Calcium triggers beta-defensin (hBD-2 and hBD-3) and chemokine macrophage inflammatory protein-3 alpha (MIP-3alpha/CCL20) expression in monolayers of activated human keratinocytes.
The inducible epidermal beta-defensins and the chemokine macrophage inflammatory protein-3alpha (MIP-3alpha/CCL20) are important mediators involved in innate and adaptive immunity and in the recruitment of immune cells. The aim of our study was to determine whether calcium could trigger the induction of beta-defensins (hBD-2 and hBD-3) mRNA and the release of MIP-3alpha by normal human keratinocyte monolayers. Epidermal cells derived from foreskin were cultured in defined medium supplemented with different calcium levels (0.09, 0.8 and 1.7 mM) and were stimulated or not with the pro-inflammatory cytokines tumor necrosis factor-alpha (TNF-alpha 1-500 ng/ml) or interferon-gamma (INF-gamma 1-100 ng/ml). A high calcium concentration (1.7 mM) alone applied in culture medium for 4 days was sufficient to induce hBD-2 and hBD-3 mRNA expression. Whatever interindividual variability in the expression of hBD-2 and hBD-3 mRNA and MIP-3alpha secretion, the addition of TNF-alpha for a short duration (26h), initiated a dose-dependent and coordinated up-regulation of hBD-2 and hBD-3 mRNA and MIP-3alpha release in keratinocyte cultures. Unlike hBD-2 and hBD-3 mRNA was preferentially stimulated by IFN-gamma rather than TNF-alpha. In our experimental conditions, L-isoleucine, described to stimulate beta-defensin in bovine epithelial cells, did not exert any effect either on hBD-2 and hBD-3 transcripts or MIP-3alpha protein. Taken together, these results confirm the major role of the maturation/differentiation process of normal human keratinocytes in the induction of inducible beta-defensins and MIP-3alpha chemokine, which contribute in vivo to the immunosurveillance of the skin barrier function.
Pernet
I
I
INSERM U 36 Clinique Dermatologique, Hôpital E Herriot, Lyon, France.
Reymermier
C
C
Guezennec
A
A
Branka
J-E
JE
Guesnet
J
J
Perrier
E
E
Dezutter-Dambuyant
C
C
Schmitt
D
D
Viac
J
J
D002118:Calcium / Q000378:metabolism*; D018929:Cell Culture Techniques; D002478:Cells, Cultured; D054418:Chemokine CCL20; D019742:Chemokines, CC / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D000078404:Epidermal Cells; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D007532:Isoleucine / Q000494:pharmacology; D015603:Keratinocytes / Q000378:metabolism*; D019402:Macrophage Inflammatory Proteins / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012867:Skin / Q000378:metabolism; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D015854:Up-Regulation; D023083:beta-Defensins / Q000378:metabolism*
C108355:CCL20 protein, human; D054418:Chemokine CCL20; D019742:Chemokines, CC; C422950:DEFB103A protein, human; C106641:DEFB4A protein, human; D019402:Macrophage Inflammatory Proteins; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D023083:beta-Defensins; D007532:Isoleucine; D007371:Interferon-gamma; D002118:Calcium