Label-free photoacoustic microscopy for in-vivo tendon imaging using a fiber-based pulse laser

Abstract

Tendons are tough, flexible, and ubiquitous tissues that connect muscle to bone. Tendon injuries are a common musculoskeletal injury, which affect 7% of all patients and are involved in up to 50% of sportsrelated injuries in the United States. Various imaging modalities are used to evaluate tendons, and both magnetic resonance imaging and sonography are used clinically to evaluate tendons with non-invasive and non-ionizing radiation. However, these modalities cannot provide 3-dimensional (3D) structural images and are limited by angle dependency. In addition, anisotropy is an artifact that is unique to the musculoskeletal system. Thus, great care should be taken during tendon imaging. The present study evaluated a functional photoacoustic microscopy system for in-vivo tendon imaging without labeling. Tendons have a higher density of type 1 collagen in a cross-linked triple-helical formation (65-80% dryweight collagen and 1-2% elastin in a proteoglycan-water matrix) than other tissues, which provides clear endogenous absorption contrast in the near-infrared spectrum. Therefore, photoacoustic imaging with a high sensitivity to absorption contrast is a powerful tool for label-free imaging of tendons. A pulsed near-infrared fiber-based laser with a centered wavelength of 780 nm was used for the imaging, and this system successfully provided a 3D image of mouse tendons with a wide field of view (5 × 5 mm 2 ).Tendons are tough, flexible, and ubiquitous tissues that connect muscle to bone and transfer the force that is generated by muscle movement to the bone 1 . For example, muscle contraction pulls the tendon and transfers force to the connected bone. Thus, tendons play an important role in muscle-bone control of the body. Tendon injuries are a common musculoskeletal injury that affect 7% of all patients and are involved in up to 50% of sports-related injuries in the United States [2][3][4] . Various imaging modalities can be used to obtain tendon images, and both magnetic resonance imaging (MRI) 5-8 and sonography 8,9 are used clinically for tendon imaging with non-invasive and non-ionizing radiation. In addition, optical coherence tomography (OCT) has been used for tendon imaging in preclinical studies 10,11 . In the MRI sequences, normal tendons have low signal intensity because the alignment of water and collagen molecules in the tendon structure generates dipole interactions that considerably shorten the T 2 relaxation time to 1-2 ms [12][13][14] . In addition, tendons generate a characteristic angle-dependent MRI signal. The T 2 begins to increase when the angle between the tendon and the magnetic field is above 20° and maximized around 55° (the "magic angle" phenomenon) [14][15][16][17][18] . Collagen orientation of the tendon at this "magic angle" shows higher MRI signal intensity, depending on the type of sequence and the echo time. Therefore, the orientation change of tendon fibers can cause signal degradation and lead to a misinterpreted diagnosis. This problem can be reduced using a long echo time and s...

Dual-Modal Photoacoustic Imaging and Optical Coherence Tomography [Review]

“…However, this apparent difference in wavelength, does not impede applying visible light for OCT or NIR light for PAM. Recent experiments have demonstrated applications of visible OCT for high-resolution imaging and measuring metabolic rate of oxygen for clinical studies [134,135], while NIR light has been used for imaging lipid and collagen tissues in PAM [136][137][138]. Several studies explored the feasibility of using a single light source for PAM excitation and OCT imaging, which would reduce the complexity and costs of the system; in addition, it will generate synchronized and coregistered PAM and OCT images.…”

Towards non-contact photoacoustic imaging [review]

“…Reprinted with permission from [ 36 ] (B) photoacoustic images acquired from mouse small intestine cells and DNA contrast at 266 nm. Reprinted with permission from [ 79 ], fibroblast cytoplasm’s with cytochrome contrast at 422 nm, reprinted with permission from [ 80 ], PA image of mouse blood smear with hemoglobin contrast at 532 nm, reprinted with permission from [ 81 ], tyrosinase tumor expressing with melanin contrast at 680 nm, reprinted with permission from [ 12 ], photoacoustic image of mouse paw with skin removed and collagen contrast at 780 nm, reprinted with permission from [ 82 ], intramuscular fat photoacoustic image with lipid contrast at 1197 nm, reprinted with permission from [ 83 ]. …”

Multiangle Long-Axis Lateral Illumination Photoacoustic Imaging Using Linear Array Transducer

“…The average RF signal of the PA subframes at depths and in lateral direction were evaluated using distinct regions of interest (ROI). Based on the number of elements of the transducer (i.e., 128 elements), we defined a central ROI-1 within FOV, which included the five central elements (62)(63)(64)(65)(66) and extended from the phantom surface to the maximum depth, with dimensions 1.5 mm × 25.5 mm. Also, peripheral ROIs (ROI-2 and ROI-3) were defined including two sets of five elements positioned at opposite sides of transducer elements: 5-9 (ROI-2) and 119-123 (ROI-3).…”

“…At a tissue depth of 7.3 mm, ROI overlaid the tendon of flexor digitorum superficialis [63] and provided SNR = 14 dB (green rectangle in Figure 11a), while another ROI overlaid a subcutaneous blood vessel and provided SNR = 25 dB at a 2.5 mm tissue depth (cyan rectangle in Figure 11a). Differences in the SNR values between deep and shallow regions were mostly due to light attenuation; the tendon (collagen) also presents an optical absorption coefficient lower than those for melanin or hemoglobin at 800 nm [64], which reduced its SNR on the PA image. The second acquisition of PA images was obtained from the human index finger, which has a cylindrical-like shape that differs from the forearm or phantom's surface and has a maximum lateral extension of approximately 20 mm.…”