OBJECTIVES: Retinoic acid (RA) can prevent keratin formation and induce mucous differentiation in epithelia. In this study, we attempted to induce keratinizing squamous epithelium from human cholesteatoma epithelial (HCE) cells using an air-liquid interface (ALI) technique. We also examined the effect of RA on the phenotype of keratinizing HCE cells. MATERIAL AND METHODS: HCE cells were cultured in RA-free defined media at an ALI or in a submerged state. We examined the morphological differences between ALI and submerged cultures, and histologically investigated the changes of phenotype after RA treatment. We also determined the effect of RA on the mRNA expressions of the cornifin-alpha and mucin genes as indicators of squamous and mucous differentiation, respectively. RESULTS: Using an ALI technique, we were able to differentiate HCE cells into a keratinizing squamous epithelium. When we treated the keratinizing HCE cells with RA, the morphological phenotype progressively changed into mucociliary epithelium. In addition, the expression of cornifin-alpha mRNA was suppressed, and the expressions of mucin gene 5AC (MUC5AC) and MUC5B mRNA increased progressively with RA treatment. CONCLUSION: We successfully developed a culturing system for keratinizing differentiation of HCE cells using the ALI technique in a defined medium. Our study also clearly showed that RA treatment led to mucociliary differentiation of HCE cells.
OBJECTIVES:Retinoic acid (RA) can prevent keratin formation and induce mucous differentiation in epithelia. In this study, we attempted to induce keratinizing squamous epithelium from humancholesteatoma epithelial (HCE) cells using an air-liquid interface (ALI) technique. We also examined the effect of RA on the phenotype of keratinizing HCE cells. MATERIAL AND METHODS: HCE cells were cultured in RA-free defined media at an ALI or in a submerged state. We examined the morphological differences between ALI and submerged cultures, and histologically investigated the changes of phenotype after RA treatment. We also determined the effect of RA on the mRNA expressions of the cornifin-alpha and mucin genes as indicators of squamous and mucous differentiation, respectively. RESULTS: Using an ALI technique, we were able to differentiate HCE cells into a keratinizing squamous epithelium. When we treated the keratinizing HCE cells with RA, the morphological phenotype progressively changed into mucociliary epithelium. In addition, the expression of cornifin-alpha mRNA was suppressed, and the expressions of mucin gene 5AC (MUC5AC) and MUC5B mRNA increased progressively with RA treatment. CONCLUSION: We successfully developed a culturing system for keratinizing differentiation of HCE cells using the ALI technique in a defined medium. Our study also clearly showed that RA treatment led to mucociliary differentiation of HCE cells.