Robust, high-density lesion mapping in the left atrium with near-infrared spectroscopy.

Significance: Radiofrequency ablation (RFA) procedures for atrial fibrillation frequently fail to prevent recurrence, partially due to limitations in assessing extent of ablation. Optical spectroscopy shows promise in assessing RFA lesion formation but has not been validated in conditions resembling those in vivo. Aim: Catheter-based near-infrared spectroscopy (NIRS) was applied to porcine hearts to demonstrate that spectrally derived optical indices remain accurate in blood and at oblique incidence angles. Approach: Porcine left atria were ablated and mapped using a custom-fabricated NIRS catheter. Each atrium was mapped first in phosphate-buffered saline (PBS) then in porcine blood. Results: NIRS measurements showed little angle dependence up to 60 deg. A trained random forest model predicted lesions with a sensitivity of 81.7%, a specificity of 86.1%, and a receiver operating characteristic curve area of 0.921. Predicted lesion maps achieved a mean structural similarity index of 0.749 and a mean normalized inner product of 0.867 when comparing maps obtained in PBS and blood. Conclusions: Catheter-based NIRS can precisely detect RFA lesions on left atria submerged in blood. Optical parameters are reliable in blood and without perpendicular contact, confirming their ability to provide useful feedback during in vivo RFA procedures.

Image harmonization and deep learning automated classification of plus disease in retinopathy of prematurity.

Purpose: Retinopathy of prematurity (ROP) is a retinal vascular disease affecting premature infants that can culminate in blindness within days if not monitored and treated. A disease stage for scrutiny and administration of treatment within ROP is "plus disease" characterized by increased tortuosity and dilation of posterior retinal blood vessels. The monitoring of ROP occurs via routine imaging, typically using expensive instruments ($50 to $140 K) that are unavailable in low-resource settings at the point of care. Approach: As part of the smartphone-ROP program to enable referrals to expert physicians, fundus images are acquired using smartphone cameras and inexpensive lenses. We developed methods for artificial intelligence determination of plus disease, consisting of a preprocessing pipeline to enhance vessels and harmonize images followed by deep learning classification. A deep learning binary classifier (plus disease versus no plus disease) was developed using GoogLeNet. Results: Vessel contrast was enhanced by 90% after preprocessing as assessed by the contrast improvement index. In an image quality evaluation, preprocessed and original images were evaluated by pediatric ophthalmologists from the US and South America with years of experience diagnosing ROP and plus disease. All participating ophthalmologists agreed or strongly agreed that vessel visibility was improved with preprocessing. Using images from various smartphones, harmonized via preprocessing (e.g., vessel enhancement and size normalization) and augmented in physically reasonable ways (e.g., image rotation), we achieved an area under the ROC curve of 0.9754 for plus disease on a limited dataset. Conclusions: Promising results indicate the potential for developing algorithms and software to facilitate the usage of cell phone images for staging of plus disease.

Experiences of medical device innovators as they navigate the regulatory system in Uganda.

Objective: A medical device must undergo rigorous regulatory processes to verify its safety and effectiveness while in use. In low-and middle-income countries like Uganda however, medical device innovators and designers face challenges around bringing a device from ideation to being market-ready. This is mainly attributed to a lack of clear regulatory procedures among other factors. In this paper, we illustrate the current landscape of investigational medical devices regulation in Uganda. Methods: Information about the different bodies involved in regulation of medical devices in Uganda was obtained online. Nine medical device teams whose devices have gone through the Ugandan regulatory system were interviewed to gain insights into their experiences with the regulatory system. Interviews focused on the challenges they faced, how they navigated them, and factors that supported their progress towards putting their devices on the market. Results: We identified different bodies that are part of the stepwise regulatory pathway of investigational medical devices in Uganda and roles played by each in the regulatory process. Experiences of the medical device teams collected showed that navigation through the regulatory system was different for each team and progress towards market readiness was fuelled by funding, simplicity of device, and mentorship. Conclusion: Medical devices regulation exists in Uganda but is characterised by a landscape that is still in development which thereby affects the progress of investigational medical devices.

