title: Neural stem cells inhibit melanin production by activation of Wnt inhibitors.
abstract: BACKGROUND
  Melanin for skin pigmentation is synthesized from tyrosine via an enzymatic cascade that is controlled by tyrosinase (TYR), tyrosinase-related protein 1 (TRP1), and dopachrome tautomerase/tyrosinase related protein 2 (Dct/TRP2), which are the targets of microphthalmia-associated transcription factor (MITF). MITF is a master regulator of pigmentation and a target of β-catenin in Wnt/β-catenin signaling during melanocyte differentiation. Stem cells have been used in skin pigmentation studies, but the mechanisms were not determined for the conditioned medium (CM)-mediated effects.
  
  
  OBJECTIVE
  In this study, the inhibition and mechanisms of melanin synthesis were elucidated in B16 melanoma cells and UV-B irradiated C57/BL-6 mice that were treated with human neural stem cell-conditioned medium (NSC-CM).
  
  
  METHODS
  B16-F10 melanoma cells (1.5×10(4)cells/well) and the shaved dorsal skin of mice were pretreated with various amount (5, 10, 20, 50, and 100%) of NSC-CM. Melanin contents and TYR activity were measured by a Spectramax spectrophotometer. The expression of TYR, TRP1, Dct/TRP2, MITF, β-catenin and Wnt inhibitors were evaluated by RT-PCR and western blot. The dorsal skin samples were analyzed by immunofluorescence with various antibodies and compared with that control of tissues.
  
  
  RESULTS
  Marked decreases were evident in melanin content and TYR, TRP1, DCT/TRP2, MITF, and β-catenin expression in B16 cells and C57/BL-6 mice. NSC-CM negatively regulated Wnt/β-catenin signaling by decreasing the expression of β-catenin protein, which resulted from robust expression of Wnt inhibitors Dickkopf-1 (DKK1) and secreted frizzled-related protein 2 (sFRP2).
  
  
  CONCLUSIONS
  These results demonstrate that NSC-CM suppresses melanin production in vitro and in vivo, suggesting that factors in NSC-CM may play an important role in deregulation of epidermal melanogenesis.
authors.affiliation: Laboratory of Stem Cell Biology, Department of Biomedical Science, College of Health Science, Korea University, Jeongneung-dong, Sungbuk-gu, Seoul 136-703, Republic of Korea; Department of Health Science, Korea University Graduate School, Jeongneung-dong, Sungbuk-gu, Seoul 136-703, Republic of Korea.
authors.forename: Insik
authors.initials: I
authors.lastname: Hwang
authors.forename: Ju-Hwang
authors.initials: JH
authors.lastname: Park
authors.forename: Hang-Soo
authors.initials: HS
authors.lastname: Park
authors.forename: Kyung-Ah
authors.initials: KA
authors.lastname: Choi
authors.forename: Ki-Cheon
authors.initials: KC
authors.lastname: Seol
authors.forename: Seung-Ick
authors.initials: SI
authors.lastname: Oh
authors.forename: Seongman
authors.initials: S
authors.lastname: Kang
authors.forename: Sunghoi
authors.initials: S
authors.lastname: Hong
mesh_terms: D000818:Animals; D051177:Catenins / Q000378:metabolism; D045744:Cell Line, Tumor; D017077:Culture Media, Conditioned; D005786:Gene Expression Regulation; D008543:Melanins / Q000096:biosynthesis*; D008546:Melanoma, Experimental / Q000378:metabolism*; D051379:Mice; D051739:Microphthalmia-Associated Transcription Factor / Q000378:metabolism; D014442:Monophenol Monooxygenase / Q000378:metabolism; D058953:Neural Stem Cells / Q000502:physiology*; D010858:Pigmentation; D051153:Wnt Proteins / Q000378:metabolism*
chemical_list: D051177:Catenins; D017077:Culture Media, Conditioned; D008543:Melanins; D051739:Microphthalmia-Associated Transcription Factor; C497235:Mitf protein, mouse; D051153:Wnt Proteins; D014442:Monophenol Monooxygenase
keywords: (2′Z, 3′E)-6-bromoindirubin-3′-oxime; APC; BIO; CK1α; DKK1,2,3; Dct/TRP2; Dickkopf-1,2,3; Dishevelled; Dvl; GSK-3β; MITF; Melanin; NSC culture medium; NSC-CM; Neural stem cell-conditioned medium; PMSF; SCF; T-cell factor/lymphoid enhancer factor; TCF/LEF; TRP1; TYR; Tyrosinase; WIF1; Wnt inhibitors; Wnt inhibitory factor1; adenomatous polyposis coli; casein kinase1α; dopachrome tautomerase/tyrosinase related protein 2; glycogen synthase kinase 3β; microphthalmia-associated transcription factor; neural stem cell conditioned medium; non-CM; phenylmethyl sulfonyl fluoride; sFRP1,2,4,5; secreted frizzled-related protein; stem cell factor; tyrosinase; tyrosinase-related protein 1; α-MSH; α-melanocyte-stimulating hormone