Journal of Biomedical Optics最新文献

Significance: Dynamic optical coherence elastography can excite and detect propagating mechanical waves in soft tissue without physical contact and in near real time. However, most soft tissue is anisotropic, characterized by at least three independent elastic moduli. As a result, reconstructing these moduli from mechanical wave fields requires a complex procedure.

Aim: We consider a nearly incompressible transverse isotropic (NITI) material, which has been shown to locally define the symmetry of many soft tissues such as muscle, tendon, skin, cornea, heart, and brain. Reconstruction of elastic moduli in the NITI medium using Rayleigh waves is addressed here. A method to accurately compute the angular dependence of Rayleigh wave phase velocity for the most common geometries (point-like and line sources) of mechanical wave excitation is described.

Approach: When a line source is used to launch plane mechanical waves over the medium surface, the phase velocity of Rayleigh waves in the direction of propagation is directly accessible. For a point-like source, propagation of the energy flux is tracked (i.e., its group velocity), which cannot be directly used for moduli inversion. In this case, angular spectrum decomposition is used to access the phase velocity. Both numerical simulations in OnScale and experiments in a stretched PVA phantom were performed.

Results: We show that both methods (line source wave excitation and angular decomposition from a point-like source) produce similar results and accurately estimate the angular anisotropy of the Rayleigh wave phase velocity. We also explicitly show that a commonly used group velocity approach leads to inadequate moduli inversion and should not be used for reconstruction.

Conclusions: We suggest that the line source is best when a surface area must be scanned, whereas the point-like source with the proposed phase velocity reconstruction is best for single-point moduli estimation or when tissue motion is a concern.

Significance: Intralipid, a soybean oil-based lipid emulsion, is widely used in photomedicine to enhance light distribution due to its strong scattering properties. Although the optical characteristics of Intralipid are well documented, interactions with the reactive molecular species (RMS) generated during photodynamic therapy (PDT) and the impact of such interactions on therapeutic outcomes remain poorly understood. We reveal that Intralipid actively influences PDT response in vitro, beyond its role as a scattering agent.

Aim: We examined how Intralipid affects the optical and photodynamic behavior of benzoporphyrin derivative (BPD), a clinical photosensitizer, in solution and across four ovarian cancer cell lines.

Approach: The photodynamic properties of BPD, with and without Intralipid, were analyzed using fluorescence spectrometry and RMS probes, and PDT-induced oxidation of Intralipid components was characterized using LC-MS. The effects of Intralipid on BPD-PDT were evaluated under various conditions.

Results: Intralipid reduced BPD photobleaching and RMS generation, suggesting RMS quenching. Extensive oxidation of Intralipid components was observed following PDT. Finally, Intralipid significantly modified BPD-PDT efficacy across all four cell lines, depending on photosensitizer-light interval, dose, and incubation time.

Conclusions: Intralipid acts as a bioactive modulator of PDT response, highlighting the need for further investigations both in vitro and in vivo.

Significance: Noninvasive imaging to accurately measure subtle changes in tumor size is underutilized when assessing therapeutic responses in the skin. During photodynamic therapy (PDT) for basal cell carcinoma (BCC), a better definition of the tumor size threshold for PDT responsiveness is needed.

Aim: We aim to quantitatively demonstrate the first clinical evidence of tumor shrinkage after multiple rounds of PDT using a robust measurement and analysis approach.

Approach: Tumors were monitored experimentally using a 3D camera and software system (stereo photogrammetry). A total of 122 BCC tumors in 35 patients were treated with PDT (5-ALA and blue light) in three sessions. Calculated volumes and heights were used to plot changes in tumor size.

Results: In total, 70% of BCC cleared completely. Measured heights correlated with histological tumor depth; average heights were $\sim 10\%$ to 20% of actual tumor depth. From photogrammetry at baseline, an average height of $<0.15\mathrm{mm}$ was found to predict a complete therapeutic response. Thus, our 3D morphometric technique provides a surrogate measure of BCC tumor depth that predicts PDT response and is accurate to well below the millimeter level.

