Effect of photodynamic therapy with 5-aminolevulinic acid and EDTA-2Na against mixed infection of methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa.

BACKGROUND: The increasing abundance of drug-resistant bacteria is a global threat. Photodynamic therapy is an entirely new, non-invasive method for treating infections caused by antibiotic-resistant strains. We previously described the bactericidal effect of photodynamic therapy on infections caused by a single type of bacterium. We showed that gram-positive and gram-negative bacteria could be killed with 5-aminolevulic acid and 410 nm light, respectively. However, clinically, mixed infections are common and difficult to treat. OBJECTIVE: We investigated the bactericidal effects of photodynamic therapy on mixed infections of methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa. METHODS: We compared bacterial growth with and without photodynamic therapy in vitro. Then, in vivo, we studied mixed infections in a mouse skin ulcer model. We evaluated the rates of ulcer area reduction and transitions to healing in treated and untreated mice. In addition, a comparison was made between PDT and existing topical drugs. RESULTS: We found that photodynamic therapy markedly reduced the growth of both methicillin-resistant Staphylococcus aureus and Pseudomonas aeruginosa, in culture, and it reduced the skin ulcer areas in mice. PDT was also more effective than existing topical medicines. CONCLUSION: This study showed that photodynamic therapy had antibacterial effects against a mixed infection of gram-positive and gram-negative bacteria, and it promoted skin ulcer healing. These results suggested that photodynamic therapy could be effective in both single- and mixed-bacterial infections.

Wavelength-dependent threshold fluences for melanosome disruption to evaluate the treatment of pigmented lesions with 532-, 730-, 755-, 785-, and 1064-nm picosecond lasers.

BACKGROUND AND OBJECTIVES: A threshold fluence for melanosome disruption has the potential to provide a robust numerical indicator for establishing clinical endpoints for pigmented lesion treatment using a picosecond laser. Although the thresholds for a 755-nm picosecond laser were previously reported, the wavelength dependence has not been investigated. In this study, wavelength-dependent threshold fluences for melanosome disruption were determined. Using a mathematical model based on the thresholds, irradiation parameters for 532-, 730-, 755-, 785-, and 1064-nm picosecond laser treatments were evaluated quantitatively. STUDY DESIGN/MATERIALS AND METHODS: A suspension of melanosomes extracted from porcine eyes was irradiated using picosecond lasers with varying fluence. The mean particle size of the irradiated melanosomes was measured by dynamic light scattering, and their disruption was observed by scanning electron microscopy to determine the disruption thresholds. A mathematical model was developed, combined with the threshold obtained and Monte Carlo light transport to calculate irradiation parameters required to disrupt melanosomes within the skin tissue. RESULTS: The threshold fluences were determined to be 0.95, 2.25, 2.75, and 6.50 J/cm² for 532-, 730-, 785-, and 1064-nm picosecond lasers, respectively. The numerical results quantitatively revealed the relationship between irradiation wavelength, incident fluence, and spot size required to disrupt melanosomes distributed at different depths in the skin tissue. The calculated irradiation parameters were consistent with clinical parameters that showed high efficacy with a low incidence of complications. CONCLUSION: The wavelength-dependent thresholds for melanosome disruption were determined. The results of the evaluation of irradiation parameters from the threshold-based analysis provided numerical indicators for setting the clinical endpoints for 532-, 730-, 755-, 785-, and 1064-nm picosecond lasers.

Nonlinear absorption-based analysis of energy deposition in melanosomes for 532-nm short-pulsed laser skin treatment.

BACKGROUND AND OBJECTIVES: The clinical use of 532-nm short-pulsed lasers has provided effective treatment of epidermal pigmented lesions. However, the detection of significant differences in treatment effects between picosecond and nanosecond lasers has still varied among clinical studies. For robust evaluation of the differences based on the treatment mechanism, this study presents a nonlinear absorption-based analysis of energy deposition in melanosomes for 532-nm short-pulsed laser treatment. STUDY DESIGN/MATERIALS AND METHODS: Nonlinear absorption by melanin is modeled based on sequential two-photon absorption. Absorption cross-sections and nonradiative lifetimes of melanin, which are necessary for the nonlinear absorption-based analysis, are determined from transmittance measurement. Using the model and parameters, energy deposition in melanosomes was calculated with varying fluence and pulse width settings, including actual clinical parameters. RESULTS: The energy deposition in melanosomes increased with shorter laser pulses, and subnanosecond laser pulses were found to be most efficient. The comparison of energy deposition calculated using clinical parameters demonstrated the differences in treatment effects between picosecond and nanosecond lasers reported in clinical studies. CONCLUSION: The nonlinear absorption-based analysis provides quantitative evidence for the safety and efficacy evaluation of short-pulsed laser treatments, which may lead to the establishment of numerical indices for determining treatment conditions. Future studies considering the effects of the surrounding tissue on energy deposition in melanosomes will be needed.

