Antioxidant function of corneal ALDH3A1 in cultured stromal fibroblasts

Lassen, Natalie; Pappa, Aglaia; Black, William J.; Jester, James V.; Day, Brian J.; Min, Elysia; Vasiliou, Vasilis

doi:10.1016/j.freeradbiomed.2006.08.009

Cited by 60 publications

(56 citation statements)

References 53 publications

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“…However, our results showed that the size of the keratocytes maintaining transparency in the birds and rat were smaller than that in the rabbit. Corneal crystallins such as ALDH, which have been shown to protect against UV and oxidative stress induced apoptosis, are expressed abundantly in keratocytes [10,20,23,26,31]. The protein expression of crystallins in keratocytes of the birds and rat may be more abundant than that in the rabbit and not be dependent on the size of the keratocyte.…”

Section: Discussionmentioning

confidence: 99%

Microstructure Characteristics of the Cornea in Birds and Mammals

Tsukahara

Tani

Lee

et al. 2010

The Journal of Veterinary Medical Science

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ABSTRACT. In this study, the microstructure of the cornea was compared among chickens (Gallus gallus), jungle crows (Corvus macrorhynchos), rats (Rattus norvegicus) and rabbits (Oryctolagus cuniculus). The density of keratocytes in the mammals was over 3 times that in the birds. The size of the keratocytes in the birds and rat were significantly lower than those in the rabbit. Using scanning and transmission electron microscopy, the bundles of collagen fibers in the birds were found to be well arranged, while those in the mammals were arranged randomly. The collagen lamellae of the birds were significantly thicker than those of the mammals, and the numbers of collagen lamellae in the birds were significantly smaller than in the mammals. The center-to-center distances between the collagen fibrils of the chicken and rabbit were significantly larger than those of the crow and rat. The densities of collagen fibrils in the chicken and rabbit were significantly less than those of the crow and rat.

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Section: Discussionmentioning

confidence: 99%

Microstructure Characteristics of the Cornea in Birds and Mammals

Tsukahara

Tani

Lee

et al. 2010

The Journal of Veterinary Medical Science

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“…38 These enzymes are thought to play a critical role in the protection of the cornea and lens to UV-induced damage through multiple mechanisms including absorption of UV light, 39 metabolism of toxic aldehydes, 40,41 direct free radical scavenging, 32,42 and production of antioxidant NAD(P)H. 33 The sum of these actions may help block oxidative stressinduced protein modifications leading to enzyme inactivation, partial unfolding, and non-native protein aggregation. Oxidative stress and protein modification are thought to play important roles in the formation of cataract, 43 suggesting that a similar mechanism is involved in the development of corneal haze and increased light scattering after corneal injury and repair.…”

Section: Discussionmentioning

confidence: 99%

“…32 Single-cell light scattering was measured by using previously reported methods. 22 Briefly, the cells were plated onto 25-mm diameter glass coverslips containing a 10-mm diameter, 50-m-thick acrylamide gel/collagen substrate.…”

Section: Aldh Transfection and Measurement Of Single-cell Light Scattmentioning

confidence: 99%

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Myofibroblast Differentiation Modulates Keratocyte Crystallin Protein Expression, Concentration, and Cellular Light Scattering

Jester

Brown

Pappa

et al. 2012

Invest. Ophthalmol. Vis. Sci.

