Constitutive Expression of Class 3 Aldehyde Dehydrogenase in Cultured Rat Corneal Epithelium

Abstract

Mammalian Class 3 aldehyde dehydrogenase (ALDH) is normally associated with neoplastic transformation or xenobiotic induction by aromatic hydrocarbons in liver. However, Class 3 ALDH is constitutively expressed at it's highest specific activity in corneal epithelium. Tissue-specific, differential gene expression is often controlled by alternative, independent molecular pathways. We report here the development of an in vitro corneal epithelium culture system that retains constitutive high expression of the ALDH3 gene. This model system was used to establish, by enzymatic assays, Western and Northern analyses, histochemical and immunocytochemical staining, and 5'3' RACE methodologies that constitutive and xenobiotic induction of Class 3 ALDHs occurs from a single gene. Our results also provide a plausible explanation for the very high Class 3 ALDH activity in mammalian cornea, as the primary mechanism of oxidation of lipid peroxidation-derived aldehydes. Further studies with corneal epithelium suggest the presence of additional mechanisms, other than Ah-receptor-mediated, by which the ALDH3 gene can be differentially regulated in a tissue-specific manner.

Development of Selective Inhibitors for Human Aldehyde Dehydrogenase 3A1 (ALDH3A1) for the Enhancement of Cyclophosphamide Cytotoxicity

“…[17,22,27] We used the A549 (ALDH1A1 and ALDH3A1 expressing), SF767 (ALDH3A1 expressing) and CCD-13Lu (ALDH non-expressing) cells to assess the overall cytotoxicity and ability of CB29 analogs to enhance the sensitivity of cyclophosphamide and its derivatives. [20,24] The ED 50 values for mafosfamide on A549 and SF767 cells were 125 μM and 150 μM, respectively, while primary cells, CCD-13Lu were more sensitive to mafosfamide with an ED 50 of 40 μM (Figure S2). CB29 as well as compounds 17, 2, 18, 19, 11, 8, 9 and 10 were chosen because they were selective toward ALDH3A1 and showed no inhibitory potential toward ALDH1A1, ALDH1A2, ALDH1A3, ALDH1B1 and ALDH2 activity in vitro .…”

“…[20,21,22,23] It is also expressed in some normal human tissues such as cornea and keratinocytes. [18,24] Despite being present in normal cells, studies have shown that Aldh3a1(-/-) knockout mice are viable. [25] High ALDH3A1 activity in normal cells protects these cells from the products of lipid peroxidation, [26] but can lead to drug resistance in tumor cells.…”

Wounding the cornea to learn how it heals

“…Discarded limbal rims from corneal transplants or eye banks are often a source for primary human corneal epithelial cells (de Paiva et al, 2005; Kahn et al, 1993; Kim et al, 2012; Nakamura et al, 2004). Rabbit (Cha et al, 2004; Fini and Girard, 1990), rat (Boesch et al, 1996; Marshall and Hanrahan, 1991), and mouse (Kawakita et al, 2004; Li et al, 2010) primary corneal epithelial cells have also been generated and used successfully. Due to its small size, it is more difficult to generate primary corneal epithelial cells from the mouse.…”

Aldehydemetabolism in the cardiovascular system

“…157 The nonspecific ALDHs are further classified into numerous subclasses based on their conserved nucleic acid sequences and are known as Class 1, Class 2 and Class 3 ALDHs, based on cell-specific expression and sequence identity. 158 3E1. Class 1 aldehyde dehydrogenases (ALDH1).…”

“…164,177 Also, ALDH3 has been shown to metabolize aldehydes formed during lipid peroxidation. 163 The high constitutive expression of ALDH3A1 in the corneal epithelium, 20-40% of the total water-soluble protein in mammalian corneal epithelium, 158 perhaps serves as protection against volatile aldehydes and/or may be associated with the detoxification of medium-chain aldehydes produced during UV-induced lipid peroxidation by acting as a UV-sink via binding NAD + and absorbing UV light. 178 Very low levels of ALDH3 activity are detectable in normal mammalian liver, but high levels of both hepatic ALDH3 transcript and enzyme activity are obtained after exposure to certain xenobiotics such as PAHs, 3-methylcholanthrene, or dioxins, including TCDD.…”