KEYWORDS: Picosecond phenomena, Luminescence, Photodynamic therapy, Tissues, Optical properties, In vivo imaging, Animal model studies, Tumor growth modeling, Monte Carlo methods, Absorption
Employment of chlorin-based photosensitizers featuring two peaks in absorption spectrum as drugs for photodynamic therapy allows for dual-wavelength fluorescence imaging providing estimation of photosensitizer localization depth.
We present the results of combined fluorescence and optoacoustic monitoring of tumor treatment using novel photoactivatable multi-inhibitor liposomes with BPD and Irinotecan providing a synergetic effect of PDT and chemotherapeutic impact.
We report on the comparative analysis of the effect of photodynamic therapy performed with red or/and blue light on a tumor model in animals. Results of optical monitoring are in agreement with visual observations and histology studies.
Cerebrovascular imaging of rodents is one of the trending applications of optoacoustics aimed at studying brain activity and pathology. Imaging of deep brain structures is often hindered by sub-optimal arrangement of the light delivery and acoustic detection systems. In our work we revisit the physics behind opto-acoustic signal generation and perform theoretical evaluation of optimal laser wavelengths with an aim of enabling whole brain cerebrovascular optoacoustic angiography in small rodents. A comprehensive model based on diffusion approximation was developed to simulate optoacoustic signal generation using optical and acoustic parameters closely mimicking a typical murine brain and surrounding tissues. The model revealed three characteristic wavelength ranges in the visible and near-infrared spectra optimally suited for imaging cerebral vasculature of different size and depth. The theoretical conclusions are confirmed by numerical simulations while in vivo imaging experiments further validated the capacity for accurately resolving brain vasculature at depths ranging between 0.7 and 7 mm.
A novel approach to monitor photosensitizer accumulation and photobleaching in the course of photodynamic therapy (PDT) with the use of nanoconstructs based on the simultaneous fluorescence (FL) and optoacoustic (OA) imaging is implemented. A liposome nanoconstructs employed in this studies contain benzoporphyrin derivatives (BPD) which serve as a photosensitizer and secondly, as a fluorophore, and the fluorescent IRDye800 dye acting as an additional contrasting agent due to its high quantum yield. FL provides visualization of BPD and IRDye800 distribution, while OA principle allows for BPD-absorption based imaging of tumor and its vascular environment. We demonstrate the results of a preliminary in vivo study with combined FL and OA custom-made setups on a NUDE mouse with human glioblastoma U- 87. The results of this studies show a hemorrhage in the tumor area on the OA images obtained @532 nm after PDT that is not visually detected, but confirmed with the following histological verification. Fast nanoconstructs accumulation (< 10 min) was observed using FL imaging with the concentration in tumor only 10% higher than in surrounding tissues. We believe that the ratio of nanoconstructs accumulation in tumor can be significantly increased using target approach.
Two pronounced absorption peaks in blue and red ranges of the chlorin-based photosensitizer (PS) absorption spectrum provide additional benefits in photodynamic therapy (PDT) performance. Differing optical properties of biological tissues in these ranges allow for both dual-wavelength diagnostics and PDT performance. We provide a comparative analysis of different PDT regimes performed with blue and red lights and their combination, with doses varying from 50 to 150 J / cm2. The study was performed on the intact skin of a rabbit ear inner surface, with the use of chlorin e6 as a PS. PDT procedure protocol included monitoring of the treated site with fluorescence imaging technique to evaluate PS accumulation and photobleaching, as well as with optical coherence tomography (OCT) to register morphological and functional responses of the tissue. Optical diagnostic observations were compared with the results of histopathology examination. We demonstrated that PDT procedures with the considered regimes induce weaker organism reaction manifested by edema in normal tissue as compared to irradiation-only exposures with the same light doses. The light doses delivered with red light induce weaker tissue reaction as compared to the same doses delivered with blue light only or with a combination of red and blue lights in equal parts. Results of in-vivo OCT monitoring of tissue reaction are in agreement with the results of histopathology study.
