title: K6PC-5, a direct activator of sphingosine kinase 1, promotes epidermal differentiation through intracellular Ca2+ signaling.
abstract: Sphingosine-1-phosphate (S1P), a bioactive sphingolipid metabolite, regulates multiple cellular responses such as Ca(2+) signaling, growth, survival, and differentiation. Because sphingosine kinase (SphK) is the enzyme directly responsible for production of S1P, many factors have been identified that regulate its activity and subsequent S1P levels. Here we synthesized a previously unidentified SphK activator, K6PC-5, and have studied its effects on intracellular Ca(2+) signaling in HaCaT cells and epidermal differentiation in murine skin. K6PC-5, a hydrophobic compound chemically named N-(1,3-dihydroxyisopropyl)-2-hexyl-3-oxo-decanamide, activated SphK (obtained from C57BL/6 murine blood and F9-12 cell lysates) in a dose-dependent manner. K6PC-5 induced both intracellular Ca(2+) concentration ([Ca(2+)](i)) oscillations in HaCaT cells and Ca(2+) mobilization in hairless mouse epidermis. Both dimethylsphingosine (DMS) and dihydroxysphingosine (DHS), SphK inhibitors, and transfection of SphK1-siRNA blocked K6PC-5-induced increases in [Ca(2+)](i). The K6PC-5-induced [Ca(2+)](i) oscillations were dependent on thapsigargin-sensitive Ca(2+) stores and Ca(2+) entry, but independent of the classical phospholipase C-mediated pathway. In addition, K6PC-5 enhanced the expression of involucrin and filaggrin, specific differentiation-associated marker proteins in HaCaT cells, whereas transfection of SphK1-siRNA blocked the increase of involucrin. Topical K6PC-5 also enhanced the expression of involucrin, loricrin, filaggrin, and keratin 5 in intact murine epidermis. Finally, topical K6PC-5 inhibited epidermal hyperplasia by exerting antiproliferative effects on keratinocytes in murine epidermis. These results suggest that K6PC-5 acts to regulate both differentiation and proliferation of keratinocytes via [Ca(2+)](i) responses through S1P production. Thus, regulation of S1P levels may represent a novel approach for treatment of skin disorders characterized by abnormal differentiation and proliferation, such as atopic dermatitis and psoriasis.
authors.affiliation: Department of Oral Biology, Brain Korea 21 Project, Oral Science Research Center, Center for Natural Defense System, Yonsei University College of Dentistry, Seoul, Korea.
authors.forename: Jeong Hee
authors.initials: JH
authors.lastname: Hong
authors.forename: Jong-Kyung
authors.initials: JK
authors.lastname: Youm
authors.forename: Mi Jung
authors.initials: MJ
authors.lastname: Kwon
authors.forename: Byeong Deog
authors.initials: BD
authors.lastname: Park
authors.forename: Yong-Moon
authors.initials: YM
authors.lastname: Lee
authors.forename: Syng-Ill
authors.initials: SI
authors.lastname: Lee
authors.forename: Dong Min
authors.initials: DM
authors.lastname: Shin
authors.forename: Seung Hun
authors.initials: SH
authors.lastname: Lee
mesh_terms: D000577:Amides / Q000494:pharmacology*; D000818:Animals; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D020536:Enzyme Activators / Q000494:pharmacology*; D004817:Epidermis / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D005260:Female; D000091344:Filaggrin Proteins; D006801:Humans; D006965:Hyperplasia; D007381:Intermediate Filament Proteins / Q000378:metabolism; D053555:Keratin-5 / Q000378:metabolism; D015603:Keratinocytes / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D008246:Lysophospholipids / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D008812:Mice, Hairless; D008810:Mice, Inbred C57BL; D009373:Neoplasms, Germ Cell and Embryonal / Q000473:pathology; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D011498:Protein Precursors / Q000378:metabolism; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000378:metabolism; D018243:Teratocarcinoma / Q000473:pathology; D000099157:Sphingosine Kinase
chemical_list: D000577:Amides; D002118:Calcium; D020536:Enzyme Activators; D000091344:Filaggrin Proteins; D007381:Intermediate Filament Proteins; D053555:Keratin-5; D008246:Lysophospholipids; D008565:Membrane Proteins; D017853:Phosphotransferases (Alcohol Group Acceptor); D011498:Protein Precursors; D013110:Sphingosine; D000099157:Sphingosine Kinase; C000717249:FLG protein, human; C531172:N-(1,3-dihydroxyisopropyl)-2-hexyl-3-oxodecanamide; C064554:loricrin; C060506:sphingosine 1-phosphate; C032253:involucrin