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The influence of intracellular mTHPC concentration upon photobleaching dynamics
M Atif1, M R Stringer2, J E Cruse-Sawyer3
1Department of Physical Sciences, University of Hull, Cottingham Road, Hull HU6 7RX, UK.
Abstract:
In this study, we report the effect of the local photosensitizer concentration upon the dynamics of the singlet oxygen-mediated photobleaching, within formalin-fixed keratinocytes. Although the cells were incubated at a single mTHPC dose, cell-to-cell variations in concentration were defined within the perinuclear region by differences in the initial amplitude of the laser-induced fluorescence emission, located around 652nm. At a fixed laser fluence-rate, it was found that the photobleaching, when plotted as a function of delivered light dose, proceeded more rapidly at higher drug concentration. The mTHPC spectral emission profile is shown to be approximately Lorentzian and remains unchanged as the photobleaching proceeds. This indicates that there is no perturbation of the detected signal due to the inner-filter effect.
Insights
Higher concentrations of the photosensitizer mTHPC (meta-tetrahydroxyphenylchlorin) accelerate singlet oxygen-mediated photobleaching in keratinocytes. The drug concentration directly impacts photobleaching speed, independent of spectral changes.
Area of Science:
- Photodynamic Therapy
- Cell Biology
- Biophysics
Background:
- Photobleaching is a critical factor in photodynamic therapy efficacy.
- Understanding photosensitizer concentration effects is key to optimizing treatment.
- Keratinocytes are relevant cells for skin-related photodynamic applications.
Purpose of the Study:
- To investigate the relationship between local photosensitizer concentration and photobleaching dynamics.
- To determine how variations in mTHPC concentration affect singlet oxygen-mediated photobleaching in keratinocytes.
- To assess the impact of drug concentration on photobleaching rates at a fixed laser fluence-rate.
Main Methods:
- Utilizing formalin-fixed keratinocytes incubated with mTHPC.
- Measuring cell-to-cell variations in mTHPC concentration via laser-induced fluorescence at 652nm.
- Analyzing photobleaching rates as a function of delivered light dose at a fixed laser fluence-rate.
- Characterizing the mTHPC spectral emission profile during photobleaching.
Main Results:
- Photobleaching proceeded more rapidly at higher local mTHPC concentrations.
- The mTHPC spectral emission profile remained Lorentzian and unchanged during photobleaching.
- No inner-filter effect was observed, indicating the detected signal was not perturbed.
Conclusions:
- Local photosensitizer concentration is a significant determinant of photobleaching dynamics.
- Higher mTHPC concentrations lead to faster photobleaching in keratinocytes.
- The photobleaching process in this model is not affected by the inner-filter effect, simplifying analysis.