Conclusions: 3D photogrammetry can inform the selection of BCC tumors for PDT with exceptionally high spatial accuracy, dramatically better than can be quantified by a clinician.

Significance: Glaucoma is a leading cause of irreversible blindness, characterized by progressive optic nerve damage. Early detection of glaucoma is key to effective intervention, but an incomplete clinical understanding of the development of glaucoma complicates the selection of diagnostic criteria. Prolonged ocular hypertension due to glaucoma can impact the biomechanical properties of ocular tissues, including the cornea. We examine whether experimental glaucoma causes changes in the biomechanical properties of the cornea.

Aim: We determined the biomechanical properties of the cornea in a nonhuman primate model of unilateral experimental glaucoma and compared them with the fellow, untreated control eyes using optical coherence elastography (OCE) to determine if prolonged intraocular pressure (IOP) elevation causes changes in corneal stiffness.

Approach: Experimental glaucoma was induced in one eye (Macaca mulatta, $N=2$ ) by lasering the trabecular meshwork, whereas the fellow eye was used as a control. Both eyes were imaged with wave-based OCE to investigate the inter-ocular difference in stiffness. Measurements were taken at three different frequencies with quasi-harmonic excitation, and central corneal thickness was measured along with IOP in each eye.

Results: Our results show a significant ( $p<0.01$ ) increase in wave speed in the experimental glaucoma eye compared with the control eye for both subjects.

Conclusions: These results show the potential of wave-based OCE methods for assessing stiffness changes in the cornea caused by remodeling due to chronic pressure elevation.

Significance: 5-Aminolevulinic acid (5-ALA) is a medical pro-drug used to induce the intracellular production of protoporphyrin IX (PpIX) via the heme synthesis pathway. Discoveries in mechanisms and developments in novel applications still continue with this uniquely endogenous intracellular optical system.

Aim: Understanding and exploiting the growing uses can be advanced through a survey of knowledge on the mechanisms and biokinetics of 5-ALA administration, partitioning, PpIX production, localization changes, clearance mechanisms, biological interactions, and methods for unique activation methods in both diagnostic and therapeutic applications.

Approach: The current medical uses of PpIX are reviewed, separating into therapeutic and diagnostic areas, and the expansion and lateral growth areas are outlined.

Results: Initially approved for photodynamic therapy of skin lesions, fluorescence diagnostic indications later developed to guide surgical resection in bladder cancer and glioma. Today, the 5-ALA-PpIX system's spatial-temporal complexity in photophysics and pharmacokinetics continues to lead to more uses, such as photodynamic priming to alter tissue, fast intracellular tissue oxygen sensing, infection, and burn imaging and therapy.

Conclusions: The 5-ALA-PpIX system has broad potential partly because of the ubiquity of the heme synthesis across many cell/tissue types, combined with natural selectivity, unique pharmacokinetics, bright fluorescence, and sufficiently strong singlet oxygen production.

Significance: Oral cancer surgery demands precise margin delineation to ensure complete tumor resection (healthy tissue margin $>5\mathrm{mm}$ ) while preserving postoperative functionality. Inadequate margins most frequently occur at the deep surgical margins, where tumors are located beneath the tissue surface; however, current fluorescent optical imaging systems are limited by their inability to quantify subsurface structures. Combining structured light techniques with deep learning may enable intraoperative margin assessment of 3D surgical specimens.

Aim: A deep learning (DL)-enabled spatial frequency domain imaging (SFDI) system is investigated to provide subsurface depth quantification of fluorescent inclusions.

Approach: A diffusion theory-based numerical simulation of SFDI was used to generate synthetic images for DL training. ResNet and U-Net convolutional neural networks were developed to predict margin distance (subsurface depth) and fluorophore concentration from fluorescence images and optical property maps. Validation was conducted using in silico SFDI images of composite spherical harmonics, as well as simulated and phantom datasets of patient-derived tongue tumor shapes. Further testing was done in ex vivo animal tissue with fluorescent inclusions.

