Published papers

Background Lymphatic vessels are difficult to identify using existing modalities as because of their small diameter and the transparency of the lymph fluid flowing through them. Methods Here, we introduce photoacoustic lymphangiography (PAL), a new modality widely used for lymphedema treatment, to observe limb lymphatic vessels. The photoacoustic imaging system used in this study can simultaneously visualize lymphatic vessels and veins with a high resolution (0.2 mm) and can also observe their three-dimensional relationship with each other. Results High-resolution images of the lymphatic vessels, detailed structure of the dermal back flow, and the three-dimensional positional relationship between the lymphatic vessels and veins were observed by PAL. Conclusion The clear image provided by PAL could have a major application in pre- and postoperative use during lymphaticovenular anastomosis for lymphedema treatment.

A photoacoustic 3D imaging system for animal experiments was made. This system is special because it has a hemispherical detector array. To test its performance, we used a chart from the field of optics as a sample. We checked the whole imaging range using the ISO 12233 chart, which is used to test digital camera images. We found that there was no distortion in the xy-plane and the system had high resolution. We also tested it using a high image quality mode with a different scanning sequence. In this study, live albino mice with white hairs were anesthetized and photographed. Using hair removal cream, we were able to visualize the vascular network throughout their bodies, including blood vessels in organs such as the liver and kidneys. The smallest vessels we were able to visualize were less than 0.1 mm in diameter. We used photoacoustic (PA) images to relatively estimate the oxygen saturation of the mice's blood at two different wavelengths, which we refer to as the S-factor. By analyzing the PA images, we were able to estimate the arterial and venous systems of the whole body, as well as the difference in S-factor between the two systems within the liver. When the mice were euthanized and examined post-mortem, we observed that the S-factor of the whole body decreased and the difference in S-factor between the two systems within the liver was lost.

Photoacoustic (PA) technology can be used for non-invasive imaging of blood vessels. In this paper, we report on our prototype PA imaging system with a newly designed ultrasound sensor and its visualization performance of microvascular in animal. We fabricated an experimental system for animals using a high-frequency sensor. The system has two modes: still image mode by wide scanning and moving image mode by small rotation of sensor array. Optical test target, euthanized mice and rats, and live mice were used as objects. The results of optical test target showed that the spatial resolution was about two times higher than that of our conventional prototype. The image performance in vivo was evaluated in euthanized healthy mice and rats, allowing visualization of detailed blood vessels in the liver and kidneys. In tumor-bearing mice, different results of vascular induction were shown depending on the type of tumor and the method of transplantation. By utilizing the video imaging function, we were able to observe the movement of blood vessels around the tumor. We have demonstrated the feasibility of the system as a less invasive animal experimental device, as it can acquire vascular images in animals in a non-contrast and non-invasive manner.

Background Lymphaticovenular anastomosis (LVA) is a minimally invasive surgical procedure used to treat lymphedema. Volumetric measurements and quality-of-life assessments are of-ten performed to assess the effectiveness of LVA, but there is no method that provides information regarding postoperative morphological changes in lymphatic vessels and veins after LVA. Photoacoustic lymphangiography (PAL) is an optical imaging technique that visualizes the distribution of light-absorbing molecules, such as hemoglobin or indocyanine green (ICG), and provides three-dimensional images of superficial lymphatic vessels and the venous system simultaneously. In this study, we performed PAL in lymphedema patients before and after LVA and compared the images to evaluate the effect of LVA. Methods PAL was performed using the PAI-05 system in three patients (one man, two women) with lymphedema, including one primary case and two secondary cases, before LVA. ICG fluorescence lymphography was performed in all cases before PAL. Follow-up PAL was performed between 5 days and 5 months after LVA. Results PAL enabled the simultaneous visualization of clear lymphatic vessels that could not be accurately seen with ICG fluorescence lymphography and veins. We were also able to observe and analyze morphological changes such as the width and the number of lymphatic vessels and veins during the follow-up PAL after LVA. Conclusions By comparing preoperative and postoperative PAL images, it was possible to analyze the morphological changes in lymphatic vessels and veins that occurred after LVA. Our study suggests that PAL would be useful when assessing the effect of LVA surgery.

