Photodynamic Therapy Dosimetry Research Articles

Doppler optical coherence tomography monitoring of microvascular tissue response during photodynamic therapy in an animal model of Barrett's esophagus

Doppler optical coherence tomography (DOCT) is an imaging modality that allows assessment of the microvascular response during photodynamic therapy (PDT) and may be a powerful tool for treatment monitoring/optimization in conditions such as Barrett's esophagus (BE). To assess the technical feasibility of catheter-based intraluminal DOCT for monitoring the microvascular response during endoluminal PDT in an animal model of BE. Thirteen female Sprague-Dawley rats underwent esophagojejunostomy to induce enteroesophageal reflux for 35 to 42 weeks and the formation of Barrett's mucosa. Of these, 9 received PDT by using the photosensitizer Photofrin (12.5 mg/kg intravenous), followed by 635-nm intraluminal light irradiation 24 hours after drug administration. The remaining 4 surgical rats underwent light irradiation without Photofrin (controls). Another group of 5 normal rats, without esophagojejunostomy, also received PDT. DOCT imaging of the esophagus by using a catheter-based probe (1.3-mm diameter) was performed before, during, and after light irradiation in all rats. Distinct microstructural differences between normal squamous esophagus, BE, and the transition zone between the 2 tissues were observed on DOCT images. Similar submucosal microcirculatory effects (47%-73% vascular shutdown) were observed during PDT of normal esophagus and surgically induced BE. Controls displayed no significant microvascular changes. No apparent difference was observed in the PDT-induced vascular response between normal rat esophagus and the BE rat model. Real-time monitoring of PDT-induced vascular changes by DOCT may be beneficial in optimizing PDT dosimetry in patients undergoing this therapy for BE and other conditions.

Photodynamic Therapy Reduces Intimal Hyperplasia in Prosthetic Vascular Bypass Grafts in a Pig Model

Bypass surgery has a failing frequency of 30% during the first year, mainly due to intimal hyperplasia (IH). This negative effect is most pronounced in artificial grafts. Photodynamic therapy (PDT) is a technique in which light activates photosensitizer dyes to produce free-radicals resulting in an eradication of cells in the vascular wall. The aim of this study was to determine the effectiveness of PDT to reduce IH in a preclinical porcine PTFE bypass model. Ten pigs were used. After a pilot PDT dosimetry study (n=3) PTFE grafts were bilaterally placed into the circulation as bypasses from the common to the external iliac arteries (n=7). The right sides served as controls (C). Before implantation of the left grafts, the arterial connecting sites of the left distal anastomoses were PDT-treated. The arteries were pressurized at 180 mmHg for 5 minutes with the photosensitizer Methylene Blue (330 microg/ml), and thereafter endoluminally irradiated with laser light (lambda = 660 nm, 100 mW/cm(2), 150 J/cm(2)). After 4 weeks the specimens were retrieved and formalin fixed. Cross sections through the midportions of the distal anastomoses and the grafts were used for histology, immunohistochemistry to identify inflammatory cells and morphometric evaluation (n=7). No systemic side effects and no graft occlusions were noted. PDT-treated anastomoses showed reduced IH in the mid-portions of the anastomoses (Area of IH: microm(2)/microm graft: C: 6970+/-1536, PDT: 2734+/-2560; P<0.005) as well as in the grafts (C: 5391+/-4031, PDT: 777+/-1331; P<0.02). The number of inflammatory cells per microscopic field was increased after PDT (C: 24+/-16, PDT: 37+/-15; P<0.009). Adjuvant PDT, performed in an endovascular fashion, was a safe method to reduce prosthetic graftstenosis in a preclinical setting. This study underscores the clinical potential of PDT to inhibit the development of clinical bypass graftstenosis.

