1. Influence of 5-Aminolevulinic Acid and Red Light on Collagen Metabolism of Human Dermal Fibroblasts 
Sigrid Karrer, Anja Kathrin Bosserhoff, Petra Weiderer, Michael Landthaler, Rolf-Markus Szeimies 
Journal of Investigative Dermatology 
Volume 120, Issue 2, Pages (February 2003)
DOI: /j x
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

2. Figure 1
Time-dependent regulation of protein levels of MMP-1. Results of the MMP-1 ELISA from cell culture supernatants of normal fibroblasts (A) and of scleroderma fibroblasts (B) treated with ALA (75 μmol per liter) and light (40 mW per cm2, 24 J per cm2) and examined at different time points (6–72 h) following irradiation are shown. The x-fold change of protein levels of MMP-1 (black bars) and active MMP-1 as measured by an activity assay (dark gray bars) in cell supernatants of fibroblasts after treatment with ALA and light are shown as compared with the untreated control (the control is set=1 at each measured time point, controls not shown). Fibroblasts were also irradiated with sublethal doses of UVA light (10 J per cm2) as a positive regulator of MMP-1 production in this cell type. The x-fold change of total MMP-1 after irradiation with UVA light is illustrated by the light gray bars. Fibroblasts treated with red light only or ALA only did not show statistically significant changes as compared with the untreated controls and are therefore not shown. Data are mean±SEM from triplicate determinations of at least three independent experiments for each case. *A significant difference compared with the untreated control (p<0.05).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

3. Figure 2
Time-dependent regulation of MMP-3 protein levels. Results of the MMP-3 ELISA from cell culture supernatants of normal fibroblasts (black bars) and scleroderma fibroblasts (dark gray bars) treated with ALA (75 μmol per liter) and light (40 mW per cm2, 24 J per cm2) and examined at different time points (6–72 h) following irradiation are shown. Data represent fold increase of MMP-3 protein levels over the untreated control (control is set=1, controls not shown). Fibroblasts treated with light or ALA alone did not show statistically significant changes as compared with the untreated controls and are therefore not shown. Data are mean±SEM from triplicate determinations of at least three independent experiments for each case. There was no statistically significant difference between normal and scleroderma fibroblasts. *A significant difference compared with the untreated control (p<0.05).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

4. Figure 3
(MMP-1 (A) and MMP-3 (B) mRNA levels after quantitative real-time PCR using the LightCycler. Normal human fibroblasts were treated with ALA (75 μmol per liter) and light (40 mW per cm2, 24 J per cm2) and subjected to reverse transcriptase–PCR at different time points following irradiation (gray bars). Change of MMP-1 and MMP-3 mRNA as compared with the untreated control (black bars) is expressed in arbitrary units, as quantitative mRNA levels of fibroblasts prior to PDT were often beyond detection limits. Fibroblasts treated with light or ALA alone did not show statistically significant changes as compared with the untreated controls and are therefore not shown. Data are mean±SEM from duplicate determinations of three independent experiments for each case. Scleroderma fibroblasts showed the same results as compared with normal fibroblasts and are therefore not shown. *A significant difference compared with the untreated control (p<0.05).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

5. Figure 4
Collagen type I mRNA levels after quantitative real-time PCR using the LightCycler. Normal human fibroblasts (black bars) and scleroderma fibroblasts (dark gray bars) were treated with ALA (75 μmol per liter) and light (40 mW per cm2, 24 J per cm2) and subjected to reverse transcriptase–PCR at different time points (12–72 h) following irradiation. Data represent the fold decrease of collagen type I mRNA over the untreated control (control is set=1, controls are represented by the black bars). Collagen mRNA levels were reduced significantly already 12 h following PDT showing a maximal reduction at 24 h after PDT. Fibroblasts treated with light or ALA alone did not show statistically significant changes as compared with the untreated controls and are therefore not shown. Data are mean±SEM from duplicate determinations of three independent experiments for each case. *A significant difference as compared with the untreated control (p<0.05).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions

6. Figure 5
Effect of the singlet oxygen quencher sodium azide on MMP-1 induction after incubation of normal fibroblasts (A) and scleroderma fibroblasts (B) with ALA (75 μmol per liter) and addition of sodium azide (50 mmol per liter) prior to irradiation with red light (40 mW per cm2, 24 J per cm2). Untreated controls are set=1 (data not shown), dark gray bars show MMP-1 induction after PDT and light gray bars show MMP-1 protein levels after PDT plus addition of sodium azide during irradiation with red light. Addition of sodium azide to otherwise untreated fibroblasts did not alter cell viability nor MMP-1 expression as compared with the untreated controls. PDT-induced MMP-1 increase was significantly inhibited by sodium azide, indicating that singlet oxygen is an important mediator of MMP-1 expression. *A significant difference between fibroblasts treated with PDT and those treated with PDT and sodium azide (p<0.05).
Journal of Investigative Dermatology  , DOI: ( /j x)
Copyright © 2003 The Society for Investigative Dermatology, Inc Terms and Conditions