Nutrition Research and Practice

The Korean Nutrition Society (한국영양학회)
권호 표지
DOI QR코드

DOI QR Code

Analysis of ceramide metabolites in differentiating epidermal keratinocytes treated with calcium or vitamin C

  • Kim, Ju-Young (Department of Medical Nutrition, Graduate School of East-West Medical Science, Kyung Hee University) ;
  • Yun, Hye-Jeong (Department of Medical Nutrition, Graduate School of East-West Medical Science, Kyung Hee University) ;
  • Cho, Yun-Hi (Department of Medical Nutrition, Graduate School of East-West Medical Science, Kyung Hee University)
  • Received : 2011.06.30
  • Accepted : 2011.09.22
  • Published : 2011.10.31
Download PDF
⟨ Previous Next ⟩

Abstract

Ceramides (Cer) comprise the major constituent of sphingolipids in the epidermis and are known to play diverse roles in the outermost layers of the skin including water retention and provision of a physical barrier. In addition, they can be hydrolyzed into free sphingoid bases such as $C_{18}$ sphingosine (SO) and $C_{18}$ sphinganine (SA) or can be further metabolized to $C_{18}$ So-1-phosphate (S1P) and $C_{18}$ Sa-1-phosphate (Sa1P) in keratinocytes. The significance of ceramide metabolites emerged from studies reporting altered levels of SO and SA in skin disorders and the role of S1P and Sa1P as signaling lipids. However, the overall metabolism of sphingoid bases and their phosphates during keratinocyte differentiation remains not fully understood. Therefore, in this study, we analyzed these Cer metabolites in the process of keratinocyte differentiation. Three distinct keratinocyte differentiation stages were prepared using 0.07 mM calcium (Ca$^{2+}$) (proliferation stage), 1.2 mM Ca$^{2+}$ (early differentiation stage) in serum-free medium, or serum-containing medium with vitamin C (50 ${\mu}L$/mL) (late differentiation stage). Serum-containing medium was also used to determine whether vitamin C increases the concentrations of sphingoid bases and their phosphates. The production of sphingoid bases and their phosphates after hydrolysis by alkaline phosphatase was determined using high-performance liquid chromatography. Compared to cells treated with 0.07 mM Ca$^{2+}$, levels of SO, SA, S1P, and SA1P were not altered after treatment with 1.2 mM Ca$^{2+}$. However, in keratinocytes cultured in serum-containing medium with vitamin C, levels of SO, SA, S1P, and SA1P were dramatically higher than those in 0.07- and l.2-mM Ca$^{2+}$-treated cells; however, compared to serum-containing medium alone, vitamin C did not significantly enhance their production. Taken together, we demonstrate that late differentiation induced by vitamin C and serum was accompanied by dramatic increases in the concentration of sphingoid bases and their phosphates, although vitamin C alone had no effect on their production.