Monitoring Photoproduct Formation and Photobleaching by Fluorescence Spectroscopy Has the Potential to Improve PDT Dosimetry with a Verteporfin-like Photosensitizer¶

In current clinical practice, photodynamic therapy (PDT) is carried out with prescribed drug doses and light doses as well as fixed drug–light intervals and illumination fluence rates. This approach can result in undesirable treatment outcomes of either overtreatment or undertreatment because of biological variations between different lesions and patients. In this study, we explore the possibility of improving PDT dosimetry by monitoring drug photobleaching and photoproduct formation. The study involved 60 mice receiving the same drug dose of a novel verteporfin-like photosensitizer, QLT0074, at 0.3 mg/kg body weight, followed by different light doses of 5, 10, 20, 30, 40 or 50 J/cm2 at 686 nm and a fluence rate of 70 mW/cm2. Photobleaching and photoproduct formation were measured simultaneously, using fluorescence spectroscopy. A ratio technique for data processing was introduced to reliably detect the photoproduct formed by PDT on mouse skin in vivo. The study showed that the QLT0074 photoproduct is stable and can be reliably quantified. Three new parameters, photoproduct score (PPS), photobleaching score (PBS) and percentage photobleaching score (PBS%), were introduced and tested together with the conventional dosimetry parameter, light dose, for performance on predicting PDT-induced outcome, skin necrosis. The statistical analysis of experimental results was performed with an ordinal logistic regression model. We demonstrated that both PPS and PBS improved the prediction of skin necrosis dramatically compared to light dose. PPS was identified as the best single parameter for predicting the PDT outcome.

Modeling light fluence rate distribution in optically heterogeneous prostate photodynamic therapy using a kernel method.

To accurately calculate light fluence rate distribution for light dosimetry in prostate photodynamic therapy (PDT), heterogeneity of optical properties has to be taken into account. Previous study has shown that a finite-element method (FEM) can be an efficient tool to deal with the optical heterogeneity. However, the calculation speed of the FEM is not suitable for real time treatment planning. In this paper, two kernel models are developed. Because the kernels are based on analytic solutions of the diffusion equation, calculations are much faster. We derived our extensions of kernel from homogeneous medium to heterogeneous medium assuming spherically symmetrical heterogeneity of optical properties. The kernel models are first developed for a point source and then extended for a linear source, which is considered a summation of point sources uniformly spaced along a line. The kernel models are compared with the FEM calculation. In application of the two kernel models to a heterogeneous prostate PDT case, both kernel models give improved light fluence rate results compared with those derived assuming homogeneous medium. In addition, kernel model 2 predicts reasonable light fluence rates and is deemed suitable for treatment planning.

A method to improve reconstruction of the distribution of hemoglobin, oxygenation, and MLu concentration in the human prostate before and after photodynamic therapy.

Explicit dosimetry of photodynamic therapy requires detailed knowledge of the light, drug, and oxygenation distributions within the target tissue. We present a method for the optical detection and three-dimensional reconstruction of hemoglobin concentration and oxygenation and sensitizer concentration within the human prostate. Spectrally resolved diffuse transmission measurements were made using a small isotropic fiber-based white light source and an isotropic detector inserted into the prostate via parallel closed transparent catheters. The spectra were modeled using the diffusion approximation appropriate for infinite media. The optical absorption of the prostate was assumed to be a linear combination of the absorption spectra of oxy- and deoxyhemoglobin and MLu, and the scattering was assumed to be of the form A(λ/λ0)-b. The separation of absorption and scattering coefficients was accomplished based on the spectral shape of the diffuse transmission, rather than the spatial variation in intensity. By making multiple measurements at various source-detector separations, we investigate the signal-to-noise sensitivity of our algorithm. In addition, the redundancy in our source-detector position matrix creates several positions in which the tissue parameters can be reconstructed from multiple independent measurements, allowing an assessment of the repeatability of the algorithm. We find significant heterogeneity in the reconstructed optical properties; however the recovery of spectrally consistent absorption and scattering spectra is improved compared to wavelength-wise reconstruction algorithms.

