title: Expression of NF-kappaB in epidermis and the relationship between NF-kappaB activation and inhibition of keratinocyte growth.
abstract: BACKGROUND
  Nuclear factor-kappaB (NF-kappaB) is a transcription factor involved in a number of signalling pathways in many cell types. NF-kappaB in mice has been implicated as an important regulator of keratinocyte proliferation and differentiation.
  
  
  OBJECTIVE
  To evaluate the role of NF-kappaB in keratinocyte growth in human beings, we examined its expression in keratinocytes both in culture and in situ, and studied the relationship between NF-kappaB activation and the inhibition of keratinocyte proliferation induced by known modulators of keratinocyte growth.
  
  
  METHODS
  The expression of subunits of the NF-kappaB family was examined in human skin, primary cultured keratinocytes and an immortalized keratinocyte line by immunohistochemistry and reverse transcriptase-polymerase chain reaction analysis. NF-kB activation was examined in keratinocytes treated with various modulating agents by electrophoretic mobility shift assay (for DNA-binding activity) and by immunocytochemistry (nuclear translocation). The proliferative capacity of treated keratinocytes was also examined by 3H-thymidine incorporation, cell cycle analysis, and expression of Ki-67, a nuclear marker for cell proliferation. The involvement of NF-kappaB was assessed using sodium salicylate, which inhibits NF-kappaB activation.
  
  
  RESULTS
  The NF-kappaB subunits, p50, p65, RelB, and c-Rel (but not p52), were detected in keratinocytes and in normal epidermis at mRNA and protein levels. The four subunits were expressed in a cytoplasmic (rather than a nuclear) pattern in both basal and suprabasal keratinocytes. Phorbol myristate acetate (PMA), tumour necrosis factor alpha, and interferon gamma each activated NF-kappaB and inhibited keratinocyte proliferation. Lipopolysaccharide and dexamethasone did not activate NF-kappaB and had the least effect on proliferation. Finally, a high concentration of calcium (Ca2+) and retinoic acid each failed to activate NF-kappaB, but were potent inhibitors of keratinocyte proliferation, respectively. PMA-induced cell cycle arrest of keratinocytes was blocked by pretreatment with sodium salicylate.
  
  
  CONCLUSIONS
  NF-kappaB is constitutively expressed in a resting state in both human cultured keratinocytes and the epidermis. Activation of NF-kappaB is required for PMA-induced keratinocyte growth arrest.
authors.affiliation: Department of Dermatology, The University of Texas Southwestern Medical Center, and the Dallas Veterans Affairs Medical Center, Dallas, TX 75390, USA.
authors.forename: J
authors.initials: J
authors.lastname: Takao
authors.forename: T
authors.initials: T
authors.lastname: Yudate
authors.forename: A
authors.initials: A
authors.lastname: Das
authors.forename: S
authors.initials: S
authors.lastname: Shikano
authors.forename: M
authors.initials: M
authors.lastname: Bonkobara
authors.forename: K
authors.initials: K
authors.lastname: Ariizumi
authors.forename: P D
authors.initials: PD
authors.lastname: Cruz
mesh_terms: D002453:Cell Cycle / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D004817:Epidermis / Q000378:metabolism*; D015870:Gene Expression; D006801:Humans; D007124:Immunoenzyme Techniques; D007231:Infant, Newborn; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism*; D008297:Male; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012980:Sodium Salicylate / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000037:antagonists & inhibitors / Q000494:pharmacology
chemical_list: D016328:NF-kappa B; D012333:RNA, Messenger; D013755:Tetradecanoylphorbol Acetate; D012980:Sodium Salicylate