Keywords

References

  1. Houben E, De Paepe K, Rogiers V. A keratinocyte's course of life. Skin Pharmacol Physiol 2007;20:122-32. https://doi.org/10.1159/000098163
  2. Downing DT, Stewart ME, Wertz PW, Colton SW, Abraham W, Strauss JS. Skin lipids: an update. J Invest Dermatol 1987;88:2s-6s. https://doi.org/10.1111/1523-1747.ep12468850
  3. Elias PM, Menon GK. Structural and lipid biochemical correlates of the epidermal permeability barrier. Adv Lipid Res 1991;24:1-26.
  4. Hennings H, Michael D, Cheng C, Steinert P, Holbrook K, Yuspa SH. Calcium regulation of growth and differentiation of mouse epidermal cells in culture. Cell 1980;19:245-54. https://doi.org/10.1016/0092-8674(80)90406-7
  5. Ponec M, Weerheim A, Kempenaar J, Mulder A, Gooris GS, Bouwstra J, Mommaas AM. The formation of competent barrier lipids in reconstructed human epidermis requires the presence of vitamin C. J Invest Dermatol 1997;109:348-55. https://doi.org/10.1111/1523-1747.ep12336024
  6. Uchida Y, Behne M, Quiec D, Elias PM, Holleran WM. Vitamin C stimulates sphingolipid production and markers of barrier formation in submerged human keratinocyte cultures. J Invest Dermatol 2001;117:1307-13. https://doi.org/10.1046/j.0022-202x.2001.01555.x
  7. Menon GK, Grayson S, Elias PM. Ionic calcium reservoirs in mammalian epidermis: ultrastructural localization by ion-capture cytochemistry. J Invest Dermatol 1985;84:508-12. https://doi.org/10.1111/1523-1747.ep12273485
  8. Holleran WM, Takagi Y, Uchida Y. Epidermal sphingolipids: metabolism, function, and roles in skin disorders. FEBS Lett 2006;580:5456-66. https://doi.org/10.1016/j.febslet.2006.08.039
  9. Mizutani Y, Mitsutake S, Tsuji K, Kihara A, Igarashi Y. Ceramide biosynthesis in keratinocyte and its role in skin function. Biochimie 2009;91:784-90. https://doi.org/10.1016/j.biochi.2009.04.001
  10. Geilen CC, Barz S, Bektas M. Sphingolipid signaling in epidermal homeostasis. Current knowledge and new therapeutic approaches in dermatology. Skin Pharmacol Appl Skin Physiol 2001;14:261-71. https://doi.org/10.1159/000056356
  11. Wertz PW, Downing DT. Free sphingosine in human epidermis. J Invest Dermatol 1990;94:159-61. https://doi.org/10.1111/1523-1747.ep12874122
  12. Wertz PW, Downing DT. Free sphingosines in porcine epidermis. Biochim Biophys Acta 1989;1002:213-7. https://doi.org/10.1016/0005-2760(89)90289-0
  13. Wakita H, Nishimura K, Takigawa M. Composition of free long-chain (sphingoid) bases in stratum corneum of normal and pathologic human skin conditions. J Invest Dermatol 1992;99:617-22. https://doi.org/10.1111/1523-1747.ep12668019
  14. Bibel DJ, Aly R, Shinefield HR. Antimicrobial activity of sphingosines. J Invest Dermatol 1992;98:269-73. https://doi.org/10.1111/1523-1747.ep12497842
  15. Arikawa J, Ishibashi M, Kawashima M, Takagi Y, Ichikawa Y, Imokawa G. Decreased levels of sphingosine, a natural antimicrobial agent, may be associated with vulnerability of the stratum corneum from patients with atopic dermatitis to colonization by Staphylococcus aureus. J Invest Dermatol 2002;119:433-9. https://doi.org/10.1046/j.1523-1747.2002.01846.x
  16. Gupta AK, Fisher GJ, Elder JT, Nickoloff BJ, Voorhees JJ. Sphingosine inhibits phorbol ester-induced inflammation, ornithine decarboxylase activity, and activation of protein kinase C in mouse skin. J Invest Dermatol 1988;91:486-91. https://doi.org/10.1111/1523-1747.ep12476635
  17. Kawanabe T, Kawakami T, Yatomi Y, Shimada S, Soma Y. Sphingosine 1-phosphate accelerates wound healing in diabetic mice. J Dermatol Sci 2007;48:53-60. https://doi.org/10.1016/j.jdermsci.2007.06.002
  18. Vogler R, Sauer B, Kim DS, Schafer-Korting M, Kleuser B. Sphingosine-1-phosphate and its potentially paradoxical effects on critical parameters of cutaneous wound healing. J Invest Dermatol 2003;120:693-700. https://doi.org/10.1046/j.1523-1747.2003.12096.x