Purpose: Varicose veins in the lower extremities are dilated subcutaneous varicose veins with a diameter of ≥ 3 mm, caused by increased venous pressure resulting from backflow of blood due to venous valve insufficiency (Gloviczki in Handbook of venous disorders: guidelines of the American venous forum, Hodder Arnold, London, 2009). When diagnosing varicose veins, the shape and thickness of the blood vessels should be accurately visualized in three dimensions. In this study, we investigated a new method for numerical evaluation of vascular morphology related to varicose veins in the lower extremities, using a photoacoustic imaging (PAI) system, which can acquire high-resolution and three-dimensional images noninvasively. Methods: Nine patients with varicose veins participated in the study, and their images were captured using an optical camera and PAI system. We visualized the vascular structure, created a blood presence density (BPD) heat map, and examined the correlation between BPD and location of varicose veins. Results: The obtained photoacoustic (PA) images demonstrated the ability of this method to visualize vessels ranging from as small as 0.2 mm in diameter to large, dilated vessels in three dimensions. Furthermore, the study revealed a correlation between the high-density part of the BPD heat map generated from the PAI images and the presence of varicose veins. Conclusion: PAI is a promising technique for noninvasive and accurate diagnosis of varicose veins in the lower extremities. By providing valuable information on the morphology and hemodynamics of the varicose veins, PAI may facilitate their early detection and treatment.

Spectral photoacoustic imaging (PAI) is a new technology that is able to provide 3D geometric structure associated with 1D wavelength-dependent absorption information of the interior of a target in a non-invasive manner. It has potentially broad applications in clinical and medical diagnosis. Unfortunately, the usability of spectral PAI is severely affected by a time-consuming data scanning process and complex noise. Therefore in this study, we propose a reliability-aware restoration framework to recover clean 4D data from incomplete and noisy observations. To the best of our knowledge, this is the first attempt for the 4D spectral PA data restoration problem that solves data completion and denoising simultaneously. We first present a sequence of analyses, including modeling of data reliability in the depth and spectral domains, developing an adaptive correlation graph, and analyzing local patch orientation. On the basis of these analyses, we explore global sparsity and local self-similarity for restoration. We demonstrated the effectiveness of our proposed approach through experiments on real data captured from patients, where our approach outperformed the state-of-the-art methods in both objective evaluation and subjective assessment.

Lymphaticovenular anastomosis (LVA) is a widely performed surgical procedure for the treatment of lymphedema. For good LVA outcomes, identifying lymphatic vessels and venules is crucial. Photoacoustic lymphangiography (PAL) is a new technology for visualizing lymphatic vessels. It can depict lymphatic vessels at high resolution; therefore, this study focused on how to apply PAL for lymphatic surgery. To visualize lymphatic vessels, indocyanine green was injected as a color agent. PAI-05 was used as the photoacoustic imaging device. Lymphatic vessels and veins were visualized at 797-and 835-nm wavelengths. First, it was confirmed whether the branching of the vasculature as depicted by the PAL was consistent with the actual branching of the vasculature as confirmed intraoperatively. Second, to use PAL images for surgical planning, preoperative photoacoustic images were superimposed onto the patient limb through augmented reality (AR) glasses (MOVERIO Smart Glass BT-30E). Lymphatics and venule markings drawn using AR glasses were consistent with the actual intraoperative images obtained during LVA. To anastomose multiple lymphatic vessels, a site with abundant venous branching was selected as the incision site; and selecting the incision site became easier. The anatomical morphology obtained by PAL matched the surgical field. AR-based marking could be very useful in future LVA.

