title: Keratinocytes in culture accumulate phagocytosed melanosomes in the perinuclear area.
abstract: There are many techniques for evaluating melanosome transfer to keratinocytes but the spectrophotometric quantification of melanosomes incorporated by keratinocyte phagocytosis has not been previously reported. Here we describe a new method that allows the spectrophotometric visualization of melanosome uptake by normal human keratinocytes in culture. Fontana-Masson staining of keratinocytes incubated with isolated melanosomes showed the accumulation of incorporated melanosomes in the perinuclear areas of keratinocytes within 48 h. Electron microscopic observations of melanosomes ingested by keratinocytes revealed that many phagosomes containing clusters of melanosomes or their fragments were localized in the perinuclear area. A known inhibitor of keratinocyte phagocytosis which inhibits protease-activated receptor-2, i.e., soybean trypsin inhibitor, decreased melanosome uptake by keratinocytes in a dose-dependent manner. These data suggest that our method is a useful model to quantitate keratinocyte phagocytosis of melanosomes visually in vitro.
authors.affiliation: Skin Aging and Photo-aging Research Center, Doshisha University, Kizugawa, Kyoto, Japan. hando@mail.doshisha.ac.jp
authors.forename: Hideya
authors.initials: H
authors.lastname: Ando
authors.forename: Yoko
authors.initials: Y
authors.lastname: Niki
authors.forename: Masaki
authors.initials: M
authors.lastname: Yoshida
authors.forename: Masaaki
authors.initials: M
authors.lastname: Ito
authors.forename: Kaoru
authors.initials: K
authors.lastname: Akiyama
authors.forename: Jin-Hwa
authors.initials: JH
authors.lastname: Kim
authors.forename: Tae-Jin
authors.initials: TJ
authors.lastname: Yoon
authors.forename: Jeung-Hoon
authors.initials: JH
authors.lastname: Lee
authors.forename: Mary S
authors.initials: MS
authors.lastname: Matsui
authors.forename: Masamitsu
authors.initials: M
authors.lastname: Ichihashi
mesh_terms: D018929:Cell Culture Techniques; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000378:metabolism / Q000648:ultrastructure*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism / Q000648:ultrastructure*; D008543:Melanins / Q000378:metabolism; D020460:Melanosomes / Q000378:metabolism / Q000648:ultrastructure*; D046529:Microscopy, Electron, Transmission; D008954:Models, Biological; D010587:Phagocytosis / Q000502:physiology*; D010588:Phagosomes / Q000378:metabolism / Q000648:ultrastructure*; D044464:Receptor, PAR-2 / Q000037:antagonists & inhibitors / Q000378:metabolism; D013053:Spectrophotometry / Q000379:methods; D013194:Staining and Labeling
chemical_list: D004791:Enzyme Inhibitors; D008543:Melanins; D044464:Receptor, PAR-2