Multiple scattering suppression for in vivo optical coherence tomography measurement using the B-scan-wise multi-focus averaging method.

We demonstrate a method that reduces the noise caused by multi-scattering (MS) photons in an in vivo optical coherence tomography image. This method combines a specially designed image acquisition (i.e., optical coherence tomography scan) scheme and subsequent complex signal processing. For the acquisition, multiple cross-sectional images (frames) are sequentially acquired while the depth position of the focus is altered for each frame by an electrically tunable lens. In the signal processing, the frames are numerically defocus-corrected, and complex averaged. Because of the inconsistency in the MS-photon trajectories among the different electrically tunable lens-induced defocus, this averaging reduces the MS signal. Unlike the previously demonstrated volume-wise multi-focus averaging method, our approach requires the sample to remain stable for only a brief period, approximately 70 ms, thus making it compatible with in vivo imaging. This method was validated using a scattering phantom and in vivo unanesthetized small fish samples, and was found to reduce MS noise even for unanesthetized in vivo measurement.

Multimodal imaging analysis of autosomal recessive bestrophinopathy: Case series.

RATIONALE: Autosomal recessive bestrophinopathy (ARB) is a subtype of bestrophinopathy caused by biallelic mutations of the BEST1 gene, which affect the retinal pigment epithelium (RPE). Studying RPE abnormalities through imaging is essential for understanding ARB. This case series involved the use of multimodal imaging techniques, namely autofluorescence (AF) imaging at 488 nm [short-wavelength AF] and 785 nm [near-infrared AF (NIR-AF)] and polarization-sensitive optical coherence tomography (PS-OCT), to investigate RPE changes in 2 siblings with ARB. PATIENT CONCERNS: Two Japanese siblings (Case 1: male, followed for 20-23 years; Case 2: female, followed for 13-17 years) carried compound heterozygous mutations of the BEST1 gene. DIAGNOSIS: Both siblings were diagnosed with ARB. INTERVENTIONS AND OUTCOMES: Multimodal imaging techniques were used to evaluate RPE changes. Both siblings had funduscopic changes similar to those seen in the vitelliruptive stage of Best vitelliform macular dystrophy during the follow-up period. NIR-AF imaging showed hypo-AF of the entire macular lesion in both cases, and this hypo-AF remained stable over time. PS-OCT confirmed reduced RPE melanin content in these hypo-AF areas. Additionally, hyper-NIR-AF dots were observed within hypo-NIR-AF areas. Concomitant identification of focally thickened RPE melanin on PS-OCT imaging and hyper-AF on short-wavelength AF imaging at the sites containing hyper-NIR-AF dots indicated that the hyper-NIR-AF dots had originated from either stacked RPE cells or RPE dysmorphia. LESSONS: We confirmed RPE abnormalities in ARB, including diffuse RPE melanin damage in the macula alongside evidence of RPE activity-related changes. This case series demonstrates that multimodal imaging, particularly NIR-AF and PS-OCT, provides detailed insights into RPE alterations in ARB.

Optical-coherence-tomography-based deep-learning scatterer-density estimator using physically accurate noise model.

We demonstrate a deep-learning-based scatterer density estimator (SDE) that processes local speckle patterns of optical coherence tomography (OCT) images and estimates the scatterer density behind each speckle pattern. The SDE is trained using large quantities of numerically simulated OCT images and their associated scatterer densities. The numerical simulation uses a noise model that incorporates the spatial properties of three types of noise, i.e., shot noise, relative-intensity noise, and non-optical noise. The SDE's performance was evaluated numerically and experimentally using two types of scattering phantom and in vitro tumor spheroids. The results confirmed that the SDE estimates scatterer densities accurately. The estimation accuracy improved significantly when compared with our previous deep-learning-based SDE, which was trained using numerical speckle patterns generated from a noise model that did not account for the spatial properties of noise.

Neural-network based high-speed volumetric dynamic optical coherence tomography.

We demonstrate deep-learning neural network (NN)-based dynamic optical coherence tomography (DOCT), which generates high-quality logarithmic-intensity-variance (LIV) DOCT images from only four OCT frames. The NN model is trained for tumor spheroid samples using a customized loss function: the weighted mean absolute error. This loss function enables highly accurate LIV image generation. The fidelity of the generated LIV images to the ground truth LIV images generated using 32 OCT frames is examined via subjective image observation and statistical analysis of image-based metrics. Fast volumetric DOCT imaging with an acquisition time of 6.55 s/volume is demonstrated using this NN-based method.

