title: Inhibition of tumor necrosis factor-alpha stimulated NFkappaB/p65 in human keratinocytes by alpha-melanocyte stimulating hormone and adrenocorticotropic hormone peptides.
abstract: Alpha-melanocyte stimulating hormone (alpha-MSH) has pigmentary, anti-inflammatory, antipyretic, and general immunomodulatory roles. It can oppose several cytokines including tumor necrosis factor-alpha in a number of tissues, including skin. We have previously shown that alpha-MSH can inhibit tumor necrosis factor-alpha stimulated intercellular adhesion molecule 1 upregulation and nuclear factor kappaB (NFkappaB) transcription factor activation in melanocyte and melanoma cells. It is thought, however, that this MSH biology may also extend to other cells of the skin and in this study we extend our work to keratinocytes. We have investigated in detail the ability of three alpha-MSH peptides to inhibit tumor necrosis factor alpha stimulated NFkappaB activation in nonpigmentary HaCaT keratinocytes (alpha-MSH, L-Lys-L-Pro-L-Val, and L-Lys-L-Pro-D-Val) and two adrenocorticotropic hormone (ACTH) peptides (1-17 and 1-39), reported to be present in skin tissue. NFkappaB/p65 activation was analyzed by electrophoretic mobility shift assay and immunofluorescent microscopy. alpha-MSH, L-Lys-L-Pro-L-Val, and L-Lys-L-Pro-D-Val all significantly inhibited tumor necrosis factor alpha stimulated NFkappaB activation, whereas ACTH 1-17 and 1-39 did not, in the HaCaT keratinocytes. MSH peptides and ACTH 1-39 were effective, however, at inhibiting NFkappaB activation in normal human keratinocytes. Immunolabeling of inhibitor kappaBalpha of NFkappaB (IkappaBalpha) revealed an abnormal localization to the nucleus of HaCaT cells, which was unaffected by MSH/ACTH peptides. In contrast, normal human keratinocytes showed a normal IkappaBalpha distribution that responded to MSH/ACTH with nuclear translocation. Our data support previous work on the role of MSH/ACTH peptides as immunomodulatory/anti-inflammatory regulators, and extend this work to keratinocytes identifying a novel IkappaBalpha mechanism and extends findings to ACTH peptides, identifying an abnormal IkappaBalpha mechanism in the immortal HaCaT versus normal keratinocyte.
authors.affiliation: University of Sheffield, Section of Medicine, Division of Clinical Sciences, Northern General Hospital, Sheffield, UK.
authors.forename: Manar
authors.initials: M
authors.lastname: Moustafa
authors.forename: Marika
authors.initials: M
authors.lastname: Szabo
authors.forename: Ghanem E
authors.initials: GE
authors.lastname: Ghanem
authors.forename: Renato
authors.initials: R
authors.lastname: Morandini
authors.forename: E Helen
authors.initials: EH
authors.lastname: Kemp
authors.forename: Sheila
authors.initials: S
authors.lastname: MacNeil
authors.forename: John W
authors.initials: JW
authors.lastname: Haycock
mesh_terms: D000324:Adrenocorticotropic Hormone / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D015153:Blotting, Western; D002460:Cell Line; D004347:Drug Interactions; D006801:Humans; D020672:I-kappa B Proteins / Q000378:metabolism; D015603:Keratinocytes / Q000737:chemistry / Q000166:cytology / Q000378:metabolism*; D007668:Kidney / Q000166:cytology; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D010446:Peptide Fragments / Q000494:pharmacology*; D011333:Pro-Opiomelanocortin / Q000494:pharmacology; D011869:Radioligand Assay; D044103:Receptor, Melanocortin, Type 2; D018043:Receptors, Corticotropin / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D044101:Receptors, Melanocortin; D051996:Transcription Factor RelA; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D000521:alpha-MSH / Q000494:pharmacology*
chemical_list: D000970:Antineoplastic Agents; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D010446:Peptide Fragments; D044103:Receptor, Melanocortin, Type 2; D018043:Receptors, Corticotropin; D044101:Receptors, Melanocortin; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; D000072000:NF-KappaB Inhibitor alpha; D000521:alpha-MSH; D011333:Pro-Opiomelanocortin; D000324:Adrenocorticotropic Hormone; C028238:ACTH (1-17)