Dual-wavelength photodynamic therapy is a photodynamic therapy (PDT) modality combining therapeutic effects of irradiation at two wavelengths of different region of visible range. In this paper we report on comparative analysis of single- and dual-wavelength PDT regimes based on multimodal optical monitoring of tissue response with histologic verification. Morphological and functional tissue responses to PDT procedure with chlorin-based photosensitizer (PS) were studied at the normal skin of a rabbit ear inner surface. Multimodal optical monitoring was performed by OCT and dual-wavelength fluorescence technique. The studied doses vary from 50 to 150 J/cm2 delivered separately at 405 or 660 nm, or at both wavelengths together in the equal dose.
In this study we present a complex approach to photodynamic therapy (PDT) with chlorin based photosensitizers including Monte Carlo based planning and prediction of optical diagnostics results, intra-procedure dual-wavelength fluorescence monitoring allowing to evaluate PS accumulation and photobleaching, and monitoring of tissue response with optical coherence tomography (OCT). The approach was employed to compare the effects of different PDT regimens in normal and tumor tissues and the results of non-invasive optical diagnostics were matched with results of histologic examination, including hematoxylin-eosin and Mallory staining. The considered doses are in the range 50-150 J/cm2 for normal tissues and 150-275 J/cm2 for tumor tissues, single wavelength (405 and 660 nm) and dual-wavelength regimes are studied.
Photodynamic therapy (PDT) proved itself as a powerful tumor and non-tumor pathologies treatment tool. Photosensitizers (PS), which are employed as therapeutic agents for PDT, feature fluorescent properties. Thus, PDT provides the principles of theranostics when diagnostics is performed during treatment. The estimation of PS localization within the tissue is of critical importance for PDT planning. Fluorescence imaging is commonly applied for the monitoring of PS accumulation within the tissue, however, it does not provide with the in-depth PS distribution. Chlorinbased PS feature two pronounced peaks in their excitation spectra corresponding to 402 nm and 662 nm, which provides additional diagnostic possibilities. The ratio of fluorescence signals corresponding to different excitation wavelengths is shown to be a criterion for the evaluation of PS penetration depth after topical application and PS localization within the tissue after intravenous injection. The study is based on numerical simulation applying Monte Carlo technique. The results of numerical simulations are verified with phantom experiments results.
We propose a new approach to monitoring of photodynamic therapy (PDT) of glioblastoma with the use of targeted nanoconstructs containing a photosensitizer (PS) benzoporphyrin derivative (BPD) and IRDye800 dye, antibodies for efficient accumulation of the drug in a tumor, and a chemotherapeutic agent for combined effect on tumor cells. Monitoring of PDT is based on the simultaneous fluorescent and optoacoustic (OA) imaging. Fluorescent imaging provides visualization of fluorescence agents with high molecular sensitivity, and monitoring of the effectiveness of PDT by PS photobleaching. OA allows to examine the vascular pattern of the tumor environment, as well as assess the tumor depth. IRDye800 is a better contrast agent in comparison to BPD due to red shifted spectral characteristics and higher fluorescence quantum yield. The results of numerical simulations have been verified in phantom studies using fluorescence and optoacoustic experimental setups and an agar phantom with optical characteristics similar to those of murine brain.
KEYWORDS: In vivo imaging, In vitro testing, Blood, Absorption, Blood oxygen saturation, Signal attenuation, Optoacoustics, Blood vessels, Calibration, Veins
We present the comparison of two approaches of blood oxygen saturation determination from multispectral optoacoustic measurements: a calibration-free approach based on evaluation of the effective optical attenuation coefficient derived from in-depth OA signal decay, and an approach based on determination of optical absorption coefficient from OA signal amplitudes. Both approaches were tested in in vitro and in vivo experiments. The results of in vitro and in vivo experiments demonstrated the large difference between experimentally obtained μeff spectra and the literature data, that indicates much lower potential of the OA signal decay approach as compared to OA amplitudes approach. In vivo measurements of the μa spectrum experimentally obtained from OA signal amplitudes give the saturation values of 0.57±0.08 and 0.50±0.07 for two veins of the thoracic spine that agree well with physiological values for venous blood oxygenation in rat. Instead of multiple wavelengths measurements, a pair of wavelengths can be employed for OA measurements. In this case, the saturation maps were obtained at all wavelength pairs from the 658‒1069 nm range. The results demonstrated that the most accurate oxygenation values can be achieved at wavelength pairs of 700 nm and a wavelength from the range 850-1069 nm.