Erratum: Deep convolutional neural networks-based scatterer density and resolution estimators in optical coherence tomography: erratum.

[This corrects the article on p. 168 in vol. 13, PMID: 35154862.].

Label-free visualization and quantification of the drug-type-dependent response of tumor spheroids by dynamic optical coherence tomography.

We demonstrate label-free dynamic optical coherence tomography (D-OCT)-based visualization and quantitative assessment of patterns of tumor spheroid response to three anti-cancer drugs. The study involved treating human breast adenocarcinoma (MCF-7 cell-line) with paclitaxel (PTX), tamoxifen citrate (TAM), and doxorubicin (DOX) at concentrations of 0 (control), 0.1, 1, and 10 µM for 1, 3, and 6 days. In addition, fluorescence microscopy imaging was performed for reference. The D-OCT imaging was performed using a custom-built OCT device. Two algorithms, namely logarithmic intensity variance (LIV) and late OCT correlation decay speed (OCDS[Formula: see text]) were used to visualize the tissue dynamics. The spheroids treated with 0.1 and 1 µM TAM appeared similar to the control spheroid, whereas those treated with 10 µM TAM had significant structural corruption and decreasing LIV and OCDS[Formula: see text] over treatment time. The spheroids treated with PTX had decreasing volumes and decrease of LIV and OCDS[Formula: see text] signals over time at most PTX concentrations. The spheroids treated with DOX had decreasing and increasing volumes over time at DOX concentrations of 1 and 10 µM, respectively. Meanwhile, the LIV and OCDS[Formula: see text] signals decreased over treatment time at all DOX concentrations. The D-OCT, particularly OCDS[Formula: see text], patterns were consistent with the fluorescence microscopic patterns. The diversity in the structural and D-OCT results among the drug types and among the concentrations are explained by the mechanisms of the drugs. The presented results suggest that D-OCT is useful for evaluating the difference in the tumor spheroid response to different drugs and it can be a useful tool for anti-cancer drug testing.

Polarization-artifact reduction and accuracy improvement of Jones-matrix polarization-sensitive optical coherence tomography by multi-focus-averaging based multiple scattering reduction.

Polarization-sensitive optical coherence tomography (PS-OCT) is a promising biomedical imaging tool for the differentiation of various tissue properties. However, the presence of multiple-scattering (MS) signals can degrade the quantitative polarization measurement accuracy. We demonstrate a method to reduce MS signals and increase the measurement accuracy of Jones matrix PS-OCT. This method suppresses MS signals by averaging multiple Jones matrix volumes measured using different focal positions. The MS signals are decorrelated among the volumes by focus position modulation and are thus reduced by averaging. However, the single scattering signals are kept consistent among the focus-modulated volumes by computational refocusing. We validated the proposed method using a scattering phantom and a postmortem medaka fish. The results showed reduced artifacts in birefringence and degree-of-polarization uniformity measurements, particularly in deeper regions in the samples. This method offers a practical solution to mitigate MS-induced artifacts in PS-OCT imaging and improves quantitative polarization measurement accuracy.

Polarization-Diversity Optical Coherence Tomography Assessment of Choroidal Nevi.

Purpose: The purpose of this study was to demonstrate the utility of polarization-diversity optical coherence tomography (PD-OCT), a noninvasive imaging technique with melanin-specific contrast, in the quantitative and qualitative assessment of choroidal nevi. Methods: Nevi were imaged with a custom-built 55-degree field-of-view (FOV) 400 kHz PD-OCT system. Imaging features on PD-OCT were compared to those on fundus photography, auto-fluorescence, ultrasound, and non-PD-OCT images. Lesions were manually segmented for size measurement and metrics for objective assessment of melanin distributions were calculated, including degree of polarization uniformity (DOPU), attenuation coefficient, and melanin occupancy rate (MOR). Results: We imaged 17 patients (mean age = 69.5 years, range = 37-90) with 11 pigmented, 3 non-pigmented, and 3 mixed pigmentation nevi. Nevi with full margin acquisition had an average longest basal diameter of 5.1 mm (range = 2.99-8.72 mm) and average height of 0.72 mm (range = 0.37 mm-2.09 mm). PD-OCT provided clear contrast of choroidal melanin content, distribution, and delineation of nevus margins for melanotic nevi. Pigmented nevi were found to have lower DOPU, higher attenuation coefficient, and higher MOR than non-pigmented lesions. Melanin content on PD-OCT was consistent with pigmentation on fundus in 15 of 17 nevi (88%). Conclusions: PD-OCT allows objective assessment of choroidal nevi melanin content and distribution. In addition, melanin-specific contrast by PD-OCT enables clear nevus margin delineation and may improve serial growth surveillance. Further investigation is needed to determine the clinical significance and prognostic value of melanin characterization by PD-OCT in the evaluation of choroidal nevi.