Singlet Oxygen Luminescence Dosimetry (SOLD) for Photodynamic Therapy: Current Status, Challenges and Future Prospects

As photodynamic therapy (PDT) continues to develop and find new clinical indications, robust individualized dosimetry is warranted to achieve effective treatments. We posit that the most direct PDT dosimetry is achieved by monitoring singlet oxygen (1O2), the major cytotoxic species generated photochemically during PDT. Its detection and quantification during PDT have been long-term goals for PDT dosimetry and the development of techniques for this, based on detection of its near-infrared luminescence emission (1270 nm), is at a noteworthy stage of development. We begin by discussing the theory behind singlet-oxygen luminescence dosimetry (SOLD) and the seminal contributions that have brought SOLD to its current status. Subsequently, technology developments that could potentially improve SOLD are discussed, together with future areas of research, as well as the potential limitations of this method. We conclude by examining the major thrusts for future SOLD applications: as a tool for quantitative photobiological studies, a point of reference to evaluate other PDT dosimetry techniques, the optimal means to evaluate new photosensitizers and delivery methods and, potentially, a direct and robust clinical dosimetry system.

Light distribution and calibration of commercial PDT LED arrays

Commercial photodynamic therapy (PDT) light sources are supplied with no evidence of traceability to national measurements standards to validate indicated delivered dose. Also, the spatial distribution of the radiant energy is not disclosed. This means that there is no way for the user to be able to verify that the required dose is being delivered to the appropriate area of the lesion. To this end, a simple method, traceable to national standards, is described and applied to an investigation of two commercial LED arrays. In one case, the dose fell to 38% of that received at the central area at a distance of only 2 cm. In the other, the output was more uniform over a larger area but the maximum irradiance was not at the centre of the field. All of these inhomogenieties were taken into account when the actual light dose delivered to the patient was calculated in the method described. This ensured transparent traceability in PDT dosimetry.

O-025 Tumor ablation through self-expandable metal stent (SEMS) forairway neoplasm: The dosimetry and efficacy of photodynamic therapy

O-025 Tumor ablation through self-expandable metal stent (SEMS) forairway neoplasm: The dosimetry and efficacy of photodynamic therapy

TH-A-T-6C-01: Photodynamic Therapy: Fundamentals and Dosimetry

Photodynamic therapy (PDT) is an emerging cancer treatment modality based on the interaction of light, a photosensitizing drug, and oxygen. PDT has been approved by the US Food and Drug Administration for the treatment of microinvasive lung cancer, obstructing lung cancer, and obstructing esophageal cancer. Studies have shown some efficacy in the treatment of a variety of malignant and premalignant conditions including head and neck cancer, lung cancer, mesothelioma, Barrett's esophagus, prostate, and brain tumors. Unlike radiation therapy, PDT is a non-ionizing radiation that can be used repeatedly without cumulative long-term complications since it does not appear to target DNA. Various photosensitizer drugs have been developed. Most of the drugs are so called Type II photosensitizers, where the active agent of phototoxicity is induced by the production of singlet oxygen or active oxygen derivatives through light activation. The first-generation photosensitizer, haematoporphyrin derivative (HPD), is a mixture of porphyrin monomers and oligomers that is partially purified to produce the commercially available product, Photofrin®. HPD is photochemically activated by the absorption of tissue-penetrating light at λ 630 nm (red) and is the only FDA approved photosensitizer. HPD-mediated PDT has several clinical disadvantages, including prolonged skin photosensitivity (4 weeks), relatively low quantum yield of singlet oxygen, and a limited depth of associated tissue damage of 2–5 mm. Second generation photosensitizers (e.g., mTHPC, TOOKAD, and MLu) overcome these shortcomings with higher quantum yield, lower skin toxicities, and deeper penetration. There has been tremendous progress in photodynamic therapy dosimetry. The simplest clinical light dose prescription is to quantify the incident fluence for patients treated with a given photosensitizer injection per body weight. However, light dose given in this way do not take into account the light scattering by tissue and usually underestimate light fluence rate. Techniques have been developed to characterize the tissue optical properties and the light fluence rate in-vivo. Other optical spectroscopic methods have been developed to characterize tissue absorption and scattering properties, which in term provide information about tissue oxygenation and drug concentration. Fluorescence techniques can be used to quantify drug concentration and potentially photobleaching rate of photosensitizers. The objective of this course is to present a brief review of the issues related to the application of photodynamic therapy. In particular, we review the current start of art of techniques to quantify light fluence, drug concentration, and tissue oxygenations, and PDT outcome. Educational Objectives: 1. To explain the basic principle of photodynamic therapy. 2. To discuss the various photosensitizer drugs. 3. To review techniques for PDT light delivery for surface and interstitial applications. 4. To review current state of art in PDT in-vivo light dosimetry, light-tissue interaction, and the light fluence distributions in turbid medium. 5. To review PDT dosimetry techniques to characterize tissue optical properties, drug concentration, tissue oxygen concentration, and PDT efficacy.

