Lymphedema, either primary or secondary, is a chronic and debilitating disorder affecting millions of persons around the world. Although compression therapy is the first choice in the treatment of lymphedema, lymphatic surgery is indicated when conservative therapy fails. While dynamic contrast magnetic resonance (MR) lymphangiography readily visualizes the central lymphatic anatomy [1
], reliable and non-invasive visualization of peripheral lymphatic vessels remains challenging.
Identification of lymphatic vessels by means of indocyanine green (ICG) lymphography in patients with severe lymphedema is limited due to the overlying dermal backflow. Moreover, deep and/or small vessels cannot be detected with the current technology. Detection of lymphatic vessels in obese patients with lymphedema is even more troublesome. Although ultra-high frequency ultrasound can detect small vessels, due to the inability of coaptating ICG contrast, finding lymphatics in cases of severe lymphedema can be demanding. In addition, differentiating veins from lymphatic vessels can be difficult and depends on the operator’s experience. Such hurdles motivate the search for novel diagnostic approaches to patients with severe lymphedema particularly in view of subsequent surgical treatment.
In optoacoustic tomography a laser-induced localized thermoelastic expansion generates acoustic waves from which a 3D image is reconstructed. This emerging technology has been applied in preclinical models [2
] and in humans, mainly in the field of oncology to detect tumors and metastases [3
], but also in muscular and inflammatory diseases [6
]. Blood and lymph vessels can be visualized and traced without ionizing radiation thanks to the unique light absorption spectral profile by both endogenous tissue chromophores and exogenous contrast agents including indocyanine green. Kajita et al. [8
] were the first to describe lymphatic vessels up to a diameter of 0.2 mm using a 3D photoacoustic visualization system (Luxonus Inc., Kawasaki, Kanagawa, Japan). However, the need to immerse the body part in water and the size of the device’s configuration with dedicated bed [9
], hamper widespread application of a promising technology.
In contrast, Multispectral Optoacoustic Tomography (MSOT) in handheld mode operating at video rates can be performed at the bedside without water immersion of the limb. In addition, the handheld MSOT device in the present work can be used to apply light at several wavelengths to optimally unmix tissue and injected chromophores, while other devices are restricted to two wavelengths [2
]. While MSOT with a handheld scanner was found suitable for clinical imaging of major blood vessels and microvasculature in healthy persons [10
] and in patients with vascular malformations [11
], its use to visualize lymphatic vessels in patients with lymphedema has never been described.
We conducted a pilot study in 11 lymphedema patients in order to test whether lymphatic vessels, particularly in areas of dermal backflow, could be identified and traced by multispectral optoacoustic tomography.
Following MSOT examination, no side effects, particularly skin damage, were observed in any of the patients.
In eight out of 11 patients, both lymphatic and blood vessels could be visualized with MSOT in handheld mode, while in three patients only blood vessels were identified (Table 1
In eight patients, lymphatic vessels were clearly differentiated from veins during the MSOT examination by applying a slight pressure onto the skin (Figure 2
, Video S2
). Lymphatic contractility was observed in two patients (Video S3
Furthermore, lymphatic vessels could be identified in regions with linear pattern as well as in regions without visible fluorescence (Figure 3
In patients with primary (Figure 4
) and secondary lower limb lymphedema, lymphatic vessels were identified in areas exhibiting a diffuse ICG pattern.
Likewise, in the upper limb, lymphatic vessels and veins were identified by MSOT in areas of pronounced dermal backflow (Figure 5
In patient 1, scheduled for microsurgical LVA treatment, a custom-made 3D pointer aided in marking the incision site (Figure 6
Indeed, vessel position based on MSOT images corresponded well with intra-operative findings (Figure 7
). Consequently, an end-to-end lympho-venous anastomosis between a 0.30 mm lymphatic vessel and a 0.35 mm vein was performed. Patency of the lympho-venous anastomosis was confirmed intra-operatively by ICG lymphography (Video S4
). Lymphatic structure was confirmed by postoperative histopathologic inspection.
In this pilot study we investigated the performance of a mobile handheld MSOT device for imaging lymphatic vessels in patients with lymphedema in view of possible microsurgical treatment.
Lymphatic vessels and their characteristics could be visualized in eight out of 11 patients. In one patient, hair at the scanned region interfered with the analysis of the images. This has been reported previously [9
] and is not surprising since MSOT has also been applied to hair follicle imaging [15
]. We believe therefore that this case represents a purely technical issue caused by absorption of light by the melanin present in the hair. Since shaving did not eliminate the interference, the application of depilatory creams to completely remove the hair shaft, is advisable in future studies. In the two other patients where only blood vessels were detected, structural issues or inexperience with the novel technique may have played a role.