Multi-focus averaging for multiple scattering suppression in optical coherence tomography.

Multiple scattering is one of the main factors that limits the penetration depth of optical coherence tomography (OCT) in scattering samples. We propose a method termed multi-focus averaging (MFA) to suppress the multiple-scattering signals and improve the image contrast of OCT in deep regions. The MFA method captures multiple OCT volumes with various focal positions and averages them in complex form after correcting the varying defocus through computational refocusing. Because the multiple-scattering takes different trajectories among the different focal position configurations, this averaging suppresses the multiple-scattering signal. Meanwhile, the single-scattering takes a consistent trajectory regardless of the focal position configuration and is not suppressed. Hence, the MFA method improves the ratio between the single-scattering signal and multiple-scattering signal, resulting in an enhancement in the image contrast. A scattering phantom and a postmortem zebrafish were measured to validate the proposed method. The results showed that the contrast of intensity images of both the phantom and zebrafish were improved using the MFA method, such that they were better than the contrast provided by the standard single focus averaging method. The MFA method provides a cost-effective solution for contrast enhancement through multiple-scattering reduction in tissue imaging using OCT systems.

Extended and adjustable field-of-view of variable interscan time analysis by ammonite-scanning swept-source optical coherence tomography angiography.

A novel scanning protocol, ammonite scan, is proposed for widefield optical coherence tomography angiography (OCTA) and relative retinal blood flow velocity imaging in the human retina using variable interscan time analysis (VISTA). A repeated circle scan using a 400 kHz swept-source was employed to achieve an interscan time of 1.28 ms. The center of the repeated circular scan continuously moved spirally towards the peripheral region, ensuring an extended and adjustable scan range while preserving the short interscan time. Image artifacts due to eye movement were eliminated via extra motion-correction processing using data redundancy. The relative blood flow velocity in superficial and deep plexus layers was calculated from the VISTA image, and their ratio was used to explore the microvascular flow parameter in the healthy human eye.

Birefringence-derived artifact in optical coherence tomography imaging of the lamina cribrosa in eyes with glaucoma.

We investigated birefringence-derived artifacts that potentially mimic focal defects of the lamina cribrosa (focal LC defects) in optical coherence tomography (OCT) imaging of eyes with glaucoma. This study included 74 eyes of 48 patients with glaucoma. Five horizontal line B-scan images of the optic disc were obtained using commercial swept-source OCT. From a dataset of prototype swept-source polarization-diversity OCT, we calculated the following types of OCT images: polarization-dependent, polarization-dependent attenuation-coefficient, polarization-independent, and polarization-independent attenuation-coefficient. We assessed the commercial OCT images for the presence of birefringence-derived artifacts by comparison with the polarization-diversity OCT images. Commercial OCT showed suggestive findings of focal LC defects in 17 of 74 eyes. Reevaluation using polarization-independent OCT revealed that the focal LC defects in one of 17 eyes (5.9%) were actually birefringence-derived artifacts. This study demonstrated the existence of birefringence-derived artifacts mimicking focal LC defects in commercial OCT imaging and indicated that polarization-diversity OCT is an effective tool to evaluate the presence of these artifacts.

Renal tubular function and morphology revealed in kidney without labeling using three-dimensional dynamic optical coherence tomography.

Renal tubule has distinct metabolic features and functional activity that may be altered during kidney disease. In this paper, we present label-free functional activity imaging of renal tubule in normal and obstructed mouse kidney models using three-dimensional (3D) dynamic optical coherence tomography (OCT) ex vivo. To create an obstructed kidney model, we ligated the ureter of the left kidney for either 7 or 14 days. Two different dynamic OCT (DOCT) methods were implemented to access the slow and fast activity of the renal tubules: a logarithmic intensity variance (LIV) method and a complex-correlation-based method. Three-dimensional DOCT data were acquired with a 1.3 [Formula: see text]m swept-source OCT system and repeating raster scan protocols. In the normal kidney, the renal tubule appeared as a convoluted pipe-like structure in the DOCT projection image. Such pipe-like structures were not observed in the kidneys subjected to obstruction of the ureter for several days. Instead of any anatomical structures, a superficial high dynamics appearance was observed in the perirenal cortex region of the obstructed kidneys. These findings suggest that volumetric LIV can be used as a tool to investigate kidney function during kidney diseases.