  19. Houben E, Holleran WM, Yaginuma T, Mao C, Obeid LM, Rogiers V, Takagi Y, Elias PM, Uchida Y. Differentiationassociated expression of ceramidase isoforms in cultured keratinocytes and epidermis. J Lipid Res 2006;47:1063-70. https://doi.org/10.1194/jlr.M600001-JLR200
  20. Labarca C, Paigen K. A simple, rapid, and sensitive DNA assay procedure. Anal Biochem 1980;102:344-52. https://doi.org/10.1016/0003-2697(80)90165-7
  21. Ruwisch L, Schafer-Korting M, Kleuser B. An improved highperformance liquid chromatographic method for the determination of sphingosine-1-phosphate in complex biological materials. Naunyn Schmiedebergs Arch Pharmacol 2001;363:358-63. https://doi.org/10.1007/s002100000365
  22. Min JK, Yoo HS, Lee EY, Lee WJ, Lee YM. Simultaneous quantitative analysis of sphingoid base 1-phosphates in biological samples by o-phthalaldehyde precolumn derivatization after dephosphorylation with alkaline phosphatase. Anal Biochem 2002;303:167-75. https://doi.org/10.1006/abio.2002.5579
  23. Yoon HT, Yoo HS, Shin BK, Lee WJ, Kim HM, Hong SP, Moon DC, Lee YM. Improved fluorescent determination method of cellular sphingoid bases in high-performance liquid chromatography. Arch Pharm Res 1999;22:294-9. https://doi.org/10.1007/BF02976365
  24. Pillai S, Bikle DD, Hincenbergs M, Elias PM. Biochemical and morphological characterization of growth and differentiation of normal human neonatal keratinocytes in a serum-free medium. J Cell Physiol 1988;134:229-37. https://doi.org/10.1002/jcp.1041340208
  25. Herzinger T, Kleuser B, Schafer-Korting M, Korting HC. Sphingosine-1-phosphate signaling and the skin. Am J Clin Dermatol 2007;8:329-36. https://doi.org/10.2165/00128071-200708060-00002
  26. Youm JK, Jo H, Hong JH, Shin DM, Kwon MJ, Jeong SK, Park BD, Choi EH, Lee SH. K6PC-5, a sphingosine kinase activator, induces anti-aging effects in intrinsically aged skin through intracellular $Ca^{2+}$ signaling. J Dermatol Sci 2008;51:89-102. https://doi.org/10.1016/j.jdermsci.2008.03.002
  27. Manggau M, Kim DS, Ruwisch L, Vogler R, Korting HC, Schafer-Korting M, Kleuser B. 1Alpha,25-dihydroxyvitamin D3 protects human keratinocytes from apoptosis by the formation of sphingosine-1-phosphate. J Invest Dermatol 2001;117:1241-9. https://doi.org/10.1046/j.0022-202x.2001.01496.x
  28. Mao C, Obeid LM. Ceramidases: regulators of cellular responses mediated by ceramide, sphingosine, and sphingosine-1-phosphate. Biochim Biophys Acta 2008;1781:424-34. https://doi.org/10.1016/j.bbalip.2008.06.002
  29. Sun W, Xu R, Hu W, Jin J, Crellin HA, Bielawski J, Szulc ZM, Thiers BH, Obeid LM, Mao C. Upregulation of the human alkaline ceramidase 1 and acid ceramidase mediates calciuminduced differentiation of epidermal keratinocytes. J Invest Dermatol 2008;128:389-97.
  30. Coderch L, Lopez O, de la Maza A, Parra JL. Ceramides and skin function. Am J Clin Dermatol 2003;4:107-29. https://doi.org/10.2165/00128071-200304020-00004
  31. Castiel-Higounenc I, Chopart M, Ferraris C. Stratum corneum lipids: specificity, role, deficiencies and modulation. Eur J Dermatol 2004;11:401-6.
  32. Sando GN, Howard EJ, Madison KC. Induction of ceramide glucosyltransferase activity in cultured human keratinocytes. Correlation with culture differentiation. J Biol Chem 1996;271:22044-51. https://doi.org/10.1074/jbc.271.36.22044
  33. Uchida Y, Holleran WM. Omega-O-acylceramide, a lipid essential for mammalian survival. J Dermatol Sci 2008;51:77-87. https://doi.org/10.1016/j.jdermsci.2008.01.002
  34. Behne M, Uchida Y, Seki T, de Montellano PO, Elias PM, Holleran WM. Omega-hydroxyceramides are required for corneocyte lipid envelope (CLE) formation and normal epidermal permeability barrier function. J Invest Dermatol 2000;114:185-92. https://doi.org/10.1046/j.1523-1747.2000.00846.x
