Influence of 5-aminolevulinic acid and red light on collagen metabolism of human dermal fibroblasts.
Patients with localized scleroderma receiving topical photodynamic therapy with 5-aminolevulinic acid show a reduction in skin tightness, suggesting that this therapy reduces skin sclerosis. To investigate potential mechanisms, the effects of 5-aminolevulinic acid and light on collagen metabolism were studied in vitro. Normal and scleroderma fibroblasts were treated with sublethal doses of 5-aminolevulinic acid and red light and transferred to three-dimensional collagen lattices. Cell supernatants were taken 6-72 h after photodynamic therapy to determine protein levels of the matrix metalloproteinases 1, 2, and 3, and of their inhibitors, tissue inhibitor of metalloproteinase 1 and 2 by enzyme-linked immunosorbent assay. Cellular mRNA expression of these proteins and of collagen type I and III was measured by quantitative real-time polymerase chain reaction. A significant, time-dependent induction of matrix metalloproteinase 1 (up to 2.4-fold after 48 h) and matrix metalloproteinase 3 (up to 4.3-fold after 48 h) protein levels was seen after 5-aminolevulinic acid-photodynamic therapy. Irradiation with ultraviolet A light, used as a positive control, showed a similar induction of matrix metalloproteinase 1 (2.3-fold after 48 h). The mRNA levels of matrix metalloproteinase 1 and matrix metalloproteinase 3 were significantly increased 12 h after irradiation, whereas collagen type I mRNA was strongly decreased already 6 h following irradiation. Collagen type III, tissue inhibitor of metalloproteinase 1, and matrix metalloproteinase 2 did not change after photodynamic therapy. Addition of nontoxic concentrations of sodium azide, a singlet-oxygen quencher, significantly inhibited induction of matrix metalloproteinase 1 by 5-aminolevulinic acid and light. These data show that 5-aminolevulinic acid and light induce matrix metalloproteinase 1 and matrix metalloproteinase 3 expression in normal and scleroderma fibroblasts in a singlet oxygen-dependent way while reducing collagen type I mRNA expression. Induction of collagen-degrading enzymes together with reduction of collagen production might be responsible for the anti-sclerotic effects of 5-aminolevulinic acid-photodynamic therapy observed in vivo.
Karrer
Sigrid
S
Department of Dermatology, University of Regensburg, Regensburg, Germany. sigrid.karrer@klinik.uni-regensburg.de
Bosserhoff
Anja Kathrin
AK
Weiderer
Petra
P
Landthaler
Michael
M
Szeimies
Rolf-Markus
RM
D000622:Aminolevulinic Acid / Q000494:pharmacology*; D002478:Cells, Cultured; D024042:Collagen Type I / Q000378:metabolism*; D024061:Collagen Type III / Q000378:metabolism; D020405:Dermis / Q000166:cytology* / Q000187:drug effects / Q000528:radiation effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000528:radiation effects; D006801:Humans; D008027:Light; D020781:Matrix Metalloproteinase 1 / Q000235:genetics / Q000378:metabolism; D020778:Matrix Metalloproteinase 2 / Q000235:genetics / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000235:genetics / Q000378:metabolism; D017319:Photosensitizing Agents / Q000494:pharmacology*; D012333:RNA, Messenger / Q000032:analysis; D012594:Scleroderma, Localized / Q000378:metabolism; D019810:Sodium Azide / Q000494:pharmacology; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics / Q000378:metabolism; D019716:Tissue Inhibitor of Metalloproteinase-2 / Q000235:genetics / Q000378:metabolism
D024042:Collagen Type I; D024061:Collagen Type III; D004791:Enzyme Inhibitors; D017319:Photosensitizing Agents; D012333:RNA, Messenger; D019715:Tissue Inhibitor of Metalloproteinase-1; D019716:Tissue Inhibitor of Metalloproteinase-2; D000622:Aminolevulinic Acid; D019810:Sodium Azide; D019278:Matrix Metalloproteinase 3; D020778:Matrix Metalloproteinase 2; D020781:Matrix Metalloproteinase 1