Summary: It is difficult to evaluate the postoperative patency of lymphaticovenular anastomosis, but this evaluation is essential for determining surgical results. When using the current standard modality, near-infrared fluorescent lymphography, it is difficult to observe patency if the anastomotic point is veiled by dermal backflow. In this study, we used a new photoacoustic imaging device, PAI-05, to check the patency of anastomosis. We performed photoacoustic lymphangiography after lymphaticovenular anastomosis surgery. By digitally subtracting the superficial area, we can examine an area deeper than the dermal backflow, which is not visible by near-infrared fluorescent lymphography. The connection between the lymphatic vessel and the venule observed in the image is an indication of the patency of anastomosis. However, in a non-patent anastomosed site, the lymphatic vessel has a gap that separates it from the venule at the anastomosed site. Although photoacoustic lymphangiography cannot be used to visualize the lymphatic vessels that are not contrasted by indocyanine green, the resulting high-resolution images and clear anastomosis evaluation afforded by it will contribute to the development of future lymphedema treatments.

Background: Dermal backflow (DBF), which refers to lymphatic reflux due to lymphatic valve insufficiency, is a diagnostic finding in lymphedema. However, the three-dimensional structure of DBF remains unknown. Photoacoustic lymphangiography (PAL) is a new technique that enables the visualization of the distribution of light-absorbing molecules, such as hemoglobin or indocyanine green (ICG), and can provide three-dimensional images of superficial lymphatic vessels and the venous system. This study reports the use of PAL to visualize DBF structures in the extremities of patients with lymphedema after cancer surgery. Methods: Patients with a clinical or lymphographic diagnosis of lymphedema who previously underwent surgery for cancer at one of two participating hospitals were included in this study. PAL was performed using the PAI-05 system. ICG was administered subcutaneously in the affected hand or foot, and ICG fluorescence lymphography was performed using a nearnfrared camera system prior to PAL. Results: Between April 2018 and January 2019, 21 patients were enrolled and examined usng PAL. The DBF was composed of dense, interconnecting, three-dimensional lymphatic vessels. It was classified into three patterns according to the composition of the lymphatic vessels: A linear structure of lymphatic collectors (pattern 1), a network of lymphatic capillaries and lymphatic collectors in an underlying layer (pattern 2), and lymphatic capillaries and precollectors with no lymphatic collectors (pattern 3). Conclusions: PAL showed the structure of DBF more precisely than ICG fluorescence lymphography. The use of PAL to visualize DBF assists in understanding the pathophysiology and assessing the severity of cancer-related lymphedema.

Photoacoustic (PA) imaging is a new imaging technology that can non-invasively visualize blood vessels and body hair in 3D. It is useful in cosmetic surgery for detecting body hair and computing metrics such as the number and thicknesses of hairs. Previous supervised body hair detection methods often do not work if the imaging conditions change from training data. We propose an unsupervised hair detection method. Hair samples were automatically extracted from unlabeled samples using prior knowledge about spatial structure. If hair (positive) samples and unlabeled samples are obtained, Positive Unlabeled (PU) learning becomes possible. PU methods can learn a binary classifier from positive samples and unlabeled samples. The advantage of the proposed method is that it can estimate an appropriate decision boundary in accordance with the distribution of the test data. Experimental results using real PA data demonstrate that the proposed approach effectively detects body hairs.

