The ion balance of Shotokuseki extract promotes filaggrin fragmentation and increases amino acid production and pyrrolidone carboxylic acid content in three-dimensional cultured human epidermis

Natural moisturizing factor (NMF) in the stratum corneum contributes to the retention of moisture there. The purpose of this study was to determine the penetration of ions in Shotokuseki extract (SE) into the three-dimensional cultured epidermis and the effect of NMF on the biosynthesis of amino acids and pyrrolidone carboxylic acid formation. Various ions, amino acids and pyrrolidone carboxylic acid were quantified by inductively coupled plasma mass spectrometry, fully automatic amino acid analyzer or high-performance liquid chromatography (HPLC) in three-dimensional cultured epidermis after application of SE. Gene expression levels of profilaggrin, calpain1, caspase14, and bleomycin hydrolase, which are involved in NMF production, were determined by reverse-transcription qPCR and bleomycin hydrolase activity was determined by aminopeptidase assay. The application of SE increased Na, K, Mg, Ca, Al, and Fe levels in three-dimensional cultured epidermis. The mRNA levels of the starting material of amino acid synthesis profilaggrin, and calpain1 and bleomycin hydrolase, which are involved in its fragmentation, increased. The activity of bleomycin hydrolase also increased. Furthermore, the levels of amino acids and pyrrolidone carboxylic acid increased in the three-dimensional cultured epidermis. This suggests that the ionic composition of SE may be involved in its moisturizing effect on the stratum corneum. GRAPHICAL ABSTRACT: [Image: see text]