Segmentation of beating embryonic heart structures from 4-D OCT images using deep learning.

Optical coherence tomography (OCT) has been used to investigate heart development because of its capability to image both structure and function of beating embryonic hearts. Cardiac structure segmentation is a prerequisite for the quantification of embryonic heart motion and function using OCT. Since manual segmentation is time-consuming and labor-intensive, an automatic method is needed to facilitate high-throughput studies. The purpose of this study is to develop an image-processing pipeline to facilitate the segmentation of beating embryonic heart structures from a 4-D OCT dataset. Sequential OCT images were obtained at multiple planes of a beating quail embryonic heart and reassembled to a 4-D dataset using image-based retrospective gating. Multiple image volumes at different time points were selected as key-volumes, and their cardiac structures including myocardium, cardiac jelly, and lumen, were manually labeled. Registration-based data augmentation was used to synthesize additional labeled image volumes by learning transformations between key-volumes and other unlabeled volumes. The synthesized labeled images were then used to train a fully convolutional network (U-Net) for heart structure segmentation. The proposed deep learning-based pipeline achieved high segmentation accuracy with only two labeled image volumes and reduced the time cost of segmenting one 4-D OCT dataset from a week to two hours. Using this method, one could carry out cohort studies that quantify complex cardiac motion and function in developing hearts.

Automated analysis framework for in vivo cardiac ablation therapy monitoring with optical coherence tomography.

Radiofrequency ablation (RFA) is a minimally invasive procedure that is commonly used for the treatment of atrial fibrillation. However, it is associated with a significant risk of arrhythmia recurrence and complications owing to the lack of direct visualization of cardiac substrates and real-time feedback on ablation lesion transmurality. Within this manuscript, we present an automated deep learning framework for in vivo intracardiac optical coherence tomography (OCT) analysis of swine left atria. Our model can accurately identify cardiac substrates, monitor catheter-tissue contact stability, and assess lesion transmurality on both OCT intensity and polarization-sensitive OCT data. To the best of our knowledge, we have developed the first automatic framework for in vivo cardiac OCT analysis, which holds promise for real-time monitoring and guidance of cardiac RFA therapy..

Phase-Decorrelation Optical Coherence Tomography Measurement of Cold-Induced Nuclear Cataract.

Purpose: The purpose of this work is to determine the sensitivity of phase-decorrelation optical coherence tomography (OCT) to protein aggregation associated with cataracts in the ocular lens, as compared to OCT signal intensity. Methods: Six fresh porcine globes were held at 4°C until cold cataracts developed. As the globes were re-warmed to ambient temperature, reversing the cold cataract, each lens was imaged repeatedly using a conventional OCT system. Throughout each experiment, the internal temperature of the globe was recorded using a needle-mounted thermocouple. OCT scans were acquired, their temporal fluctuations were analyzed, and the rates of decorrelation were spatially mapped. Both decorrelation and intensity were evaluated as a function of recorded temperature. Results: Both signal decorrelation and intensity were found to change with lens temperature, a surrogate of protein aggregation. However, the relationship between signal intensity and temperature was not consistent across different samples. In contrast, the relationship between decorrelation and temperature was found to be consistent across samples. Conclusions: In this study, signal decorrelation was shown to be a more repeatable metric for quantification of crystallin protein aggregation in the ocular lens than OCT intensity-based metrics. Thus, OCT signal decorrelation measurements could enable more detailed and sensitive study of methods to prevent cataract formation. Translational Relevance: This dynamic light scattering-based approach to early cataract assessment can be implemented on existing clinical OCT systems without hardware additions, so it could quickly become part of a clinical study workflow or an indication for use for a pharmaceutical cataract intervention.

