Measuring tissue oxygenation in vivo is of interest in fundamental biological as well as medical applications. One minimally invasive approach to assess the oxygen partial pressure in tissue (pO2) is to measure the oxygen-dependent luminescence lifetime of molecular probes. The relation between tissue pO2 and the probes’ luminescence lifetime is governed by the Stern-Volmer equation. Unfortunately, virtually all oxygen-sensitive probes based on this principle induce some degree of phototoxicity. For that reason, we studied the oxygen sensitivity and phototoxicity of dichlorotris(1, 10-phenanthroline)-ruthenium(II) hydrate [Ru(Phen)] using a dedicated optical fiber–based, time-resolved spectrometer in the chicken embryo chorioallantoic membrane. We demonstrated that, after intravenous injection, Ru(Phen)’s luminescence lifetime presents an easily detectable pO2 dependence at a low drug dose (1 mg/kg) and low fluence (120 mJ/cm2 at 470 nm). The phototoxic threshold was found to be at 10 J/cm2 with the same wavelength and drug dose, i.e., about two orders of magnitude larger than the fluence necessary to perform a pO2 measurement. Finally, an illustrative application of this pO2 measurement approach in a hypoxic tumor environment is presented.
Tissular oxygen concentration plays a key role during photodynamic therapy (PDT). Therefore, monitoring its local oxygen partial pressure (pO2) may help predict and/or control the outcome of a PDT treatment. The first real-time, in vivo measurements of the pO2 in the chicken egg's chorioallantoic membrane, using the delayed fluorescence of photoactivable porphyrins (PAPs), including protoporphyrin IX (PpIX), as monitored with a dedicated optical, fiber-based, time-resolved spectrometer, are reported here. The formation of PAPs/PpIX, photosensitizers of extensive clinical use, was induced in the chicken egg's chorioallantoic membrane (CAM) with aminolevulinic acid. An excellent correlation between the vascular damage induced by PDT and the reduction in tissular pO2 is found. This study suggests that clinical measurement of the pO2 using the PAPs'/PpIX's delayed fluorescence (DF) may be used to individualize in real time the PDT light dose applied.
The concentration of oxygen and its rate of consumption are important factors playing a role in PDT and radiotherapy.
One of the methods for measuring the tissular oxygen partial pressure (pO2) is based on the use of luminophores
presenting an oxygen-dependent quenching of their phosphorescence. The time-resolved luminescence spectroscopy of
palladium (PdTCPP) or ruthenium (RuDPP) porphyrin complexes is used for this purpose. Unfortunately, these
porphyrin derivatives are phototoxic and leak rapidly out of the blood vessels, making them unsuitable for measuring
tissular and or intravascular pO2.
Therefore, this research aimed at developing and testing new biocompatible, non-phototoxic oxygen sensors based on
palladium complexes incorporated into oxygen permeable, polysaccharide-based nanoparticles appropriate for noninvasive
in situ and in vivo measurements of the pO2.
In vitro studies, performed with an optical fiber-based time-resolved spectrophotometer, showed that the incorporation of
such pO2 probes in nanovectors reduces their sensitivity to oxygen as well as their photobleaching by less than one order
of magnitude.
However, in vivo biocompatibility studies performed on the chick's embryo chorioallantoic membrane (CAM) model
demonstrated that the luminescence of those oxygen probes tends to be heterogeneously distributed within the
vasculature. In addition, these probes induce a 'clumping tendency', resulting in a more or less decreased viability of the
embryos.
The chicken embryo's chorioallantoic membrane (CAM) is widely used as an in vivo model to study the vascular effects
induced by agents administrated topically or intravenously. Hence, in the vascular plexus of this respiratory membrane,
angiogenic and anti-angiogenic agents, as well as phototoxic effects have been studied.
The main goal of this study was to characterize the capillary network of the CAM after topical administration of
dimethyl sulfoxid (DMSO), a frequently used solvent of lipophylic drugs, including potent anti-VEGF agents. The CAM
capillaries were observed between days 8 and 9 of the embryo development, with an epi-fluorescence microscope
equipped with a sensitive camera by intravenous injection of a fluorescent agent and a non-fluorescing absorber (in the
extra-embryonic cavity) to screen the tissue background fluorescence. The fluorescence images of the CAM vasculature
were then processed in order to obtain a skeleton of the vessels and capillaries. This was done to quantify descriptors
such as the number of branching points/mm2, the mean area value of the vessels network meshes, and the mean of the 3rd
quartile of the histogram of these meshes, were then extracted.
Our results demonstrate that the topical administration of an aqueous solution of 20 μl of DMSO at concentrations equal
or larger than 0.1% turned out to modify the capillary network morphology in a dose-dependent manner as compared to
the control (20 μl of 0.9% NaCl).
Conference Committee Involvement (1)
17th International Photodynamic Association World Congress
28 June 2019 | Cambridge, Massachusetts, United States
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