Optimization of light dosimetry for photodynamic therapy of Barrett's esophagus: efficacy vs. incidence of stricture after treatment

Photodynamic therapy (PDT) may be used to ablate high-grade dysplasia/early stage cancer (HGD/T1) in patients with Barrett's esophagus. PDT may result in esophageal stricture. This nonrandomized, unblinded, dose de-escalation study in consecutive patients was designed to determine the lowest light dose effective for ablation of HGD/T1 while reducing the incidence of stricture. A total of 113 patients received an injection of porfimer sodium (2 mg/kg). Three days later, 630 nm light was delivered by using a 20-mm-diameter PDT balloon at doses of 115 J/cm (n=59), 105 J/cm (n=18), 95 J/cm (n=17), or 85 J/cm (n=19). Treatment efficacy was determined by obtaining biopsy specimens of the treated area 3 months later. The incidence of stricture was determined by the need for esophageal dilation to treat dysphagia. A stricture was considered severe if 6 or more dilations were required. The incidence of severe stricture was related to the light dose. At 115 J/cm, 15.3% of patients developed severe strictures compared with 5.3% to 5.6% of those treated with the lower doses. At a light dose of 115 J/cm, 17.0% of patients had residual HGD/T1. Light doses of 105 J/cm, 95 J/cm, and 85 J/cm resulted in residual HGD/T1 in 33.3%, 29.4%, and 31.6% of patients, respectively. None of the observations were statistically significant. Decreasing the light dose below 115 J/cm appeared to result in a reduced incidence rate of severe stricture but higher relative frequencies of residual HGD/T1 in Barrett's esophagus.

Photodynamic therapy on keloid fibroblasts in tissue-engineered keratinocyte-fibroblast co-culture

Keloids are disfiguring, proliferative scars that are a pathologic response to cutaneous injury. An organotypic tissue culture system (the Raft model 1-10) was used to investigate the feasibility of using photodynamic therapy (PDT) as an adjunctive therapy to treat keloids following surgical excision. The Raft co-culture system mimics skin by layering keratinocytes on top of fibroblasts embedded in a collagen matrix. PDT uses drugs that produce singlet oxygen in situ when irradiated by light, and may lead to a number of effects in living tissues varying from the modulation of growth to apoptosis. PDT is already used to treat several benign and malignant diseases in organs such as the skin, retina, and esophagus. Normal adult, neonatal, and keloid fibroblasts and keratinocytes were isolated from skin obtained from patients undergoing elective procedures and used to construct the Rafts. Mature Rafts (after 4 days) were incubated with 5-amino levulinic acid (5-ALA), a photosensitizer, for 3 hours and were laser-irradiated (635 nm) for total energy delivery of 5 J/cm2, 10 J/cm2, or 20 J/cm2. Rafts were examined 24 hours and 14 days later. Cell viability was determined using confocal imaging combined with live-dead fluorescent dyes. Multi-photon microscope (MPM) imaged collagen structure and density. As Rafts contract over time, surface area was measured using optical micrometry daily. At 10 and 20 J/cm2, near-total cell death was observed in all constructs, while at 5 J/cm2 cell viability was comparable to controls. Cell viability in keloid and neonatal Rafts was greater than that observed in normal adult Rafts. Treated Rafts contracted less over the 14-day period compared to controls. Contraction and collagen density were greatest in keloid and neonatal Rafts. A PDT dosimetry range was established, which reduces tissue contraction and collagen density while minimizing injury to fibroblasts.