Automated endocardial cushion segmentation and cellularization quantification in developing hearts using optical coherence tomography.

Of all congenital heart defects (CHDs), anomalies in heart valves and septa are among the most common and contribute about fifty percent to the total burden of CHDs. Progenitors to heart valves and septa are endocardial cushions formed in looping hearts through a multi-step process that includes localized expansion of cardiac jelly, endothelial-to-mesenchymal transition, cell migration and proliferation. To characterize the development of endocardial cushions, previous studies manually measured cushion size or cushion cell density from images obtained using histology, immunohistochemistry, or optical coherence tomography (OCT). Manual methods are time-consuming and labor-intensive, impeding their applications in cohort studies that require large sample sizes. This study presents an automated strategy to rapidly characterize the anatomy of endocardial cushions from OCT images. A two-step deep learning technique was used to detect the location of the heart and segment endocardial cushions. The acellular and cellular cushion regions were then segregated by K-means clustering. The proposed method can quantify cushion development by measuring the cushion volume and cellularized fraction, and also map 3D spatial organization of the acellular and cellular cushion regions. The application of this method to study the developing looping hearts allowed us to discover a spatial asymmetry of the acellular cardiac jelly in endocardial cushions during these critical stages, which has not been reported before.

Polarization-sensitive optical coherence tomography monitoring of percutaneous radiofrequency ablation in left atrium of living swine.

Radiofrequency ablation (RFA) is commonly used to treat atrial fibrillation (AF). However, the outcome is often compromised due to the lack of direct real-time feedback to assess lesion transmurality. In this work, we evaluated the ability of polarization-sensitive optical coherence tomography (PSOCT) to measure cardiac wall thickness and assess RF lesion transmurality during left atrium (LA) RFA procedures. Quantitative transmural lesion criteria using PSOCT images were determined ex vivo using an integrated PSOCT-RFA catheter and fresh swine hearts. LA wall thickness of living swine was measured with PSOCT and validated with a micrometer after harvesting the heart. A total of 38 point lesions were created in the LA of 5 living swine with the integrated PSOCT-RFA catheter using standard clinical RFA procedures. For all lesions with analyzable PSOCT images, lesion transmurality was assessed with a sensitivity of 89% (17 of 19 tested positive) and a specificity of 100% (5 of 5 tested negative) using the quantitative transmural criteria. This is the first report of using PSOCT to assess LA RFA lesion transmurality in vivo. The results indicate that PSOCT may potentially provide direct real-time feedback for LA wall thickness and lesion transmurality.

CompassLSM: axially swept light-sheet microscopy made simple.

Axially swept light-sheet microscopy (ASLM) is an effective method of generating a uniform light sheet across a large field of view (FOV). However, current ASLM designs are more complicated than conventional light-sheet systems, limiting their adaptation in less experienced labs. By eliminating difficult-to-align components and reducing the total number of components, we show that high-performance ASLM can be accomplished much simpler than existing designs, requiring less expertise and effort to construct, align, and operate. Despite the high simplicity, our design achieved 3.5-µm uniform optical sectioning across a >6-mm FOV, surpassing existing light-sheet designs with similar optical sectioning. With well-corrected chromatic aberration, multi-channel fluorescence imaging can be performed without realignment. This manuscript provides a comprehensive tutorial on building the system and demonstrates the imaging performance with optically cleared whole-mount tissue samples.

Biomechanics of Ophthalmic Crosslinking.