Incident Fluence Analysis for 755-nm Picosecond Laser Treatment of Pigmented Skin Lesions Based on Threshold Fluences for Melanosome Disruption.

BACKGROUND AND OBJECTIVES: In this study, the threshold fluences for disrupting the melanosomes for pigmented skin lesion treatment were determined using a 755-nm picosecond laser for clinical use. Based on the melanosome disruption thresholds, incident fluences corresponding to the target lesion depths were evaluated in silico for different laser spot sizes. STUDY DESIGN/MATERIALS AND METHODS: Melanosome samples were isolated from porcine eyes as alternative samples for human cutaneous melanosomes. The isolated melanosomes were exposed to 755-nm picosecond laser pulses to measure the mean particle sizes by dynamic light scattering and confirm their disruption by scanning electron microscopy. The threshold fluences were statistically determined from the relationships between the irradiated fluences and the mean particle sizes. Incident fluences of picosecond laser pulses for the disruption of melanosomes located at different depths in skin tissue were calculated through a light transport simulation using the obtained thresholds. RESULTS: The threshold fluences of 550- and 750-picosecond laser pulses were determined to be 2.19 and 2.49 J/cm2 , respectively. The numerical simulation indicated that appropriate incident fluences of picosecond laser pulses differ depending on the depth distribution of the melanosomes in the skin tissue, and large spot sizes are desirable for disrupting the melanosomes more deeply located within the skin tissue. CONCLUSION: The threshold fluences of picosecond laser pulses for melanosome disruption were determined. The incident fluence analysis based on the thresholds for melanosome disruption provides valuable information for the selection of irradiation endpoints for picosecond laser treatment of pigmented skin lesions. Lasers Surg. Med. © 2021 The Authors. Lasers in Surgery and Medicine published by Wiley Periodicals LLC.

Ultralow radiant exposure of a short-pulsed laser to disrupt melanosomes with localized thermal damage through a turbid medium.

Short-pulsed lasers can treat dermal pigmented lesions through selective photothermolysis. The irradiated light experiences multiple scattering by the skin and is absorbed by abnormal melanosomes as well as by normal blood vessels above the target. Because the fluence is extremely high, the absorbed light can cause thermal damage to the adjacent tissue components, leading to complications. To minimize radiant exposure and reduce the risk of burns, a model of the melanosome-disruption threshold fluence (MDTF) has been developed that accounts for the light-propagation efficiency in the skin. However, the light-propagation efficiency is attenuated because of multiple scattering, which limits the extent to which the radiant exposure required for treatment can be reduced. Here, this study demonstrates the principle of melanosome disruption with localized thermal damage through a turbid medium by ultralow radiant exposure of a short-pulsed laser. The MDTF model was combined with a wavefront-shaping technique to design an irradiation condition that can increase the light-propagation efficiency to the target. Under this irradiation condition, melanosomes were disrupted at a radiant exposure 25 times lower than the minimal value used in conventional laser treatments. Furthermore, almost no thermal damage to the skin was confirmed through a numerical simulation. These experimental and numerical results show the potential for noninvasive melanosome disruption and may lead to the improvement of the safety of short-pulsed laser treatment.

Deep learning based depth map estimation of protoporphyrin IX in turbid media using dual wavelength excitation fluorescence.

This study presents a depth map estimation of fluorescent objects in turbid media, such as biological tissue based on fluorescence observation by two-wavelength excitation and deep learning-based processing. A U-Net-based convolutional neural network is adapted for fluorophore depth maps from the ratiometric information of the two-wavelength excitation fluorescence. The proposed method offers depth map estimation from wide-field fluorescence images with rapid processing. The feasibility of the proposed method was demonstrated experimentally by estimating the depth map of protoporphyrin IX, a recognized cancer biomarker, at different depths within an optical phantom.

Transient simulation of laser ablation based on Monte Carlo light transport with dynamic optical properties model.

Laser ablation is a minimally invasive therapeutic technique to denature tumors through coagulation and/or vaporization. Computational simulations of laser ablation can evaluate treatment outcomes quantitatively and provide numerical indices to determine treatment conditions, thus accelerating the technique's clinical application. These simulations involve calculations of light transport, thermal diffusion, and the extent of thermal damage. The optical properties of tissue, which govern light transport through the tissue, vary during heating, and this affects the treatment outcomes. Nevertheless, the optical properties in conventional simulations of coagulation and vaporization remain constant. Here, we propose a laser ablation simulation based on Monte Carlo light transport with a dynamic optical properties (DOP) model. The proposed simulation is validated by performing optical properties measurements and laser irradiation experiments on porcine liver tissue. The DOP model showed the replicability of the changes in tissue optical properties during heating. Furthermore, the proposed simulation estimated coagulation areas that were comparable to experimental results at low-power irradiation settings and provided more than 2.5 times higher accuracy when calculating coagulation and vaporization areas than simulations using static optical properties at high-power irradiation settings. Our results demonstrate the proposed simulation's applicability to coagulation and vaporization region calculations in tissue for retrospectively evaluating the treatment effects of laser ablation.