Photodynamic therapy on keloid fibroblasts in engineered keratinocyte-fibroblast co-culture

Objectives: Keloids are disfiguring proliferative scars that are a pathologic response to cutaneous trauma. An organotypic tissue culture system (the RAFT model) was used to investigate the feasibility of treating keloids using photodynamic therapy (PDT). The RAFT co-culture recreates skin by layering keratinocytes on top of fibroblasts embedded in a collagen matrix. PDT uses drugs that produce singlet oxygen in situ when irradiated by light and may lead to a number of effects in living tissues varying from the modulation of growth to apoptosis. Methods: Normal adult, neonatal, and keloid fibroblasts and keratinocytes were used to construct the RAFTs. Mature RAFTS were incubated with 5-ALA, a photosensitizer, and were laser-irradiated (635 nm) for energy delivery of 5 J/cm2, 10 J/cm2, or 20 J/cm2. RAFTS were examined 24 hours and 14 days later. Cell viability was determined using confocal imaging combined with live-dead fluorescent dyes. Multiphoton microscopy (MPM) imaged collagen structure and density. As RAFTs contracted over time, surface area was measured using optical micrometry daily. Results: At 20 J/cm2, near-total cell death was observed in all constructs. At 10 J/cm2 some cell viability was maintained, while at 5 J/cm2 cell viability was comparable to controls. After 14 days, cell viability in keloid and neonatal RAFTs was greater than that observed in normal adult RAFTs. Treated RAFTs contracted less over the 14-day period compared to controls. Contraction and collagen density were greatest in keloid and neonatal RAFTS. Conclusions: A PDT dosimetry range has been established that reduces tissue contraction and collagen synthesis while preserving cell viability.

Assessment of Photosensitizer Dosimetry and Tissue Damage Assay for Photodynamic Therapy in Advanced‐stage Tumors¶

ABSTRACTPhotodynamic therapy (PDT) efficacy is a complex function of tissue sensitivity, photosensitizer (PS) uptake, tissue oxygen concentration, delivered light dose and some other parameters. To better understand the mechanisms and optimization of PDT treatment, we assessed two techniques for quantifying tissue PS concentration and two methods for quantifying pathological tumor damage. The two methods used to determine tissue PS concentration kinetic were in vivo fluorescence probe and ex vivo chemical extraction. Both methods show that the highest tumor to normal tissue PS uptake ratio appears 4 h after PS administration. Two different histopathologic techniques were used to quantify tumor and normal tissue damage. A planimetry assessment of regional tumor necrosis demonstrated a linear relationship with increasing light dose. However, in large murine tumors this finding was complicated by the presence of significant spontaneous necrosis. A second method (densitometry) assessed cell death by nuclear size and density. With some exceptions the densitometry method generally supported the planimetry results. Although the densitometry method is potentially more accurate, it has greater potential subjectivity. Finally, our research suggests that the tools or methods we are studying for quantifying PS levels and tissue damage are necessary for the understanding of PDT effect and therapeutic ratio in experimental in vivo tumor research.