Crosslinking involves the formation of bonds between polymer chains, such as proteins. In biological tissues, these bonds tend to stiffen the tissue, making it more resistant to mechanical degradation and deformation. In ophthalmology, the crosslinking phenomenon is being increasingly harnessed and explored as a treatment strategy for treating corneal ectasias, keratitis, degenerative myopia, and glaucoma. This review surveys the multitude of exogenous crosslinking strategies reported in the literature, both "light" (involving light energy) and "dark" (involving non-photic chemical processes), and explores their mechanisms, cytotoxicity, and stage of translational development. The spectrum of ophthalmic applications described in the literature is then discussed, with particular attention to proposed therapeutic mechanisms in the cornea and sclera. The mechanical effects of crosslinking are then discussed in the context of their proposed site and scale of action. Biomechanical characterization of the crosslinking effect is needed to more thoroughly address knowledge gaps in this area, and a review of reported methods for biomechanical characterization is presented with an attempt to assess the sensitivity of each method to crosslinking-mediated changes using data from the experimental and clinical literature. Biomechanical measurement methods differ in spatial resolution, mechanical sensitivity, suitability for detecting crosslinking subtypes, and translational readiness and are central to the effort to understand the mechanistic link between crosslinking methods and clinical outcomes of candidate therapies. Data on differences in the biomechanical effect of different crosslinking protocols and their correspondence to clinical outcomes are reviewed, and strategies for leveraging measurement advances predicting clinical outcomes of crosslinking procedures are discussed. Advancing the understanding of ophthalmic crosslinking, its biomechanical underpinnings, and its applications supports the development of next-generation crosslinking procedures that optimize therapeutic effect while reducing complications.

Prenatal ethanol exposure impairs the conduction delay at the atrioventricular junction in the looping heart.

The etiology of ethanol-related congenital heart defects has been the focus of much study, but most research has concentrated on cellular and molecular mechanisms. We have shown with optical coherence tomography (OCT) that ethanol exposure led to increased retrograde flow and smaller atrioventricular (AV) cushions compared with controls. Since AV cushions play a role in patterning the conduction delay at the atrioventricular junction (AVJ), this study aims to investigate whether ethanol exposure alters the AVJ conduction in early looping hearts and whether this alteration is related to the decreased cushion size. Quail embryos were exposed to a single dose of ethanol at gastrulation, and Hamburger-Hamilton stage 19-20 hearts were dissected for imaging. Cardiac conduction was measured using an optical mapping microscope and we imaged the endocardial cushions using OCT. Our results showed that, compared with controls, ethanol-exposed embryos exhibited abnormally fast AVJ conduction and reduced cushion size. However, this increased conduction velocity (CV) did not strictly correlate with decreased cushion volume and thickness. By matching the CV map to the cushion-size map along the inflow heart tube, we found that the slowest conduction location was consistently at the atrial side of the AVJ, which had the thinner cushions, not at the thickest cushion location at the ventricular side as expected. Our findings reveal regional differences in the AVJ myocardium even at this early stage in heart development. These findings reveal the early steps leading to the heterogeneity and complexity of conduction at the mature AVJ, a site where arrhythmias can be initiated.NEW & NOTEWORTHY To the best of our knowledge, this is the first study investigating the impact of ethanol exposure on the early cardiac conduction system. Our results showed that ethanol-exposed embryos exhibited abnormally fast atrioventricular conduction. In addition, our findings, in CV measurements and endocardial cushion thickness, reveal regional differences in the AVJ myocardium even at this early stage in heart development, suggesting that the differentiation and maturation at this site are complex and warrant further studies.

Pocket MUSE: an affordable, versatile and high-performance fluorescence microscope using a smartphone.

Smartphone microscopes can be useful tools for a broad range of imaging applications. This manuscript demonstrates the first practical implementation of Microscopy with Ultraviolet Surface Excitation (MUSE) in a compact smartphone microscope called Pocket MUSE, resulting in a remarkably effective design. Fabricated with parts from consumer electronics that are readily available at low cost, the small optical module attaches directly over the rear lens in a smartphone. It enables high-quality multichannel fluorescence microscopy with submicron resolution over a 10× equivalent field of view. In addition to the novel optical configuration, Pocket MUSE is compatible with a series of simple, portable, and user-friendly sample preparation strategies that can be directly implemented for various microscopy applications for point-of-care diagnostics, at-home health monitoring, plant biology, STEM education, environmental studies, etc.