Results: For oral cancer optical properties, the U-Net DL model predicted the overall depth, concentration, and closest depth with errors of $1.43\pm 1.84\mathrm{mm}$ , $2.26\pm 1.63\mu g/\mathrm{ml}$ , and $0.33\pm 0.31\mathrm{mm}$ , respectively, using in silico patient-derived tongue shapes with closest depths below 10 mm. In PpIX fluorescent phantoms of inclusion depths up to 8 mm, the closest subsurface depth was predicted with an error of $0.57\pm 0.38\mathrm{mm}$ . For ex vivo tissue, the closest distance to the fluorescent inclusions with depths up to 6 mm was predicted with an error of $0.59\pm 0.53\mathrm{mm}$ .

Conclusions: A DL-enabled SFDI system trained with in silico images demonstrates promise in providing margin assessment of oral cancer tumors.

Significance: As fluorescence-guided surgery (FGS) gains clinical adoption, robust and experimentally validated computational models for tissue fluorescence are increasingly essential. Although there have been several developments in modeling fluorescence with Monte Carlo simulations, the scope of the experimental validation has been limited in the parameters tested and phantoms used.

Aim: We aim to present and experimentally validate a graphics processing unit (GPU)-accelerated, voxel-based Monte Carlo fluorescence framework capable of modeling varying fluorophore concentrations, optical properties, and complex three-dimensional (3D) geometries.

Approach: A two-step approach (MCX-ExEm) based on Monte Carlo eXtreme was developed for simulating fluorescence. Both commercial reference targets and custom 3D-printed phantoms with well-characterized optical properties were imaged for varying parameters-including absorption, scattering, fluorophore concentrations, and geometries-and compared against simulations.

Results: Strong agreement is observed between simulated and experimental fluorescence across all tested parameters. MCX-ExEm accurately captures nonlinear quenching at high fluorophore concentrations, variations driven by scattering and absorption, intensity scaling with volume, and depth-dependent attenuation and resolution. Minor deviations occur primarily under low-scattering or low-absorption regimes, where optical characterization presents greater uncertainties.

Conclusions: By integrating experimentally validated simulations with a broad range of solid phantoms, this framework establishes a foundation for developing fluorescence digital twins, enabling faster and more systemic testing of fluorescence imaging systems. These findings can help accelerate the design and optimization of FGS and other fluorescence-based biomedical applications.

Significance: Molecular photoacoustic (PA) imaging with exogenous dyes faces a significant challenge due to the photobleaching of the dye that can compromise tissue visualization, particularly in 3D imaging. Addressing this limitation can revolutionize the field by enabling safer, more reliable imaging and improve real-time visualization, quantitative analysis, and clinical decision-making in various molecular PA imaging applications such as image-guided surgeries.

Aim: We tackle photobleaching in molecular PA imaging by introducing a platform-flexible deep learning framework that enhances SNR from single-laser pulse data, preserving contrast and signal integrity without requiring averaging of signals from multiple laser pulses.

Approach: The generative deep learning network was trained with an LED-illuminated PA image dataset and tested on acoustic resolution PA microscopy images obtained with single-laser pulse illumination. In vitro and ex vivo samples were first tested for demonstrating SNR improvement, and then, a 3D-scanning experiment with an ICG-filled tube was conducted to depict the usability of the technique in reducing the impact of photobleaching during PA imaging.

Results: Our generative deep learning model outperformed traditional nonlearning, filter-based algorithms and the U-Net deep learning network when tested with in vitro and ex vivo single pulse-illuminated images, showing superior performance in terms of signal-to-noise ratio ( $93.54\pm 6.07$ , and $92.77\pm 10.74$ compared with $86.35\pm 3.97$ , and $84.52\pm 11.82$ with U-Net for kidney, and tumor, respectively) and contrast-to-noise ratio ( $11.82\pm 4.42$ , and $9.9\pm 4.41$ compared with $7.59\pm 0.82$ , and $6.82\pm 2.12$ with U-Net for kidney, and tumor respectively). The use of cGAN with single-pulse rapid imaging has the potential to prevent photobleaching ( $9.51\pm 3.69\%$ with cGAN, and $35.14\pm 5.38\%$ with long-time laser exposure by averaging 30 pulses), enabling accurate, quantitative imaging suitable for real-time implementation, and improved clinical decision support.

