Time-resolved singlet oxygen luminescence detection under photodynamic therapy relevant conditions: comparison of ex vivo application of two photosensitizer formulations

Abstract. Singlet oxygen plays a crucial role in photo-dermatology and photodynamic therapy (PDT) of cancer. Its direct observation by measuring the phosphorescence at 1270 nm, however, is still challenging due to the very low emission probability. It is especially challenging for the time-resolved detection of singlet oxygen kinetics in vivo which is of special interest for biomedical applications. Photosensitized generation of singlet oxygen, in pig ear skin as model for human skin, is investigated here. Two photosensitizers (PS) were topically applied to the pig ear skin and examined in a comparative study, which include the amphiphilic pheophorbide-a and the highly hydrophobic perfluoroalkylated zinc phthalocyanine (F64PcZn). Fluorescence microscopy indicates the exclusive accumulation of pheophorbide-a in the stratum corneum, while F64PcZn can also accumulate in deeper layers of the epidermis of the pig ear skin. The kinetics obtained with phosphorescence measurements show the singlet oxygen interaction with the PS microenvironment. Different generation sites of singlet oxygen correlate with the luminescence kinetics. The results show that singlet oxygen luminescence detection can be used as a diagnostic tool, not only for research, but also during treatment. The detection methodology is suitable for the monitoring of chemical quenchers’ oxidation as well as O2 saturation at singlet oxygen concentration levels relevant to PDT treatment protocols.

[1]  B. Röder,et al.  Determination of the electron transfer parameters of a covalently linked porphyrin-quinone with mesogenic substituents — optical spectroscopic studies in solution , 1997 .

[2]  B. Pogue,et al.  A Singlet Molecular Oxygen Imaging Sensor for Photodynamic Therapy , 2008 .

[3]  P. Mullineaux,et al.  Imaging the production of singlet oxygen in vivo using a new fluorescent sensor, Singlet Oxygen Sensor Green. , 2006, Journal of experimental botany.

[4]  J. Kennedy,et al.  Photodynamic therapy with endogenous protoporphyrin IX: basic principles and present clinical experience. , 1990, Journal of photochemistry and photobiology. B, Biology.

[5]  S. Hackbarth,et al.  A new benchmark for time-resolved detection of singlet oxygen luminescence – revealing the evolution of lifetime in living cells with low dose illumination , 2009 .

[6]  Michael S Patterson,et al.  Direct Near-infrared Luminescence Detection of Singlet Oxygen Generated by Photodynamic Therapy in Cells In Vitro and Tissues In Vivo¶ , 2002, Photochemistry and photobiology.

[7]  L. Norlén.  Stratum corneum keratin structure, function and formation – a comprehensive review , 2006, International journal of cosmetic science.

[8]  P. Wertz,et al.  Molecular models of the intercellular lipid lamellae from epidermal stratum corneum. , 2003, Biochimica et biophysica acta.

[9]  R. Warner,et al.  Electron probe analysis of human skin: determination of the water concentration profile. , 1988, The Journal of investigative dermatology.

[10]  Hidetake Imasato,et al.  A combination of techniques to evaluate photodynamic efficiency of photosensitizers , 2008 .

[11]  Reinhard Schmidt,et al.  Physical Mechanisms of Generation and Deactivation of Singlet Oxygen , 2003 .

[12]  Hyun-jin Lee,et al.  Enhanced Acidity, Photophysical Properties and Liposome Binding of Perfluoroalkylated Phthalocyanines Lacking C-H Bonds , 2006, Photochemistry and photobiology.

[13]  S. Nonell,et al.  Kinetics of singlet oxygen photosensitization in human skin fibroblasts. , 2008, Free radical biology & medicine.

[14]  J. Thiele.  Oxidative Targets in the Stratum corneum , 2001, Skin Pharmacology and Physiology.

[15]  R. Tyrrell,et al.  UVA radiation-induced oxidative damage to lipids and proteins in vitro and in human skin fibroblasts is dependent on iron and singlet oxygen. , 1995, Free radical biology & medicine.