We introduce a portable hand-held hyperspectral imaging system for the functional diagnostics of skin and vascular system. Hyperspectral image analysis aided by artificial neural networks (ANN) allows to reconstruct major physiological parameters of human skin nearly in real-time. The developed device provides spatial distribution of blood volume fraction, oxygenation and melanin content within skin. Special attention has been paid on the system validation and calibration using specially developed skin mimicking phantoms with confirmed optical properties.
The device was built on the basis of unique hyperspectral snapshot camera utilizing a micro Fabry-Perot filter providing real spectral response in each pixel (no interpolation is used in image formation). A broadband illumination unit combined with the camera is based on the fiber-optic illuminator providing uniform distribution of light intensity and utilizes halogen lamp.
The specially developed ANN algorithm was used to perform the inverse problem solution for quantitative assessment of major parameters of skin based on the measured hyperspectral images. A set of diffuse reflectance spectra of human skin imitated by the Monte Carlo method developed in-house has been used extensively for the training of ANN. The volume fraction of blood, oxygen saturation, melanin content and thickness of the epidermal layer were used variable parameters in the utilized seven-layer Monte Carlo-based skin model. The total training set contained 45,198 spectra in the range of 505–800 nm simulated with a step of 5 nm. The developed imaging system has been successfully used to perform the occlusion test measurements with healthy volunteers.
Employment of chlorin-based photosensitizers (PSs) provides additional advantages to photodynamic therapy (PDT) due to absorption peak around 405 nm allowing for superficial impact and efficient antimicrobial therapy. We report on the morphological and clinical study of the efficiency of PDT at 405 nm employing chlorin-based PS. Numerical studies demonstrated difference in the distribution of absorbed dose at 405 nm in comparison with traditionally employed wavelength of 660 nm and difference in the in-depth absorbed dose distribution for skin and mucous tissues. Morphological study was performed at the inner surface of rabbit ear with histological examinations at different periods after PDT procedure. Animal study revealed tissue reaction to PDT consisting in edema manifested most in 3 days after the procedure and neoangiogenesis. OCT diagnostics was confirmed by histological examination. Clinical study included antimicrobial PDT of pharynx chronic inflammatory diseases. It revealed no side effects or complications of the PDT procedure. Pharyngoscopy indicated reduction of inflammatory manifestations, and, in particular cases, hypervascularization was observed. Morphological changes were also detected in the course of monitoring, which are in agreement with pharyngoscopy results. Microbiologic study after PDT revealed no pathogenic bacteria; however, in particular cases, saprophytic flora was detected.
In this work we studied the efficiency of spatially modulated illumination in optical diffuse reflectometry (ODR) and analyzed various probing patterns. The optimal illumination pattern was determined from the series of Monte Carlo numerical experiments on structured illumination and comparison of the parameters of fluence distribution within tissue. We considered the following illumination profiles: sinusoidal patterns with different spatial frequencies k (1 - 2.5 mm-1); piecewise constant patterns with the fixed duty cycle w = 2 and various strip width (0.2 – 1 mm); piecewise constant patterns with the fixed strip width (0.1 mm) and various duty cycle (3 – 11). Assuming the same total incident intensity for all patterns, we observed the growth in intensity at depth with decreasing value of spatial frequency for sinusoidal pattern, the similar tendency of intensity distribution was for piecewise constant patterns with the fixed strip width (or duty cycle) and duty cycle increase (or increase of a strip width, respectively). The intensity distributions within the sample are almost identical for sinusoidal and piecewise constant patterns with matching peaks and lows. However, probing by piecewise constant profile of illumination provide more local information about sample’s properties. A prototype of an ODR system for probing with the structured illumination was designed. The system consists of PC-controlled optical projection system, polarization filters and a CCD camera. The approbation of the system was carried out on the optical phantoms with optical properties close to those of biological tissues and on skin of human volunteers.
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