Photodynamic therapy (PDT) using protoporpyrin IX (PpIX) precursors like 5-aminolevulinic acid (ALA) or methyl-aminolevulinate (MAL) has shown to be effective in the treatment of various skin diseases. Using ALA we have shown in numerous studies a significantly improved efficacy by applying light fractionation with a long dark interval. In contrast, in the hairless mouse model, the PDT efficacy using MAL is unaffected by adopting this approach. More acute edema is found after ALA-PDT suggesting a difference in response of endothelial cells to PDT. To investigate the role of endothelial cells, cryo-sections of hairless mouse skin after 4 hours of topical MAL or ALA application were stained with a fluorescent endothelial cell marker (CD31). Co-localization of this marker with the PpIX fluorescence was performed using the spectral imaging function of the confocal microscope. We have also used intra-vital confocal microscopy to image the PpIX fluorescence distribution in correlation with the vasculature of live mouse skin. Our results show PpIX fluorescence at depth in cryo-sections of mouse skin after 4 hours of topical application. Co-localization has shown to be difficult due to the changes in tissue organization caused by the staining procedure. As expected we found high PpIX fluorescence levels in the epidermis after both MAL and ALA application using intra-vital microscopy. After ALA application more PpIX fluorescence was found deep in the dermal layer of the skin than after MAL. Furthermore we detected localized fluorescence in unidentified structures that could not be correlated to blood vessels or nerves.

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Keywords ALA, in-vivo microscopy, MAL, PDT, PpIX, vasculature
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Conference 12th World Congress of the International Photodynamic Association - Photodynamic Therapy: Back to the Future
de Bruijn, H.S, de Vijlder, H.C, de Haas, E.R.M, van der Ploeg-van den Heuvel, A, Kruijt, B, Poel-Dirks, D, … Robinson, D.J. (2009). Light fractionation increases the efficacy of ALA-PDT but not of MAL-PDT: What is the role of (vascular) endothelial cells?. Presented at the 12th World Congress of the International Photodynamic Association - Photodynamic Therapy: Back to the Future. doi:10.1117/12.822973