  35. Ternes P, Franke S, Zahringer U, Sperling P, Heinz E. Identification and characterization of a sphingolipid delta 4-desaturase family. J Biol Chem 2002;277:25512-8. https://doi.org/10.1074/jbc.M202947200
  36. Holland WL, Brozinick JT, Wang LP, Hawkins ED, Sargent KM, Liu Y, Narra K, Hoehn KL, Knotts TA, Siesky A, Nelson DH, Karathanasis SK, Fontenot GK, Birnbaum MJ, Summers SA. Inhibition of ceramide synthesis ameliorates glucocorticoid-, saturated-fat-, and obesity-induced insulin resistance. Cell Metab 2007;5:167-79. https://doi.org/10.1016/j.cmet.2007.01.002
  37. Motta S, Monti M, Sesana S, Caputo R, Carelli S, Ghidoni R. Ceramide composition of the psoriatic scale. Biochim Biophys Acta 1993;1182:147-51. https://doi.org/10.1016/0925-4439(93)90135-N
  38. Pitson SM, D'andrea RJ, Vandeleur L, Moretti PA, Xia P, Gamble JR, Vadas MA, Wattenberg BW. Human sphingosine kinase: purification, molecular cloning and characterization of the native and recombinant enzymes. Biochem J 2000;350:429-41. https://doi.org/10.1042/0264-6021:3500429
  39. Olivera A, Kohama T, Tu Z, Milstien S, Spiegel S. Purification and characterization of rat kidney sphingosine kinase. J Biol Chem 1998;273:12576-83. https://doi.org/10.1074/jbc.273.20.12576
  40. Meyer zu Heringdorf D, Lass H, Alemany R, Laser KT, Neumann E, Zhang C, Schmidt M, Rauen U, Jakobs KH, van Koppen CJ. Sphingosine kinase-mediated $Ca^{2+}$ signalling by G-protein-coupled receptors. EMBO J 1998;17:2830-7. https://doi.org/10.1093/emboj/17.10.2830
  41. Hong JH, Youm JK, Kwon MJ, Park BD, Lee YM, Lee SI, Shin DM, Lee SH. K6PC-5, a direct activator of sphingosine kinase 1, promotes epidermal differentiation through intracellular $Ca^{2+}$ signaling. J Invest Dermatol 2008;128:2166-78. https://doi.org/10.1038/jid.2008.66
  42. Xu SZ, Muraki K, Zeng F, Li J, Sukumar P, Shah S, Dedman AM, Flemming PK, McHugh D, Naylor J, Cheong A, Bateson AN, Munsch CM, Porter KE, Beech DJ. A sphingosine-1-phosphate-activated calcium channel controlling vascular smooth muscle cell motility. Circ Res 2006;98:1381-9. https://doi.org/10.1161/01.RES.0000225284.36490.a2
  43. Bornfeldt KE, Graves LM, Raines EW, Igarashi Y, Wayman G, Yamamura S, Yatomi Y, Sidhu JS, Krebs EG, Hakomori S, Ross R. Sphingosine-1-phosphate inhibits PDGF-induced chemotaxis of human arterial smooth muscle cells: spatial and temporal modulation of PDGF chemotactic signal transduction. J Cell Biol 1995;130:193-206. https://doi.org/10.1083/jcb.130.1.193
  44. Sauer B, Vogler R, von Wenckstern H, Fujii M, Anzano MB, Glick AB, Schafer-Korting M, Roberts AB, Kleuser B. Involvement of Smad signaling in sphingosine 1-phosphate-mediated biological responses of keratinocytes. J Biol Chem 2004;279:38471-9. https://doi.org/10.1074/jbc.M313557200
  45. Kim J, Kim Y, Yun H, Park H, Kim SY, Lee KG, Han SM, Cho Y. Royal jelly enhances migration of human dermal fibroblasts and alters the levels of cholesterol and sphinganine in an in vitro wound healing model. Nutr Res Pract 2010;4:362-8. https://doi.org/10.4162/nrp.2010.4.5.362

Cited by

  1. Topical apigenin improves epidermal permeability barrier homoeostasis in normal murine skin by divergent mechanisms vol.22, pp.3, 2013, https://doi.org/10.1111/exd.12102
  2. Vitamin C Stimulates Epidermal Ceramide Production by Regulating Its Metabolic Enzymes vol.23, pp.6, 2015, https://doi.org/10.4062/biomolther.2015.044
  3. The concept of sphingolipid rheostat in skin: a driving force for new active ingredients in cosmetic applications pp.2257-6614, 2018, https://doi.org/10.1051/ocl/2018043
  4. Role of Vitamin C in Skin Diseases vol.9, pp.1664-042X, 2018, https://doi.org/10.3389/fphys.2018.00819