Objective: Aging is one of the causes of primary lymphedema. However, the effects of aging on the lymphatic system are still not completely understood. We investigated the effects of aging on the lymphatic vessels in the lower extremities of healthy volunteers using photoacoustic imaging. Methods: Healthy volunteers who underwent photoacoustic lymphangiography between March 2018 and January 2019 were enrolled. To visualize lymphatics, indocyanine green (ICG, 5.0 mg/mL) was injected subcutaneously into the first and fourth web spaces of the foot and under the lateral malleolus. Subsequently, near-infrared fluorescence lymphography was performed to confirm good ICG flow, and photoacoustic lymphangiography was performed on the medial side of the lower leg. Ti sapphire laser irradiation at 797 and 835 nm, the optimal wavelengths for visualizing ICG and blood, was applied. The number of lymphatic vessels shown at areas 10 cm (L10) and 20 cm (L20) cranially from the internal malleolus was counted. Results: Nineteen healthy volunteers (4 males and 15 females) were enrolled in the study. Their mean age was 42.9 ± 12.8 years. One volunteer was bilaterally imaged; 15 left lower limbs and 5 right lower limbs were imaged. The number of lymphatic vessels visualized increased with age. There were strong positive correlations between age and L10 (R = 0.729, P <.001) and between age and L20 (R = 0.570, P =.009). Conclusions: Photoacoustic imaging indicates that the number of lymphatic vessels increases with age. Lymphatic stasis resulted in visualization of not only normal drainage pathways but also nonfunctional lymphatic pathways.

Background: Photoacoustic microscopy is expected to have clinical applications as a noninvasive and three-dimensional (3D) method of observing intradermal structures. Objective: Investigate the applicability of a photoacoustic microscope equipped with two types of pulsed lasers that can simultaneously recognize hemoglobin and melanin. Methods: 16 skin lesions including erythema, pigmented lesions, vitiligo and purpura, were analyzed to visualize 3D structure of melanin granule distribution and dermal blood vessels. 13 cases of livedo racemosa in cutaneous polyarteritis nodosa (cPN) were further analyzed to visualize the 3D structure of dermal blood vessels in detail. Vascular structure was also analyzed in the biopsy specimens obtained from tender indurated erythema of cPN by CD34 immunostaining. Results: Hemoglobin-recognition signal clearly visualized the 3D structure of dermal blood vessels and melanin-recognition signal was consistently reduced in vitiligo. In livedo racemosa, the hemoglobin-recognition signal revealed a relatively thick and large reticular structure in the deeper layers that became denser and finer toward the upper layers. The numerical analysis revealed that the number of dermal blood vessels was 1.29-fold higher (p<0.05) in the deeper region of the lesion than that of normal skin. The CD34 immunohistochemical analysis in tender indurated erythema revealed an increased number of dermal vessels compared with normal skin in 88.9% (8/9) of the cases, suggesting that vascular network remodeling had occurred in cPN. Conclusion: The photoacoustic system has an advantage in noninvasively detecting dermal blood vessel structures that are difficult to recognize by two-dimensional histopathology specimen examination and is worth evaluating in various skin diseases.

Lymphedema occurs when interstitial fluid and fibroadipose tissues accumulate abnormally because of decreased drainage of lymphatic fluid as a result of injury, infection, or congenital abnormalities of the lymphatic system drainage pathway. An accurate anatomical map of the lymphatic vasculature is needed not only for understanding the pathophysiology of lymphedema but also for surgical planning. However, because of their limited spatial resolution, no imaging modalities are currently able to non-invasively provide a clear visualization of the lymphatic vessels. Photoacoustic imaging is an emerging medical imaging technique that provides unique scalability of optical resolution and acoustic depth of penetration. Moreover, light-absorbing biomolecules, including oxy-and deoxyhemoglobin, lipids, water, and melanin, can be imaged. Using exogenous contrast agents that are taken up by lymphatic vessels, e.g., indocyanine green, photoacoustic lymphangiography, which has a higher spatial resolution than previous imaging modalities, is possible. Using a new prototype of a photoacoustic imaging system with a wide field of view developed by a Japanese research group, high-resolution three-dimensional structural information of the vasculatures was successfully obtained over a large area in both healthy and lymphedematous extremities. Anatomical information on the lymphatic vessels and adjacent veins provided by photoacoustic lymphangiography is helpful for the management of lymphedema. In par-ticular, such knowledge will facilitate the planning of microsurgical lymphaticovenular anastomoses to bypass the excess fluid component by joining with the circulatory system peripherally. Although challenges remain to establish its implementation in clinical practice, photoacoustic lymphangiography may contribute to improved treatments for lymphedema patients in the near future.