Differences between cytotoxicity in photodynamic therapy using a pulsed laser and a continuous wave laser: study of oxygen consumption and photobleaching

Oxygen consumption at the targeted site has a significant effect on dosimetry in photodynamic therapy (PDT). However, oxygen consumption in PDT using a pulsed laser as a light source has not been clarified. We therefore investigated the dependence of cytotoxicity on the oxygen consumption and the photosensitizer photobleaching of PDT using a pulsed laser by comparing with that using a continuous wave (CW) laser. Mouse renal carcinoma cells (Renca) were incubated with a second-generation photosensitizer, PAD-S31. The cells were then irradiated with either a 670-nm nanosecond pulsed light from the 3rd harmonics of a Nd:YAG laser-pumped optical parametric oscillator with a peak fluence rate of approximately 1 MW/cm(2) at 30 Hz or a 670-nm CW diode laser with a total light dose of 40 J/cm(2). Regardless of laser source, cytotoxic effects exhibited cumulative dose responses to the photosensitizer ranging from 12 to 96 microg/ml. However, cytotoxic effect of PDT using the pulsed light was significantly less than that using the CW light with the photosensitizer concentrations of 24 and 48 microg/ml under identical fluence rates. During PDT, the cells exposed to the pulsed light consumed oxygen more slowly, resulting in a lower amount of oxygen consumption when compared with PDT using CW light. In accordance with oxygen consumption, the pulsed light induced significantly less photobleaching of the photosensitizer than the CW light did. These results indicate that the efficiency of PDT using pulsed light is less when compared with CW light, probably being related to suppressed oxygen consumption during the pulsed light irradiation.

Assessment of photosensitizer dosimetry and tissue damage assay for photodynamic therapy in advanced-stage tumors.

Photodynamic therapy (PDT) efficacy is a complex function of tissue sensitivity, photosensitizer (PS) uptake, tissue oxygen concentration, delivered light dose and some other parameters. To better understand the mechanisms and optimization of PDT treatment, we assessed two techniques for quantifying tissue PS concentration and two methods for quantifying pathological tumor damage. The two methods used to determine tissue PS concentration kinetic were in vivo fluorescence probe and ex vivo chemical extraction. Both methods show that the highest tumor to normal tissue PS uptake ratio appears 4 h after PS administration. Two different histopathologic techniques were used to quantify tumor and normal tissue damage. A planimetry assessment of regional tumor necrosis demonstrated a linear relationship with increasing light dose. However, in large murine tumors this finding was complicated by the presence of significant spontaneous necrosis. A second method (densitometry) assessed cell death by nuclear size and density. With some exceptions the densitometry method generally supported the planimetry results. Although the densitometry method is potentially more accurate, it has greater potential subjectivity. Finally, our research suggests that the tools or methods we are studying for quantifying PS levels and tissue damage are necessary for the understanding of PDT effect and therapeutic ratio in experimental in vivo tumor research.

Photodynamic Therapy

Photodynamic Therapy

Intracellular Photobleaching of 5,10,15,20-Tetrakis(m-hydroxyphenyl) chlorin (Foscan®) Exhibits a Complex Dependence on Oxygen Level and Fluence Rate¶

The understanding of photosensitizer photobleaching is important not only for mechanistic studies, but also for the development of monitoring techniques for clinical dosimetry in photodynamic therapy. In this study, we investigated the intracellular photobleaching of 5,10,15,20-tetrakis(m-hydroxyphenyl)chlorin (mTHPC, Foscan) in the murine macrophage cell line J774A.1, using quantitative fluorescence imaging microscopy, microspectrofluorometry and microspectrophotometry. Using 652 nm laser irradiation, it was found that mTHPC exhibits oxygen- and fluence rate-dependent intracellular photobleaching. The kinetics showed an inverse dose-rate behavior, i.e. a reduction of fluence rate resulted in more photobleaching at comparable fluences. The effect of deoxygenation was found to be more complex, with decreased bleaching at low fluence rates and increased bleaching at higher fluence rates. The intracellular formation of reactive oxygen species was measured using 2',7'-dichlorodihydrofluorescein diacetate. The results are analyzed in terms of competitive Type-I and Type-II mechanisms.