Endoscopic image-guided laser treatment system based on fiber bundle laser steering.

A miniaturized endoscopic laser system with laser steering has great potential to expand the application of minimally invasive laser treatment for micro-lesions inside narrow organs. The conventional systems require separate optical paths for endoscopic imaging and laser steering, which limits their application inside narrower organs. Herein, we present a novel endoscopic image-guided laser treatment system with a thin tip that can access inside narrow organs. The system uses a single fiber bundle to simultaneously acquire endoscopic images and modulate the laser-irradiated area. The insertion and operation of the system in a narrow space were demonstrated using an artificial vascular model. Repeated laser steering along set targets demonstrated accurate laser irradiation within a root-mean-square error of 28 [Formula: see text]m, and static repeatability such that the laser irradiation position was controlled within a 12 [Formula: see text]m radius of dispersion about the mean trajectory. Unexpected irradiation on the distal irradiated plane due to fiber bundle crosstalk was reduced by selecting the appropriate laser input diameter. The laser steering trajectory spatially controlled the photothermal effects, vaporization, and coagulation of chicken liver tissue. This novel system achieves minimally invasive endoscopic laser treatment with high lesion-selectivity in narrow organs, such as the peripheral lung and coronary arteries.

Mathematical modelling for antimicrobial photodynamic therapy mediated by 5-aminolaevulinic acid: An in vitro study.

BACKGROUND: Antimicrobial photodynamic therapy (aPDT) using aminolaevulinic acid (ALA) is a promising alternative to antibiotic therapy. ALA administration induces protoporphyrin IX (PpIX) accumulation in bacteria, and light excitation of the accumulated PpIX generates singlet oxygen to bacterial toxicity. Several factors, including drug administration and light irradiation conditions, contribute to the antibiotic effect. Such multiple parameters should be determined moderately for effective aPDT in clinical practice. METHODS: A mathematical model to predict bacterial dynamics in ALA-aPDT following clinical conditions was constructed. Applying a pharmacokineticspharmacodynamics (PK-PD) approach, which is widely used in antimicrobial drug evaluation, viable bacteria count by defining the bactericidal rate as the concentration of singlet oxygen produced when PpIX in bacteria is irradiated by light. RESULTS: The in vitro experimental results of ALA-aPDT for Pseudomonas aeruginosa demonstrated the PK-PD model validity. The killing rate has an upper limit, and the lower power density for a long irradiation time can suppress the viable bacteria number when the light dosages are the same. CONCLUSIONS: This study proposed a model of bacterial viability change in ALA-aPDT based on the PK-PD model and confirmed, by in vitro experiments using PA, that the variation of bacterial viability with light-sensitive substance concentration and light irradiation power densities could be expressed. Further validation of the PK-PD model with other gram negative and gram positive strains will be needed.

Measurement of absorption and reduced scattering coefficients in Asian human epidermis, dermis, and subcutaneous fat tissues in the 400- to 1100-nm wavelength range for optical penetration depth and energy deposition analysis (Errata).

The errata correct errors that appeared in Table 2 of the published article.

Measurement of absorption and reduced scattering coefficients in Asian human epidermis, dermis, and subcutaneous fat tissues in the 400- to 1100-nm wavelength range for optical penetration depth and energy deposition analysis.

SIGNIFICANCE: In laser therapy and diagnosis of skin diseases, the irradiated light distribution, which is determined by the absorption coefficient µa and reduced scattering coefficient µs' of the epidermis, dermis, and subcutaneous fat, affects the treatment outcome and diagnosis accuracy. Although values for µa and µs' have been reported, detailed analysis for Asian skin tissues is still lacking. AIM: We present µa and µs' measurements of Asian skin tissues in the 400- to 1100-nm wavelength range for evaluating optical penetration depth and energy deposition. APPROACH: The measurements with Asian human skin samples are performed employing a double integrating sphere spectrometric system and an inverse Monte Carlo technique. Using the measured parameters, the optical penetration depth and energy deposition are quantitatively analyzed. RESULTS: The µa of the epidermis layer varies among different ethnic groups, while the µa of the other layers and the µs' of all of the layers exhibit almost no differences. The analysis reveals that the optical penetration depth and the energy deposition affect the photodynamic therapy treatment depth and the heat production in skin tissue, respectively. CONCLUSIONS: The experimentally measured values of µa and µs' for Asian skin tissues are presented, and the light behavior in Asian skin tissues is analyzed using a layered tissue model.