Label-free drug response evaluation of human derived tumor spheroids using three-dimensional dynamic optical coherence tomography.

This study aims at demonstrating label-free drug-response-patterns assessment of different tumor spheroids and drug types by dynamic optical coherence tomography (D-OCT). The study involved human breast cancer (MCF-7) and colon cancer (HT-29) spheroids. The MCF-7 and HT-29 spheroids were treated with paclitaxel (Taxol; PTX) and the active metabolite of irinotecan SN-38, respectively. The drugs were applied with 0 (control), 0.1, 1, and 10 µM concentrations and the treatment durations were 1, 3, and 6 days. A swept-source OCT microscope equipped with a repeated raster scanning protocol was used to scan the spheroids. Logarithmic intensity variance (LIV) and late OCT correlation decay speed (OCDS[Formula: see text]) algorithms were used to visualize the tumor spheroid dynamics. LIV and OCDS[Formula: see text] images visualized different response patterns of the two types of spheroids. In addition, spheroid morphology, LIV, and OCDS[Formula: see text] quantification showed different time-courses among the spheroid and drug types. These results may indicate different action mechanisms of the drugs. The results showed the feasibility of D-OCT for the evaluation of drug response patterns of different cell spheroids and drug types and suggest that D-OCT can perform anti-cancer drug testing.

Theoretical model for en face optical coherence tomography imaging and its application to volumetric differential contrast imaging.

A new formulation of the lateral imaging process of point-scanning optical coherence tomography (OCT) and a new differential contrast method designed by using this formulation are presented. The formulation is based on a mathematical sample model called the dispersed scatterer model (DSM), in which the sample is represented as a material with a spatially slowly varying refractive index and randomly distributed scatterers embedded in the material. It is shown that the formulation represents a meaningful OCT image and speckle as two independent mathematical quantities. The new differential contrast method is based on complex signal processing of OCT images, and the physical and numerical imaging processes of this method are jointly formulated using the same theoretical strategy as in the case of OCT. The formula shows that the method provides a spatially differential image of the sample structure. This differential imaging method is validated by measuring in vivo and in vitro samples.

Label-free intratissue activity imaging of alveolar organoids with dynamic optical coherence tomography.

An organoid is a three-dimensional (3D) in vitro cell culture emulating human organs. We applied 3D dynamic optical coherence tomography (DOCT) to visualize the intratissue and intracellular activities of human induced pluripotent stem cells (hiPSCs)-derived alveolar organoids in normal and fibrosis models. 3D DOCT data were acquired with an 840-nm spectral domain optical coherence tomography with axial and lateral resolutions of 3.8 µm (in tissue) and 4.9 µm, respectively. The DOCT images were obtained by the logarithmic-intensity-variance (LIV) algorithm, which is sensitive to the signal fluctuation magnitude. The LIV images revealed cystic structures surrounded by high-LIV borders and mesh-like structures with low LIV. The former may be alveoli with a highly dynamics epithelium, while the latter may be fibroblasts. The LIV images also demonstrated the abnormal repair of the alveolar epithelium.

Synthesizing the degree of polarization uniformity from non-polarization-sensitive optical coherence tomography signals using a neural network.

Degree of polarization uniformity (DOPU) imaging obtained by polarization-sensitive optical coherence tomography (PS-OCT) has the potential to provide biomarkers for retinal diseases. It highlights abnormalities in the retinal pigment epithelium that are not always clear in the OCT intensity images. However, a PS-OCT system is more complicated than conventional OCT. We present a neural-network-based approach to estimate the DOPU from standard OCT images. DOPU images were used to train a neural network to synthesize the DOPU from single-polarization-component OCT intensity images. DOPU images were then synthesized by the neural network, and the clinical findings from ground truth DOPU and synthesized DOPU were compared. There is a good agreement in the findings for RPE abnormalities: recall was 0.869 and precision was 0.920 for 20 cases with retinal diseases. In five cases of healthy volunteers, no abnormalities were found in either the synthesized or ground truth DOPU images. The proposed neural-network-based DOPU synthesis method demonstrates the potential of extending the features of retinal non-PS OCT.

Birefringence-derived scleral artifacts in optical coherence tomography images of eyes with pathologic myopia.

