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Biomedical Imaging and Instrumentation

Biomedical Imaging and Instrumentation Undergraduate Fire Talks

Advancing Pre-Transplant Liver Assessment with Photoacoustic Imaging and Ultrasound Localization Microscopy

Saturday, October 11, 2025
8:35 AM - 8:40 AM PDT
Location: Room 30D

Introduction: : End-stage liver disease is among major causes of death globally, with transplantation being the only cure [1]. However, the demand for donor livers far exceeds supply, partly due to the high discard rate of viable organs - approximately 76% of potential donor livers are rejected in the United States [2]. While biopsy remains the clinical gold standard for evaluating liver viability, it is invasive, prone to sampling errors, and provides limited spatial information [3-5]. Existing imaging modalities either lack sufficient molecular detection depth or are incompatible with the time-sensitive demands of pre-transplant workflows. [6,7]. Photoacoustic imaging (PAI), combining optical absorption and ultrasonic detection, offers a promising solution. PAI is inherently sensitive to endogenous chromophores, including lipids and collagen, providing crucial information on liver steatosis and fibrosis without external contrast agents [8,9]. Meanwhile, ultrasound localization microscopy (ULM) enables super-resolution blood flow imaging [10]. Recently, normothermic machine perfusion (NMP) is increasingly adopted in clinical practice for extended organ preservation [11]. Thus, advanced spatial monitoring methods are needed for comprehensive viability evaluation during perfusion. Combined with these emerging perfusion systems, PAI and ULM offer comprehensive pre-transplant liver viability quantification, from organ recovery to perfusion. This study reports the first application of PAI and ULM for evaluating human livers pre-transplantation. Our multispectral PAI method successfully images collagen and lipid in phantom experiments and ex vivo human liver tissues, validated by pathology. PAI noninvasively tracks blood oxygenation in deep tissue, while ULM monitors microvasculature perfusion and blood flow velocity during organ perfusion.

Materials and

Methods: : Human liver specimens were obtained from LifeShare of Oklahoma within 24 hours of retrieval and prepared under IRB-approved protocol. A total of nine livers were imaged using a dual-modality PA/ultrasound system (Vevo F2 LAZR-X, Fujifilm VisualSonics). As shown in Figure 1a, the assessment workflow began with procurement, followed by transportation (Fig. 1b-i, ii) and imaging setup (Fig. 1b-iii). The PAI system used a tunable laser (680-970 nm, 10-ns pulse width, 20-Hz repetition rate, 50 mJ at 750 nm), with energy maintained at 16.7 mJ/cm² for safety [12]. Five wavelengths (680, 730, 860, 926, 970 nm) were selected based on optical absorption spectra of collagen, lipid, hemoglobin (oxy-/deoxy-), and water [13]. A 20-MHz linear-array transducer (UHF29x) mounted on a translational stage scanned ~75% of the liver cortex at 0.15 mm steps (Fig. 1c-d). A custom-built water chamber ensured acoustic coupling and consistent imaging surfaces. PAI datasets were reconstructed in VEVOLAB to estimate collagen and lipid scores via spectral unmixing [14].
To validate performance, tissue-mimicking phantoms were constructed using gelatin (collagen surrogate), butter (lipid), and oxygen-modulated bovine blood [15]. For perfused liver imaging, PAI and ULM were integrated with a normothermic machine perfusion (NMP) system (Fig. 1e-f), maintaining physiological temperatures (35-37°C) and pressures [16]. ULM imaging involved microbubble injection, SVD-based clutter filtering [17], deconvolution-based localization [18], and microbubble tracking using the u-Track algorithm [19]. Histological validation (H&E, Masson's trichrome, PAS) was performed on marked regions, with blinded scoring by three board-certified pathologists [20,21].

Results, Conclusions, and Discussions::

Results:
PAI distinguished lipid and collagen in phantoms with strong linearity (R² > 0.94; Fig. 2). In human livers, spatially heterogeneous fibrosis and steatosis were visualized (Fig. 3) and validated by histology. During perfusion, ULM mapped microvasculature ( < 50 μm) and PAI monitored real-time deep-tissue oxygenation.

Conclusions:
This study presents the first successful integration of PAI and ULM for comprehensive human liver evaluation. PAI enables label-free quantification of fibrosis and steatosis, while ULM offers super-resolution visualization of microvascular perfusion during normothermic machine perfusion. Together, these modalities offer rapid, volumetric, real-time assessment of liver viability, addressing key limitations of biopsy and conventional imaging. Strong correlation with histopathology confirms diagnostic accuracy. Clinically, this approach could reduce organ discard rates, support informed transplant decisions, and improve graft outcomes. Its compatibility with current perfusion workflows underscores its translational readiness and potential to impact donor organ utilization globally.

Discussions:
Our findings demonstrate that PAI-ULM offers superior advantages as a non-invasive alternative to traditional biopsy for liver viability assessment. PAI accurately quantifies lipid and collagen content, addressing the limitations of sampling errors and spatial heterogeneity inherent to biopsies. ULM complements this by mapping microvascular perfusion at super-resolution, crucial for evaluating graft functionality during normothermic machine perfusion. The strong agreement between imaging results and histopathology confirms the reliability of this dual-modal platform. Future studies will focus on optimizing imaging speed, enhancing penetration depth, and validating clinical applicability in larger donor cohorts.

Acknowledgements and/or References (Optional): : 1. Asrani SK, Devarbhavi H, Eaton J, et al. Burden of liver diseases in the world. Journal of hepatology 2019;70:151-171.
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15. Seaton B, Lloyd BB. The effects of pH on the equilibrium constants of various models for the haemoglobin-oxygen equilibrium in vitro. Respiration Physiology 1974;20:209-230.
16. Kneifel F, Wagner T, Flammang I, et al. Hyperspectral imaging for viability assessment of human liver allografts during normothermic machine perfusion. Transplantation Direct 2022;8:e1420.
17. Tang Y, Wang N, Dong Z, et al. Non-invasive deep-brain imaging with 3D integrated photoacoustic tomography and ultrasound localization microscopy (3D-PAULM). IEEE transactions on medical imaging 2024.
18. Dencks S, Lowerison M, Hansen-Shearer J, et al. Super-resolution ultrasound: From data acquisition and motion correction to localization, tracking, and evaluation. IEEE Transactions on Ultrasonics, Ferroelectrics, and Frequency Control 2025.
19. Jaqaman K, Loerke D, Mettlen M, et al. Robust single-particle tracking in live-cell time-lapse sequences. Nature methods 2008;5:695-702.
20. Clark I, Torbenson MS. Immunohistochemistry and special stains in medical liver pathology. Advances in anatomic pathology 2017;24:99-109.
21. Lefkowitch JH. Scheuer's Liver Biopsy Interpretation E-Book: Elsevier Health Sciences, 2020.