Conclusions: We demonstrate the potential of a platform-flexible generative deep learning-based approach to mitigate the effects of photobleaching in PA imaging by enhancing signal-to-noise ratio from single pulse-illuminated data, the

Significance: In previous studies, we reported on a new untargeted contrast agent, TMR-PEG1k, that exhibits high and durable diagnostic performance in glioma models starting within minutes of administration. This agent uses a fluorophore in the visible regime, which, although it helps ensure high-resolution imaging of superficial tissue, precludes the detection of subsurface structures due to the limited optical penetration. Thus, development of a near-infrared (NIR) version of the agent that retains the properties of TMR-PEG1k would combine the favorable diagnostic performance of TMR-PEG1k with sensitivity to subsurface pathologies enabled by NIR imaging.

Aim: We aim to determine whether exchanging the fluorophore on TMR-PEG1k from tetramethylrhodamine to cyanine 7 would retain the highly favorable imaging properties exhibited by TMR-PEG1k.

Approach: Mice ( $N=5$ ) with orthotopic gliomas expressing green fluorescent protein (GFP) were co-administered TMR-PEG1k and Cy7-PEG1k. After 30 min, animals were euthanized, and the whole-body specimens were imaged using fluorescence cryotomography to recover co-registered three-dimensional (3D) fluorescence distributions of all fluorescent reporters. We evaluated the two agents using tumor-to-background ratio (TBR), contrast-to-noise ratio (CNR), area under the receiver operating characteristic curve (ROC-AUC), and normalized cross-correlation with GFP fluorescence (CC to GFP).

Results: Although the imaging system exhibited higher sensitivity to Cy7-PEG1k in phantoms, the 3D cryotomography results showed dramatic differences in the properties of the two agents in vivo. TMR-PEG1k produced highly selective tumor enhancement and concordance with GFP, whereas Cy7-PEG1k showed much lower selectivity, lower signal intensity, and produced no enhancement in many tumor regions. These observations were confirmed by the evaluation metrics, which were found to be (1) TBR = 5.2 versus 1.7 ( $p=0.0037$ ), (2) CNR = 17.7 versus 3.8 ( $p=0.046$ ), (3) ROC-AUC = 0.999 versus 0.821, and (4) CC to GFP = 0.90 versus 0.52, for TMR-PEG1k versus Cy7-PEG1k, respectively.

Conclusions: Cy7-PEG1k did not retain the favorable properties exhibited by TMR-PEG1k and thus is not a suitable NIR analog for this agent.

Significance: Confocal Raman spectroscopy (CRS) is a noninvasive technique that enables detailed biochemical analysis of the skin, providing insights into its structural and molecular composition. Atopic dermatitis and hand eczema, both characterized by impaired skin barrier function, require effective monitoring tools to assess treatment efficacy.

Aim: We aimed to quantitatively evaluate changes in skin physiological and biochemical parameters following the application of a ceramide-based moisturizer (test cream) compared with an aqueous moisturizer (control cream) in healthy volunteers and eczema patients.

Approach: Skin physiological assessments and CRS measurements were performed to evaluate water content and ceramide levels in both superficial and deeper skin layers after application of the two moisturizers.

Results: CRS revealed a significant increase in water content following test cream application, indicating improved skin hydration, whereas physiological measurements detected no statistically significant changes, underscoring the greater sensitivity of CRS. The test cream also significantly enhanced ceramide levels in both superficial and deeper skin layers, whereas the control cream increased ceramide levels only in the superficial stratum corneum (SC).

Conclusions: These findings suggest that the test cream has a substantial positive impact on hand eczema, potentially improving both skin barrier function and biochemical properties.