transfected lenses to oxidation-induced opacification were measured. RESULTS. Rat lenses transfected with ALDH1A1-specific SiRNA, or cultured in the presence of the ALDH inhibitor cyanamide/disulfiram and subjected to oxidative stress displayed accelerated loss of transparency and a diminished capacity to oxidize HNE. Similarly, inhibition of ALDH1A1 in HLECs by ALDH1A1-specific antisense RNA or SiRNA was associated with decreased oxidation of 3H-HNE and increased susceptibility of the cells to oxidative damage, including apoptosis. Furthermore, 3H-HNE metabolism and HNE-induced toxicity were not affected in ALDH3A1-specific SiRNA- or antisense RNA-treated rat lenses, HLECs, or ALDH3A1-null mouse lenses. CONCLUSIONS. The results suggest that, under oxidative stress, HNE produced in the lens epithelium can cause toxicity and thus contribute to oxidation-induced cataractogenesis. Furthermore, the studies indicate that ALDH1A1 is a critical isozyme for maintaining clarity in human, rat, and mouse lenses.

AB - PURPOSE. 4-Hydroxynonenal (HNE), a metastable lipid peroxidation product, is highly toxic to various cell types if not detoxified. Because of its constant exposure to light, the ocular lens continuously generates reactive oxygen species which, under conditions of oxidative stress, may lead to excessive lipid peroxidation and consequent formation of lipid-derived aldehydes (LDAs) such as HNE. The contribution of various isozymes of aldehyde dehydrogenase (ALDH) to the oxidation of LDAs has never been systematically investigated in the lens. The present study was undertaken to ascertain the role of ALDH1A1 and -3A1 in HNE metabolism and HNE-induced toxicity in cultured human lens epithelial cells (HLECs) and in rat and mouse lenses. METHODS. The metabolism of 3H-HNE was studied in ALDH3A1-knockout mouse lens and in HLECs transfected with ALDH1A1- or -3A1-specific antisense RNA and short interfering (Si)RNA. Appropriate controls were used, including wild-type mouse lens, scrambled oligonucleotides, and a transfection reagent. Transfected HLECs were exposed to oxidative stress (Fenton reaction) or HNE (30 μM) for 3 hours. Toxicity parameters, such as cell viability, apoptosis, and protein-HNE adducts and oxidation of exogenously added 3H-HNE were measured. Rat lenses were transfected with the SiRNA specific to ALDH1A1, and oxidation of 3H-HNE and the susceptibility of the transfected lenses to oxidation-induced opacification were measured. RESULTS. Rat lenses transfected with ALDH1A1-specific SiRNA, or cultured in the presence of the ALDH inhibitor cyanamide/disulfiram and subjected to oxidative stress displayed accelerated loss of transparency and a diminished capacity to oxidize HNE. Similarly, inhibition of ALDH1A1 in HLECs by ALDH1A1-specific antisense RNA or SiRNA was associated with decreased oxidation of 3H-HNE and increased susceptibility of the cells to oxidative damage, including apoptosis. Furthermore, 3H-HNE metabolism and HNE-induced toxicity were not affected in ALDH3A1-specific SiRNA- or antisense RNA-treated rat lenses, HLECs, or ALDH3A1-null mouse lenses. CONCLUSIONS. The results suggest that, under oxidative stress, HNE produced in the lens epithelium can cause toxicity and thus contribute to oxidation-induced cataractogenesis. Furthermore, the studies indicate that ALDH1A1 is a critical isozyme for maintaining clarity in human, rat, and mouse lenses.

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