There were three main findings in our pilot study.
The target lymphatic vessels were visible with high contrast against the background and could be clearly differentiated from blood vessels due to spectral absorption differences between hemoglobin and indocyanine green. As previously described [16
], this differentiation can be confirmed by gently pressing the 3D probe against the skin: the image of only the vein, not the lymphatic, will disappear, to reappear when pressure is lifted. Although we could also evaluate lymphatic patency by ultra-high frequency ultrasound [16
], this technique is limited to a depth of 1 cm, which precludes application in more proximal segments of an affected limb and in advanced stages of lymphedema.
Individual lymphatic vessels were visualized even in areas of dermal backflow, where near-infrared (NIR) technology falls short. While ICG lymphography can clearly visualize patent lymphatic vessels in early-stage lymphedema (linear patterns), individual lymphatics are not identifiable in more advanced-stage lymphedema (diffuse pattern). Likewise, deep lymphatics (up to ~2 cm below the skin) and microvasculature could be identified by MSOT, whereas maximal depth with NIR technology is 1.5 cm. These features, together with the ability of identifying blood vessels at the same time, are particularly important when planning microsurgery, since lympho-venous anastomoses in areas of dermal backflow have been associated with a more favorable outcome [17
MSOT is a valuable tool when evaluating feasibility of LVA surgery. Clear visualization of lymphatic vessels and adjacent veins, inspection of their features together with correct estimation of their proximity will help the surgeon select the ideal site for skin incision. In fact, using the custom-made 3D pointer, a LVA was successfully performed between a MSOT-identified lymphatic vessel and adjacent vein.
Despite the remarkable innovations in lymphatic imaging over the last years, none of the modalities has been able to combine the visualization of peripheral lymphatic vessels and (adjacent) blood vessels in view of required microsurgical treatment including lympho-venous anastomosis. Optoacoustic technology offers several advantages over other imaging techniques used in lymphedema patients. Optoacoustic devices do not require ionizing radiation (contrary to lymphoscintigraphy) or surgical dissection of lymphatic vessels (contrary to transpedal lymphangiography), provide spatial information (contrary to lymphoscintigraphy and ultrasound) and can detect deep vessels even with dermal backflow present (contrary to ICG lymphography). Furthermore, the duration of the MSOT examination is acceptable since the visualization of lymphatics and veins can be performed at the same time which is not possible by any other imaging method. Although lymphatic mapping by ICG lymphography may be faster, adjacent veins cannot be visualized. Lymphoscintigraphy takes up to 3 h and does not provide information on blood vessels either.
Our pilot study describes the first results of lymphatic vessel visualization by handheld 3D multispectral optoacoustic tomography. While MSOT has been applied in cancer, vascular and inflammatory imaging [4
], its use in other fields is still emerging [19
]. In addition, MSOT has been shown to provide investigator-independent, consistent and reproducible functional soft tissue characterization [20
]. Kajita et al. [8
] have previously described the optoacoustic visualization of lymphatic vessels. Their 3D system does not provide real-time images and is limited to the use of two laser wavelengths to differentiate between lymphatic vessels containing indocyanine green from blood vessels containing hemoglobin. In contrast, our images are acquired at seven different wavelengths, which increases their differentiation power.
Furthermore, the portable MSOT device with a handheld probe used in our study is suitable for bedside measurements. Besides mobility, MSOT offers real-time visualization of identified vessels, without the need for data postprocessing. Examination with the MSOT serves as an immediate pre-surgical work-up, and the custom-made pointer will help indicate precisely the site for incision. Future expansion of the wavelength spectrum beyond 1000 nm might result in the detection of other biological tissue properties (e.g., fat in lipedema patients). In addition, extending the acquisition field of view beyond the current 2 cm depth will result in visualization of deeper structures. Finally, as it is highly likely that optoacoustic technology will become more popular in all fields of medicine, increasing the operator’s experience will add to more refined data interpretation.
Although only 11 patients were included in this study, both primary and secondary lymphedema patients were represented, as well as varying degrees of clinical severity. Furthermore, our sample included both men and women, and both upper and lower limb edema cases. Even though our MSOT images were confirmed intra-operatively in one of our patients, our encouraging results should be replicated in larger cohorts.