Simultaneous functional photoacoustic and ultrasonic endoscopy of internal organs in vivo

Abstract

At present, clinicians routinely apply ultrasound endoscopy in a variety of interventional procedures that provide treatment solutions for diseased organs. Ultrasound endoscopy not only produces high-resolution images, but also is safe for clinical use and broadly applicable. However, for soft tissue imaging, its mechanical wave-based image contrast fundamentally limits its ability to provide physiologically specific functional information. By contrast, photoacoustic endoscopy possesses a unique combination of functional optical contrast and high spatial resolution at clinically relevant depths, ideal for imaging soft tissues. With these attributes, photoacoustic endoscopy can overcome the current limitations of ultrasound endoscopy. Moreover, the benefits of photoacoustic imaging do not come at the expense of existing ultrasound functions; photoacoustic endoscopy systems are inherently compatible with ultrasound imaging, thereby enabling multimodality imaging with complementary contrast. Here we present simultaneous photoacoustic and ultrasonic dual-mode endoscopy and show its ability to image internal organs in vivo, thus illustrating its potential clinical application.

Figure 1

Illustration of simultaneous, multi-wavelength photoacoustic (PA) and ultrasonic (US) endoscopy. (a) The endoscope performs circumferential sector scanning by rotating a scanning mirror, which reflects both the US waves and laser pulses and enables static mounting of the associated illumination and US pulse generation-detection units. At each angular step of the mirror (~1.42°), both the first (λ₁) and second (λ₂) pulsed laser beams are independently fired through the optical fiber and the acoustic pulse is generated by the US transducer with a constant time delay of ~30 µs between each of the laser and acoustic pulses. The ensuing PA and US echo waves are detected and converted into electric signals by the US transducer; the signals are then recorded and displayed on a computer. The 30-µs time delay is necessary to ensure that the sample has relaxed and that consecutive signals will not interfere with each other. However, this delay does not significantly affect co-registration of the three images because the difference between the angular positions of each signal is small and is well within the spatial resolution of the detector. (b) A photo shows the side-scanning 3.8-mm diameter probe prototype firing a 562 nm laser beam. Scale bar, 2 cm. (c) Definition of Cartesian and cylindrical coordinate systems. The +z-axis is defined along the endoscope axis (or pullback direction). (d) A volumetric image comprised of consecutive B-scan slices. (e) A representative cross-section of d along the x-y plane, which shows the endoscope’s 270° angular FOV.

Figure 2

Simultaneous, co-registered, PAE-EUS pseudo-color images from a rabbit esophagus in vivo. (a) Three-dimensionally rendered PA structural image. The left-and right-hand sides of this image correspond to the lower and upper esophagus, respectively, and the lower portion (–y axis) to the ventral side of the animal. To more clearly display the structures surrounding the esophagus, we excluded signals generated from the esophagus. (b) Co-registered US structural image for the same volume of a. (c) An overlaid image of a and b. In a–c, the horizontal and vertical scale bars represent 2 cm and 5 mm, respectively. (d) A representative PA x-y cross-sectional image (18 mm diameter) near the lung, as indicated by the left arrow in a. The ~0.8 mm thick region from the inner surface of the imaged lumen represents the signals from the esophagus. The surface of the lung is shown ~1 mm from the inner surface. (e) Corresponding US cross-sectional image of d. (f) A combined image of d and e. (g) A representative PA x-y cross-sectional image (18 mm diameter) near the trachea, as indicated by the right arrow in a. (h) Corresponding US cross-sectional image of g. In d–h, the hash marks represent 1 mm intervals. (i) Representative histology (H&E stain) of the esophagus (upper) and trachea (lower). Scale bar, 1 mm.

Figure 3

Radial-maximum amplitude projection (RMAP) images over a full 360° angular FOV (views from the inside of the esophagus). The left-and right-hand sides of these three images correspond to the lower and upper esophagus, respectively. (a) Normalized PA-RMAP image showing the total hemoglobin distribution, with the esophageal signals excluded during the RMAP construction. AL, accessory lobe; LL, left lobe; RL, right lobe of the lung; AO, aorta; CVC, caudal vena cava; CA, carina; TC, trachea. (b) Functional PA-RMAP image showing the sO₂ levels of the imaged structures in a. (c) Normalized US-RMAP image showing the echogenicity distribution. In each image, the vertical φ-axis corresponds to the angular range of 360°, and the horizontal z-axis corresponds to the pullback length of 14 cm. The approximate mid-ventral (MV) position and angular measures from the MV are marked along the vertical φ-axis, where the positive and negative values correspond to the right and left sides of the animal, and MD denotes the mid-dorsal position. The scale bars represent 1 cm for the horizontal direction only.

Figure 4

Simultaneous, co-registered, PAE-EUS colonoscopic pseudo-color images of a rat colon acquired in vivo. (a) Three-dimensionally rendered PA-US structural image. The right side of this image is closer to anus, and the negative y-axis corresponds to the ventral direction of the animal. The red and green colors correspond to PA and US signals, respectively. The dashed arrow indicates mesenteric tissue entangled around the tract, and SP denotes the sphincter. The horizontal and vertical scale bars represent 1 cm and 5 mm, respectively. (b) PA-RMAP image showing the sO₂ levels of the imaged structures in a (views from the inside of the colon). (c) Corresponding US-RMAP image showing the echogenicity distribution. In b and c, the vertical φ-axis corresponds to the angular FOV covering 270°, and the horizontal z-axis corresponds to the pullback length of 5.5 cm. The approximate mid-ventral (MV) position and angular measures from the MV are marked along the vertical φ-axis, where the positive and negative values correspond to the right and left sides of the animal. The scale bars represent 1 cm for the horizontal direction only. (d,e) PA-US cross-sectional images from the position indicated by the left and right arrows in a, respectively. In d and e, the solid arrows indicate the outer boundary of the colon, and mesenteric tissues are marked with dashed arrows. The hash marks represent 1 mm intervals. (f) A typical histology image (H&E stain) of the colon. Scale bar, 1 mm.

Figure 5

Lymphovascular system imaging in vivo near a rat colon. (a) Control PA-RMAP image before EB administration (views from the inside of the colon). The right side of this image is closer to the anus. (b) Co-registered US-RMAP image of a. (c) PA-RMAP image at ~40 min post-injection (processed from the dual-wavelength data). The red and blue colors represent PA signals from blood and EB, respectively. (d) Corresponding US-RMAP image of c. In a–d, the vertical φ-axis corresponds to the angular FOV covering 270°, and the horizontal z-axis corresponds to the pullback length of 7.2 cm. The approximate mid-dorsal (MD) position and angular measures from the MD are marked along the vertical φ-axis. (e) Merged volumetric image of c and d. SP, sphincter; CW, cavity wall. (f) A representative x-y cut near the location indicated by the arrow in e. The blue-colored regions represent a lymph node. (g) A spectrally-processed PA-RMAP image from the dataset shown in c, with the colon signals excluded. This image shows the lymph structures and sO₂ distribution of the surround vasculature outside the colon. The arrows indicate lymph vessels (LV), and the dashed circle corresponds to the left lumbar node (LN) in h. CVC, caudal vena cava; CIV, left common iliac vein. (h) A post-imaging surgical photo showing the stained lymph node and vessels located outside the colon. This area approximately corresponds to the yellow dashed rectangle in g. The horizontal and vertical scale bars in the images represent 1 cm and 5 mm, respectively.