Monitoring photoproduct formation and photobleaching by fluorescence spectroscopy has the potential to improve PDT dosimetry with a verteporfin-like photosensitizer.

In current clinical practice, photodynamic therapy (PDT) is carried out with prescribed drug doses and light doses as well as fixed drug-light intervals and illumination fluence rates. This approach can result in undesirable treatment outcomes of either overtreatment or undertreatment because of biological variations between different lesions and patients. In this study, we explore the possibility of improving PDT dosimetry by monitoring drug photobleaching and photoproduct formation. The study involved 60 mice receiving the same drug dose of a novel verteporfin-like photosensitizer, QLT0074, at 0.3 mg/kg body weight, followed by different light doses of 5, 10, 20, 30, 40 or 50 J/cm2 at 686 nm and a fluence rate of 70 mW/cm2. Photobleaching and photoproduct formation were measured simultaneously, using fluorescence spectroscopy. A ratio technique for data processing was introduced to reliably detect the photoproduct formed by PDT on mouse skin in vivo. The study showed that the QLT0074 photoproduct is stable and can be reliably quantified. Three new parameters, photoproduct score (PPS), photobleaching score (PBS) and percentage photobleaching score (PBS%), were introduced and tested together with the conventional dosimetry parameter, light dose, for performance on predicting PDT-induced outcome, skin necrosis. The statistical analysis of experimental results was performed with an ordinal logistic regression model. We demonstrated that both PPS and PBS improved the prediction of skin necrosis dramatically compared to light dose. PPS was identified as the best single parameter for predicting the PDT outcome.

The effects of oxygenation and photosensitizer substrate binding on the use of fluorescence photobleaching as a dose metric for photodynamic therapy

Photodynamic therapy (PDT) is an evolving clinical treatment that uses light-activated drugs (photosensitizers) for cell or tissue destruction, often mediated through photochemically-generated singlet oxygen ( 1 O 2 ) that is highly reactive. PDT dosimetry is complicated by the multiple factors involved: photosensitizer, light and tissue oxygenation. Photosensitizer photobleaching has been proposed as a simple method of dosimetry that obviates the need to measure these factors directly. This is based on the hypothesis that a greater degree of photobleaching should correspond to a higher effective PDT dose delivered to the tissue. Using meso-tetra hydroxyphenyl chlorin ( m-THPC) in solution as a model photosensitizer, it was found that the photobleaching rate is oxygen dependent and that there is a clear relationship between 1 O 2 concentration and the photobleaching rate. However, this oxygen dependency was markedly altered by the microenvironment, i.e. the solvent and binding to biological substrates. Hence, a measurement of the photobleaching rate alone does not predict for 1 O 2 generation and, hence, cannot be interpreted simply in terms of the PDT dose. Additional information, such as the tissue oxygenation, is required.

Some applications of ultrashort laser pulses in biology andmedicine

Ultrashort laser pulses are finding increasing applications inbiology and medicine. Such pulses can be used directly or after non-linearmodification. Direct utilization includes propagation studies in scatteringmedia with applications in optical mammography, dosimetry for photodynamictherapy and species concentration assessment. Intense continua ofelectromagnetic radiation of very brief duration are formed in the interactionof focused ultrashort and intense laser pulses with matter. Two differentkinds of experiment using such radiation are described, employing visibleradiation and x-rays, respectively. Focusing into water leads to thegeneration of a light continuum through self-phase modulation. The propagationof the light through tissue was studied, addressing questions related tospecific tissue chromophore absorption. When terawatt laser pulses are focusedonto a solid target with high nuclear charge Z, intense x-ray radiation offew ps duration and with energies exceeding hundreds of keV is emitted.Biomedical applications of this radiation are described, includingdifferential absorption and gated-viewing imaging.