title: Distinct expression patterns and roles of aldehyde dehydrogenases in normal oral mucosa keratinocytes: differential inhibitory effects of a pharmacological inhibitor and RNAi-mediated knockdown on cellular phenotype and epithelial morphology.
abstract: Aldehyde dehydrogenases (ALDHs), enzymes responsible for detoxification and retinoic acid biosynthesis, are considered a potent functional stem cell marker of normal and malignant cells in many tissues. To date, however, there are no available data on ALDH distributions and functions in oral mucosa. This study aims to clarify the levels and types of ALDH expression using immunohistochemistry with accompanying mRNA expression as well as an ALDEFLUOR assay, and to assess phenotypic and histological changes after manipulation of the ALDH activity of oral keratinocytes to increase the potency of a tissue-engineered oral mucosa by a specific ALDH inhibitor, diethylaminobenzaldehyde (DEAB), together with small interfering RNA of ALDH1A3 and ALDH3A1. Results showed the mRNA and cytoplasmic protein expression of ALDH1A3 and ALDH3A1 to be mostly localized in the upper suprabasal layer although no ALDH1A1 immunoreaction was detected throughout the epithelium. Oral keratinocytes with high ALDH activity exhibited a profile of differentiating cells. By pharmacological inhibition, the phenotypic analysis revealed the proliferating cell-population shifting to a more quiescent state compared with untreated cells. Furthermore, a well-structured epithelial layer showing a normal differentiation pattern and a decrease in Ki-67 immunopositive basal cells was developed by DEAB incubation, suggesting a slower turnover rate efficient to maintain undifferentiated cells. Histological findings of a regenerated oral epithelium by ALDH1A3 siRNA were similar to those when treated with DEAB while ALDH3A1 siRNA eradicated the epithelial regenerative capacity. These observations suggest the effects of phenotypic and morphological alterations by DEAB on oral keratinocytes are mainly consequent to the inhibition of ALDH1A3 activity.
authors.affiliation: Division of Reconstructive Surgery for Oral and Maxillofacial Region, Department of Tissue Regeneration and Reconstruction, Niigata University Graduate School of Medical and Dental Sciences, 2-5274, Gakkocho-dori, Chuo-ku, Niigata 951-8514, Japan.
authors.forename: Hiroko
authors.initials: H
authors.lastname: Kato
authors.forename: Kenji
authors.initials: K
authors.lastname: Izumi
authors.forename: Taro
authors.initials: T
authors.lastname: Saito
authors.forename: Hisashi
authors.initials: H
authors.lastname: Ohnuki
authors.forename: Michiko
authors.initials: M
authors.lastname: Terada
authors.forename: Yoshiro
authors.initials: Y
authors.lastname: Kawano
authors.forename: Kayoko
authors.initials: K
authors.lastname: Nozawa-Inoue
authors.forename: Chikara
authors.initials: C
authors.lastname: Saito
authors.forename: Takeyasu
authors.initials: T
authors.lastname: Maeda
mesh_terms: D000328:Adult; D000444:Aldehyde Dehydrogenase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D000445:Aldehyde Oxidoreductases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005260:Female; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D015603:Keratinocytes / Q000201:enzymology* / Q000473:pathology; D019394:Ki-67 Antigen / Q000378:metabolism; D008297:Male; D009061:Mouth Mucosa / Q000201:enzymology* / Q000473:pathology; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics*; D012038:Regeneration / Q000187:drug effects; D010128:p-Aminoazobenzene / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: D004791:Enzyme Inhibitors; D019394:Ki-67 Antigen; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C007368:C.I. Solvent Yellow 56; D010128:p-Aminoazobenzene; D000445:Aldehyde Oxidoreductases; C480021:ALDH3A1 protein, human; D000444:Aldehyde Dehydrogenase; C018699:aldehyde dehydrogenase (NAD(P)+)