tretinoin and conduritol-epoxide

To examine factors that regulate ceramide production during keratinization of the human stratum corneum (SC), we developed a reconstructed human epidermal keratinization model in which a fresh layer of SC is newly formed within 1 week. Addition of the UDP-glucose: ceramide glucosyltransferase inhibitor 1-phenyl-2-decanoylamino-3-morpholino-1-propanol significantly diminished SC ceramide levels (expressed as µg/mg protein) with decreased glucosylceramide levels. Desipramine hydrochloride, an inhibitor of sphingomyelinase, also significantly reduced SC ceramide levels. Similarly, conduritol B epoxide, an inhibitor of β-glucocerebrosidase, significantly down-regulated SC ceramide levels and significantly increased glucosylceramide levels. These results indicate the reliability of this model to elucidate ceramide synthesis regulating factors. Using this model, we assessed the effects of the inflammatory cytokine interleukin-1α (IL-1α), several bioactive sphingolipids and all-trans retinoic acid (RA) on ceramide levels in the SC. Whereas treatment with IL-1α (at 10 nM) significantly down-regulated ceramide levels, treatment with sphingosylphosphorylcholine (at 50 µM) or sphingosine-1-phosphate (at 10 or 20 µM) distinctly up-regulated ceramide levels. Interestingly, RA (at low as 10 nM) significantly up-regulated ceramide levels without affecting the formation of the SC or levels of keratinization-related proteins in the epidermis. The increased levels of ceramide were accompanied by a significantly increased secretion of granulocyte-macrophage colony-stimulating factor as well as by a significantly down-regulated expression of acid-ceramidase at both the gene and protein levels. Taken together, our results underscore the superiority of this reconstructed human epidermal keratinization model to analyze factors that regulate ceramide synthesis, especially in human SC.

Topics: Acid Ceramidase; Cell Culture Techniques; Cells, Cultured; Ceramides; Desipramine; Epidermis; Feasibility Studies; Gene Expression Regulation; Glucosylceramidase; Glucosyltransferases; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Inositol; Interleukin-1alpha; Keratinocytes; Models, Biological; Morpholines; Sphingolipids; Sphingomyelin Phosphodiesterase; Tretinoin

Gaucher disease (GD) is associated with upregulation of CD1d and MHC-class II expression by monocytes. While the physiological impact of CD1d upregulation remains uncertain, it has been proposed that MHC-class II upregulation is associated with inflammation. Hereby, we show that the decrease in MHC-class II expression seen in GD patients under therapy correlates positively with chitotriosidase activity, a marker of inflamed macrophages. We also show that retinoic acid (RA) and the beta-glucocerebrosidase inhibitor conduritol-B-epoxide (CBE) lead to upregulation of CD1d expression by THP-1 cells, which correlated with an increase in mRNA expression. In vitro co-culture experiments showed that RA treated THP-1 cells were more stimulatory for CD4(+) than for CD8(+) T cells, as determined by CFSE loss, in comparison to untreated THP-1 cells. Interestingly, even though addition of exogenous isoglobotrihexosylceramide (iGb3), a physiological CD1d ligand, augmented the percentage of dividing CD4(+) T cells, we could not detect a significant expansion of CD4(+)Valpha24(+) invariant Natural Killer T (iNKT) cells. In contrast, addition of alpha-galactosylceramide (alpha-GC) induced expansion of Valpha24(+) iNKT cells as determined by using alpha-GC-loaded human CD1d dimers. These results strengthen the existence of a cross-talk between monocyte lipid accumulation, inflammation and changes in cell surface CD1d and MHC-class II in monocytes, which may result in inappropriate recognition events by immune cells and perpetuate chronic inflammation.

Topics: Antigens, CD1d; Antineoplastic Agents; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Division; Cell Line; Chronic Disease; Coculture Techniques; Enzyme Inhibitors; Gaucher Disease; Globosides; Hexosaminidases; Histocompatibility Antigens Class II; Humans; Inflammation; Inositol; Monocytes; Natural Killer T-Cells; Protein Multimerization; RNA, Messenger; Tretinoin; Trihexosylceramides; Up-Regulation