Glutathione Protects the Developing Heart from Defects and Global DNA Hypomethylation Induced by Prenatal Alcohol Exposure.

BACKGROUND: Fetal alcohol spectrum disorder (FASD) is caused by prenatal alcohol exposure (PAE), the intake of ethanol (C2 H5 OH) during pregnancy. Features of FASD cover a range of structural and functional defects including congenital heart defects (CHDs). Folic acid and choline, contributors of methyl groups to one-carbon metabolism (OCM), prevent CHDs in humans. Using our avian model of FASD, we have previously reported that betaine, another methyl donor downstream of choline, prevents CHDs. The CHD preventions are substantial but incomplete. Ethanol causes oxidative stress as well as depleting methyl groups for OCM to support DNA methylation and other epigenetic alterations. To identify more compounds that can safely and effectively prevent CHDs and other effects of PAE, we tested glutathione (GSH), a compound that regulates OCM and is known as a "master antioxidant." METHODS/RESULTS: Quail embryos injected with a single dose of ethanol at gastrulation exhibited congenital defects including CHDs similar to those identified in FASD individuals. GSH injected simultaneously with ethanol not only prevented CHDs, but also improved survival and prevented other PAE-induced defects. Assays of hearts at 8 days (HH stage 34) of quail development, when the heart normally has developed 4-chambers, showed that this single dose of PAE reduced global DNA methylation. GSH supplementation concurrent with PAE normalized global DNA methylation levels. The same assays performed on quail hearts at 3 days (HH stage 19-20) of development, showed no difference in global DNA methylation between controls, ethanol-treated, GSH alone, and GSH plus ethanol-treated cohorts. CONCLUSIONS: GSH supplementation shows promise to inhibit effects of PAE by improving survival, reducing the incidence of morphological defects including CHDs, and preventing global hypomethylation of DNA in heart tissues.

Genetically Encoded Calcium Indicators for In Situ Functional Studies of Corneal Nerves.

Purpose: Millions of people suffer from diseases that involve corneal nerve dysfunction, caused by various conditions, including dry eye syndrome, neurotrophic keratopathy, diabetes, herpes simplex, glaucoma, and Alzheimer's disease. The morphology of corneal nerves has been studied extensively. However, corneal nerve function has only been studied in a limited fashion owing to a lack of tools. Here, we present a new system for studying corneal nerve function. Methods: Optical imaging was performed on the cornea of excised murine globes taken from a model animal expressing a genetically encoded calcium indicator, GCaMP6f, to record calcium transients. A custom perfusion and imaging chamber for ex vivo murine globes was designed to maintain and stabilize the cornea, while allowing the introduction of chemical stimulation during imaging. Results: Imaging of calcium signals in the ex vivo murine cornea was demonstrated. Strong calcium signals with minimal photobleaching were observed in experiments lasting up to 10 minutes. Concentrated potassium and lidocaine solutions both modulated corneal nerve activity. Similar responses were observed in the same neurons across multiple chemical stimulations, suggesting the feasibility of using chemical stimulations to test the response of the corneal nerves. Conclusions: Our studies suggest that this tool will be of great use for studying functional changes to corneal nerves in response to disease and ocular procedures. This process will enable preclinical testing of new ocular procedures to minimize damage to corneal innervation and therapies for diminished neural function.

Semi-automated shear stress measurements in developing embryonic hearts.

Blood-induced shear stress influences gene expression. Abnormal shear stress patterns on the endocardium of the early-stage heart tube can lead to congenital heart defects. To have a better understanding of these mechanisms, it is essential to include shear stress measurements in longitudinal cohort studies of cardiac development. Previously reported approaches are computationally expensive and nonpractical when assessing many animals. Here, we introduce a new approach to estimate shear stress that does not rely on recording 4D image sets and extensive post processing. Our method uses two adjacent optical coherence tomography frames (B-scans) where lumen geometry and flow direction are determined from the structural data and the velocity is measured from the Doppler OCT signal. We validated our shear stress estimate by flow phantom experiments and applied it to live quail embryo hearts where observed shear stress patterns were similar to previous studies.

