Current status, pitfalls and future directions in the diagnosis and therapy of lymphatic malformation

Abstract

Lymphatic malformations are complex congenital vascular lesions composed of dilated, abnormal lymphatic channels of varying size that can result in significant esthetic and physical impairment due to relentless growth. Lymphatic malformations comprised of micro-lymphatic channels (microcystic) integrate and infiltrate normal soft tissue, leading to a locally invasive mass. Ultrasonography and magnetic resonance imaging assist in the diagnosis but are unable to detect microvasculature present in microcystic lymphatic malformations. In this review, we examine existing tools and elaborate on alternative diagnostic methods in assessing lymphatic malformations. In particular, photoacoustics, low-toxicity nanoparticles and optical clearing can overcome existing challenges in the examination of lymphatic channels in vivo. In combination with photothermal scanning and flow cytometry, Photoacoustic techniques may provide a versatile tool for lymphatic-related clinical applications, potentially leading to a single diagnostic and therapeutic platform to overcome limitations in current imaging techniques and permit targeted theranostics of microcystic lymphatic malformations.

Keywords: lymphangioma; lymphatic malformation; macrocystic; microcystic; nanoparticles; optical clearing; photoacoustics; photothermal blood and lymph vessel therapy.

FIGURE 1

Large macrocystic lymphatic malformation of the right neck (top). Two-year postoperative follow-up after complete resection of macrocystic lesion (bottom)

FIGURE 2

Diffuse microcystic lymphatic malformation of the tongue. Disease extends into floor of mouth, mandible, and neck

FIGURE 3

Diagnostic imaging of macrocystic LM. (A) US image of a macrocystic lymphatic malformation of the right anterior neck. (B) Axial T2-weighted MRI image of a macrocystic lymphatic malformation of the right anterior neck

FIGURE 4

Principle of in vivo lymph FC. Figure is adapted from ref. [28]

FIGURE 5

In vivo images of mesenteric lymph vessel (left) and cells in lymph flow (right); LV, lymph vessel; BV, blood vessel. Figure is adapted from ref. [28]

FIGURE 6

Effect of a laser pulse (585 nm, 10 ms, 0.5–30 J/cm²) on mesenteric blood and lymph vessels in vivo. (A) Arteries (A) and vein (V) before laser pulse. (B) Destroying of the vein after 1 laser pulse. (C) Lymph vessel (L) before laser pulse (dash lines indicate edges of lymph vessel). (D) Constriction of the same lymph vessel after 1 laser pulse. Figure is adapted from ref. [34]

FIGURE 7

PA detection of GNTs in mesenteric lymph vessels in vivo: (A,B) optical and PA image of the intact (without NPs) lymph vessel; dash lines indicate edges of lymph vessel. (C) PA image of the lymph vessels after introduction of GNTs; free NPs (yellow spots) travel in lymph flow. Figure is adapted from ref. [39]

FIGURE 8

Noninvasive PA (532 nm) image of ear lymph vessel (LV) at third minute after intradermal injection of Evans Blue dye (2–3 μL of 1% solution)

FIGURE 9

In vivo theranostics of mesenteric lymph vessel using bioconjugated GNTs. (A) Workflow diagram of molecular labeling of LECs. (B) PA image of lymph vessel after labeling LECs using GNTs conjugated with antibodies to LYVE-1 receptor; dash lines show edges of lymph vessel, arrow indicates PA signal from cluster of GNTs. (C) Optical image of lymph vessel after laser therapy; red spot is laser beam; dash lines indicate edges of lymph vessel before laser exposure. Laser parameters: 850 nm (maximum absorption of GNTs), 20 mJ/cm² for diagnosis and 60 mJ/cm² for therapy. Figure is adapted from ref. [39]

FIGURE 10

(A) A 6×600-μm linear laser beam propagated through a fresh layer of mouse skin was attenuated ~3-fold and blurred into an ellipsoidal shape ~90 μm wide. (B) Topical administration of integrated skin treatment (glycerol, microdermabrasion, and sonophoresis) during just 10 minutes reduced the influence of scattering light, resulting in an ~1.7-fold decrease in blurring of the laser beam. (E, D: before and after clearing, respectively) Application of this procedure to a human subject’s hand eventually resulted in a 2.1-fold increase in PA signals from a deep 1–2 mm vein. (E, F: before and after clearing, respectively) The same procedure increased the number of PA signals from circulating tumor cells in melanoma – bearing mouse model approximately 3.5–4 times. Figure is adapted from ref. [74]

FIGURE 11

Optical imaging of microanatomy of the fresh mouse lymph node ex vivo obtained with TOC using 80% glycerol water solution. The central schematic designed by David Sabio shows a midsagital section of a lymph node containing 3 lymphoid lobules with the basic anatomical and functional units. Top and middle lobules: microanatomical schematics of lobular compartments (superficial cortex, deep cortex, and medulla) without (top) and with (middle) reticular meshwork. Bottom lobule: a lobule of a mouse lymph node as it appears in conventional histological section. C1 shows likely basophilic lymphocytes; C2 shows elongated fibroblastic reticular cells; and C3 shows B lymphocytes and follicular dendritic cells. Figure is adapted from ref. [30]

FIGURE 12

Visualization of cancer-metastasized lymph node. (A) Illustration of popliteal lymph node metastasis model using B16F10-GFP melanoma cells. (B) Photograph of lymph node with melanoma metastasis immersed in PBS or FocusClear for 12 h. (C) Serial depth images with 100 μm interval. Distribution of metastasized B16F10-GFP melanoma cells (green), blood vessels (CD31, red) and lymph vessels (LYVE-1, blue) are identifiable. (D) Projection and 3D reconstructed images generated by using Z-stack imaging data. (E) Vasculature/lymphatic network around the small-sized colony of GFP expressing cell bodies. Scale bars; (C) 500 μm, (D,E) 100 μm. Figure is adapted from ref. [69]