We investigated birefringence-derived scleral artifacts in optical coherence tomography (OCT) images of eyes with pathologic myopia. This study included 76 eyes of 42 patients with pathologic myopia. Five sets of OCT B-scan images of the macula were obtained using commercial swept-source OCT. A dataset of prototype swept-source polarization-diversity OCT images was used to identify polarization-dependent OCT images (i.e., complex averaging of OCT signals from two polarization channels) and polarization-independent OCT images (i.e., intensity averaging of two OCT signals). Polarization-dependent OCT images and commercial OCT images were assessed for the presence of birefringence-derived artifacts by comparison with polarization-independent OCT images. Both polarization-dependent OCT images and commercial OCT images contained scleral vessel artifacts. Scleral vessel artifacts were present in 46 of 76 eyes (60.5%) imaged by polarization-dependent OCT and 17 of 76 eyes (22.4%) imaged by commercial OCT. The proportion of images that showed scleral vessel artifacts was significantly greater among polarization-dependent OCT images than among commercial OCT images (P < 0.001). Additionally, polarization-dependent OCT images showed low-intensity band artifacts. This study demonstrated the existence of birefringence-derived scleral artifacts in commercial OCT images and indicated that polarization-diversity OCT is an effective tool to evaluate the presence of these artifacts.

Extending field-of-view of retinal imaging by optical coherence tomography using convolutional Lissajous and slow scan patterns.

Optical coherence tomography (OCT) is a high-speed non-invasive cross-sectional imaging technique. Although its imaging speed is high, three-dimensional high-spatial-sampling-density imaging of in vivo tissues with a wide field-of-view (FOV) is challenging. We employed convolved Lissajous and slow circular scanning patterns to extend the FOV of retinal OCT imaging with a 1-µm, 100-kHz-sweep-rate swept-source OCT prototype system. Displacements of sampling points due to eye movements are corrected by post-processing based on a Lissajous scan. Wide FOV three-dimensional retinal imaging with high sampling density and motion correction is achieved. Three-dimensional structures obtained using repeated imaging sessions of a healthy volunteer and two patients showed good agreement. The demonstrated technique will extend the FOV of simple point-scanning OCT, such as commercial ophthalmic OCT devices, without sacrificing sampling density.

Longitudinal investigation of a xenograft tumor zebrafish model using polarization-sensitive optical coherence tomography.

Breast cancer is a leading cause of death in female patients worldwide. Further research is needed to get a deeper insight into the mechanisms involved in the development of this devastating disease and to find new therapy strategies. The zebrafish is an established animal model, especially in the field of oncology, which has shown to be a promising candidate for pre-clinical research and precision-based medicine. To investigate cancer growth in vivo in zebrafish, one approach is to explore xenograft tumor models. In this article, we present the investigation of a juvenile xenograft zebrafish model using a Jones matrix optical coherence tomography (JM-OCT) prototype. Immunosuppressed wild-type fish at 1-month post-fertilization were injected with human breast cancer cells and control animals with phosphate buffered saline in the tail musculature. In a longitudinal study, the scatter, polarization, and vasculature changes over time were investigated and quantified in control versus tumor injected animals. A significant decrease in birefringence and an increase in scattering signal was detected in tumor injected zebrafish in comparison to the control once. This work shows the potential of JM-OCT as a non-invasive, label-free, three-dimensional, high-resolution, and tissue-specific imaging tool in pre-clinical cancer research based on juvenile zebrafish models.

Label-free metabolic imaging of non-alcoholic-fatty-liver-disease (NAFLD) liver by volumetric dynamic optical coherence tomography.

Label-free metabolic imaging of non-alcoholic fatty liver disease (NAFLD) mouse liver is demonstrated ex vivo by dynamic optical coherence tomography (OCT). The NAFLD mouse is a methionine choline-deficient (MCD)-diet model, and two mice fed the MCD diet for 1 and 2 weeks are involved in addition to a normal-diet mouse. The dynamic OCT is based on repeating raster scan and logarithmic intensity variance (LIV) analysis that enables volumetric metabolic imaging with a standard-speed (50,000 A-lines/s) OCT system. Metabolic domains associated with lipid droplet accumulation and inflammation are clearly visualized three-dimensionally. Particularly, the normal-diet liver exhibits highly metabolic vessel-like structures of peri-vascular hepatic zones. The 1-week MCD-diet liver shows ring-shaped highly metabolic structures formed with lipid droplets. The 2-week MCD-diet liver exhibits fragmented vessel-like structures associated with inflammation. These results imply that volumetric LIV imaging is useful for visualizing and assessing NAFLD abnormalities.