Intracardiac radiofrequency ablation in living swine guided by polarization-sensitive optical coherence tomography.

SIGNIFICANCE: Pulmonary vein isolation with catheter-based radiofrequency ablation (RFA) is carried out frequently to treat atrial fibrillation. However, RFA lesion creation is only guided by indirect information (e.g., temperature, impedance, and contact force), which may result in poor lesion quality (e.g., nontransmural) and can lead to reoccurrence or complications. AIM: The feasibility of guiding intracardiac RFA with an integrated polarization-sensitive optical coherence tomography (PSOCT)-RFA catheter in the right atria (RA) of living swine is demonstrated. APPROACH: In total, 12 sparse lesions were created in the RA of three living swine using an integrated PSOCT-RFA catheter with standard ablation protocol. PSOCT images were displayed in real time to guide catheter-tissue apposition. After experiments, post-processed PSOCT images were analyzed to assess lesion quality and were compared with triphenyltetrazolium chloride (TTC) lesion quality analysis. RESULTS: Five successful lesions identified with PSOCT images were all confirmed by TTC analysis. In two ablations, PSOCT imaging detected gas bubble formation, indicating overtreatment. Unsuccessful lesions observed with PSOCT imaging were confirmed by TTC analysis. CONCLUSIONS: The results demonstrate that the PSOCT-RFA catheter provides real-time feedback to guide catheter-tissue apposition, monitor lesion quality, and possibly help avoid complications due to overtreatment, which may enable more effective and safer RFA treatment.

Automated stent coverage analysis in intravascular OCT (IVOCT) image volumes using a support vector machine and mesh growing.

Absence of vascular-stent tissue coverage by IVOCT is a biomarker for potential stent-related thrombosis. We developed highly-automated algorithms to classify covered and uncovered struts and quantitatively evaluate stent apposition. We trained a machine learning model on 7,125 images, and included an active learning, relabeling step to improve noisy labels. We obtained uncovered strut classification sensitivity/specificity (94%/90%) comparable to analyst inter-and-intra-observer variability and AUC (0.97), and tissue coverage thickness measurement arguably better than the commercial product. By comparing classification models from regular and relabeled data sets, we observed robustness of the support vector machine to noisy data. A graph-based algorithm detected clusters of uncovered struts thought to pose a greater risk than isolated uncovered struts. The software enables highly-automated, objective, repeatable, comprehensive stent analysis.

A Review of Structural and Biomechanical Changes in the Cornea in Aging, Disease, and Photochemical Crosslinking.

The study of corneal biomechanics is motivated by the tight relationship between biomechanical properties and visual function within the ocular system. For instance, variation in collagen fibril alignment and non-enzymatic crosslinks rank high among structural factors which give rise to the cornea's particular shape and ability to properly focus light. Gradation in these and other factors engender biomechanical changes which can be quantified by a wide variety of techniques. This review summarizes what is known about both the changes in corneal structure and associated changes in corneal biomechanical properties in aging, keratoconic, and photochemically crosslinked corneas. In addition, methods for measuring corneal biomechanics are discussed and the topics are related to both clinical studies and biomechanical modeling simulations.

SLIME: robust, high-speed 3D microvascular mapping.

Three dimensional (3D) microvascular imaging of cubic millimeter to centimeter size volumes often requires much time and expensive instruments. By combining optical clearing with a novel scatter-based optical coherence tomography (OCT) contrast agent, we have greatly extended OCT imaging depth in excised tissues while maintaining a simple and low cost approach that does not require in-depth OCT knowledge. The new method enables fast 3D microvascular mapping in large tissue volumes, providing a promising tool for investigating organ level microvascular abnormalities in large cohorts.