[16]  Michael S Patterson,et al.  Imaging of Photodynamically Generated Singlet Oxygen Luminescence In Vivo¶ , 2005, Photochemistry and photobiology.

[17]  Comparative study of the photosensitization of Jurkat cells in vitro by pheophorbide-a and a pheophorbide-a diaminobutane poly-propylene-imine dendrimer complex , 2003 .

[18]  J. Fuchs,et al.  Measurements of UV-generated free radicals/reactive oxygen species (ROS) in skin. , 2006, Spectrochimica acta. Part A, Molecular and biomolecular spectroscopy.

[19]  Michael Landthaler,et al.  Direct detection of singlet oxygen generated by UVA irradiation in human cells and skin. , 2007, The Journal of investigative dermatology.

[20]  B. Ehrenberg,et al.  Kinetics and Yield of Singlet Oxygen Photosensitized by Hypericin in Organic and Biological Media , 1998, Photochemistry and photobiology.

[21]  D. Wöhrle,et al.  Rational design of a reactive yet stable organic-based photocatalyst. , 2009, Dalton transactions.

[22]  S. Lee,et al.  A singlet oxygen monitor as an in vivo photodynamic therapy dosimeter , 2009, World Congress of the International Photodynamic Association.

[23]  S. White,et al.  Structure of lamellar lipid domains and corneocyte envelopes of murine stratum corneum. An X-ray diffraction study. , 1988, Biochemistry.

[24]  M. Landthaler,et al.  Time-resolved investigations of singlet oxygen luminescence in water, in phosphatidylcholine, and in aqueous suspensions of phosphatidylcholine or HT29 cells. , 2005, The journal of physical chemistry. B.

[25]  L. Klotz,et al.  Contribution of UVB and UVA to UV-dependent stimulation of cyclooxygenase-2 expression in artificial epidermis , 2004, Photochemical & photobiological sciences : Official journal of the European Photochemistry Association and the European Society for Photobiology.

[26]  J. Søndergaard,et al.  Heat-separation of normal human skin for epidermal and dermal prostaglandin analysis , 2004, Archives of Dermatological Research.

[27]  Jan C. Schlothauer,et al.  New insights to primary photodynamic effects--Singlet oxygen kinetics in living cells. , 2010, Journal of photochemistry and photobiology. B, Biology.

[28]  J. Johnson,et al.  Activation of transcription factor AP-2 mediates UVA radiation- and singlet oxygen-induced expression of the human intercellular adhesion molecule 1 gene. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[29]  Thiele Jj.  Oxidative targets in the stratum corneum. A new basis for antioxidative strategies. , 2001 .

[30]  Esben Skovsen,et al.  Optical detection of singlet oxygen from single cells. , 2006, Physical chemistry chemical physics : PCCP.

[31]  J. Regensburger,et al.  UVA and endogenous photosensitizers — the detection of singlet oxygen by its luminescence , 2012, Photochemical & Photobiological Sciences.

[32]  M. Baptista,et al.  Generation and suppression of singlet oxygen in hair by photosensitization of melanin. , 2011, Free radical biology & medicine.

[33]  R. Dědic,et al.  Singlet oxygen quenching by oxygen in tetraphenyl-porphyrin solutions , 2006 .

[34]  A. Goetz,et al.  Strong and prolonged induction of c-jun and c-fos proto-oncogenes by photodynamic therapy. , 1996, British Journal of Cancer.

[35]  F. Cui,et al.  Mitochondria-involved apoptosis induced by MPPa mediated photodynamic therapy , 2008 .

[36]  M. Landthaler,et al.  Theoretical and experimental analysis of the luminescence signal of singlet oxygen for different photosensitizers. , 2007, Journal of photochemistry and photobiology. B, Biology.

[37]  A. Krasnovsky,et al.  Photophysical studies of pheophorbide a and pheophytin a. Phosphorescence and photosensitized singlet oxygen luminescence. , 1990, Journal of photochemistry and photobiology. B, Biology.