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PURPOSE. The purpose of this study was to determine whether myofibroblast differentiation altered keratocyte crystallin protein concentration and increased cellular light scattering. METHODS. Serum-free cultured rabbit corneal keratocytes and TGF␤ (5 ng/mL) induced myofibroblasts were harvested and counted and protein/RNA extracted. Expression of myofibroblast and keratocyte markers was determined by real-time PCR and Western blot analysis. The cell volume of calcein AMloaded keratocytes and myofibroblasts was determined by using nonlinear optical microscopy. Cellular light scattering of transformed myofibroblasts expressing human keratocyte crystallins was measured by reflectance confocal microscopy. RESULTS. Differentiated myofibroblasts showed a significant decrease in RNA levels for the keratocyte markers ALDH1A1, lumican, and keratocan and a significant increase in the myofibroblast marker ␣-smooth muscle actin. Volumetric and protein measurements showed that myofibroblast differentiation significantly increased cytoplasmic volume (293%; P Ͻ 0.001) and water-soluble and -insoluble protein content per cell (respectively, 442% and 431%; P Ͻ 0.002) compared to keratocytes. Western blot analysis showed that the level of ALDH1A1 protein per cell was similar between myofibroblasts and keratocytes, but was substantially reduced as a percentage of total water-soluble protein. Light scattering measurements showed that induced expression of corneal crystallins significantly decreased light scattering. CONCLUSIONS. These data suggest that myofibroblast differentiation leads to a marked increase in cell volume and dilution of corneal crystallins associated with an increase in cellular light scattering. (Invest Ophthalmol Vis Sci. 2012;53:770 -778) DOI: 10.1167/iovs.11-9092 P revious studies have shown that differentiation of keratocytes into myofibroblasts is regulated in part by transforming growth factor (TGF)-␤.1,2 Under serum-free culture conditions, rabbit, bovine, and human corneal keratocytes maintain a quiescent phenotype, dendritic morphology, and high expression of phenotypic markers, including keratocan, lumican, and the corneal crystallins, aldehyde dehydrogenase isozymes 3A1 and 1A1 (ALDH3A1/1A1) and transketolase (TKT). 3-5Treatment of cultured keratocytes with TGF␤ leads to cell spreading, actin filament assembly, and downregulated expression of keratocyte-specific genes that is coupled with the de novo upregulated expression of ␣-smooth muscle actin (␣-SMA), the phenotypic marker for myofibroblast differentiation. 6 Myofibroblasts play a critical role in corneal wound healing that involves deposition and organization of extracellular matrix leading to wound contraction. [7][8][9] Neutralizing antibodies to TGF␤ have been shown to block the appearance of myofibroblasts in corneal wounds and significantly reduce corneal scarring and the development of corneal haze. 10,11The loss of transparency after corneal injury and scarring has long been associated with the deposition of abnormally arranged and disorg...

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“…Cells (1 × 10 4 ) were plated in 96-well culture plates, allowed to attach overnight, and then treated in triplicate with octanal (0-1mM) in serum-free medium for 24 h. Cell viability was determined by the SRB assay, as described previously [18].…”

Section: Sulforhodamine B Assaymentioning

confidence: 99%

Expression and initial characterization of human ALDH3B1

Marchitti

Orlicky

Vasiliou

2007

Biochemical and Biophysical Research Communications

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Aldehyde dehydrogenases (ALDHs) are critical enzymes in the metabolism of endogenous and exogenous aldehydes. The human genome contains nineteen putatively functional ALDH genes; ALDH3B1 belongs to the ALDH3 family. While recent studies have linked the ALDH3B1 locus to schizophrenia, nothing was known, until now, about the properties and significance of the ALDH3B1 protein. The aim of this study was to characterize the ALDH3B1 protein. Human ALDH3B1 was baculovirus-expressed and found to be catalytically active towards medium-and long-chain aliphatic aldehydes and the aromatic aldehyde benzaldehyde. Western blot analyses indicate that ALDH3B1 is highly expressed in kidney and liver and moderately expressed in various brain regions. ALDH3B1-transfected HEK293 cells were significantly protected against cytotoxicity induced by the lipid-peroxidation product octanal when compared to vector-transfected cells. This study shows for the first time the functionality, expression and protective role of ALDH3B1 and indicates a potential physiological role of ALDH3B1 against oxidative stress.

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Antioxidant function of corneal ALDH3A1 in cultured stromal fibroblasts

Cited by 60 publications

References 53 publications

Microstructure Characteristics of the Cornea in Birds and Mammals

Microstructure Characteristics of the Cornea in Birds and Mammals

Myofibroblast Differentiation Modulates Keratocyte Crystallin Protein Expression, Concentration, and Cellular Light Scattering

Expression and initial characterization of human ALDH3B1

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