Quantification of smooth muscle in human airways by polarization-sensitive optical coherence tomography requires correction for perichondrium.

Quantifying airway smooth muscle (ASM) in patients with asthma raises the possibility of improved and personalized disease management. Endobronchial polarization-sensitive optical coherence tomography (PS-OCT) is a promising quantitative imaging approach that is in the early stages of clinical translation. To date, only animal tissues have been used to assess the accuracy of PS-OCT to quantify absolute (rather than relative) ASM in cross sections with directly matched histological cross sections as validation. We report the use of whole fresh human and pig airways to perform a detailed side-by-side qualitative and quantitative validation of PS-OCT against gold-standard histology. We matched and quantified 120 sections from five human and seven pig (small and large) airways and linked PS-OCT signatures of ASM to the tissue structural appearance in histology. Notably, we found that human cartilage perichondrium can share with ASM the properties of birefringence and circumferential alignment of fibers, making it a significant confounder for ASM detection. Measurements not corrected for perichondrium overestimated ASM content several-fold (P < 0.001, paired t test). After careful exclusion of perichondrium, we found a strong positive correlation (r = 0.96, P < 0.00001) of ASM area measured by PS-OCT and histology, supporting the method's application in human subjects. Matching human histology further indicated that PS-OCT allows conclusions on the intralayer composition and in turn potential contractile capacity of ASM bands. Together these results form a reliable basis for future clinical studies.NEW & NOTEWORTHY Polarization-sensitive optical coherence tomography (PS-OCT) may facilitate in vivo measurement of airway smooth muscle (ASM). We present a quantitative validation correlating absolute ASM area from PS-OCT to directly matched histological cross sections using human tissue. A major confounder for ASM quantification was observed and resolved: fibrous perichondrium surrounding hyaline cartilage in human airways presents a PS-OCT signature similar to ASM for birefringence and optic axis orientation. Findings impact the development of automated methods for ASM segmentation.

The distribution of breast density in women aged 18 years and older.

PURPOSE: Age and body mass index (BMI) are critical considerations when assessing individual breast cancer risk, particularly for women with dense breasts. However, age- and BMI-standardized estimates of breast density are not available for screen-aged women, and little is known about the distribution of breast density in women aged < 40. This cross-sectional study uses three different modalities: optical breast spectroscopy (OBS), dual-energy X-ray absorptiometry (DXA), and mammography, to describe the distributions of breast density across categories of age and BMI. METHODS: Breast density measures were estimated for 1,961 Australian women aged 18-97 years using OBS (%water and %water + %collagen). Of these, 935 women had DXA measures (percent and absolute fibroglandular dense volume, %FGV and FGV, respectively) and 354 had conventional mammographic measures (percent and absolute dense area). The distributions for each breast density measure were described across categories of age and BMI. RESULTS: The mean age was 38 years (standard deviation = 15). Median breast density measures decreased with age and BMI for all three modalities, except for DXA-FGV, which increased with BMI and decreased after age 30. The variation in breast density measures was largest for younger women and decreased with increasing age and BMI. CONCLUSION: This unique study describes the distribution of breast density measures for women aged 18-97 using alternative and conventional modalities of measurement. While this study is the largest of its kind, larger sample sizes are needed to provide clinically useful age-standardized measures to identify women with high breast density for their age or BMI.

Alternative methods to measure breast density in younger women.

BACKGROUND: Breast density is a strong and potentially modifiable breast cancer risk factor. Almost everything we know about breast density has been derived from mammography, and therefore, very little is known about breast density in younger women aged <40. This study examines the acceptability and performance of two alternative breast density measures, Optical Breast Spectroscopy (OBS) and Dual X-ray Absorptiometry (DXA), in women aged 18-40. METHODS: Breast tissue composition (percent water, collagen, and lipid content) was measured in 539 women aged 18-40 using OBS. For a subset of 169 women, breast density was also measured via DXA (percent fibroglandular dense volume (%FGV), absolute dense volume (FGV), and non-dense volume (NFGV)). Acceptability of the measurement procedures was assessed using an adapted validated questionnaire. Performance was assessed by examining the correlation and agreement between the measures and their associations with known determinants of mammographic breast density. RESULTS: Over 93% of participants deemed OBS and DXA to be acceptable. The correlation between OBS-%water + collagen and %FGV was 0.48. Age and BMI were inversely associated with OBS-%water + collagen and %FGV and positively associated with OBS-%lipid and NFGV. CONCLUSIONS: OBS and DXA provide acceptable and viable alternative methods to measure breast density in younger women aged 18-40 years.

In vivo imaging of the depth-resolved optic axis of birefringence in human skin.

Recent progress has enabled the reconstruction of the local (i.e., depth-resolved) optic axis (OAx) of biological tissue from measurements made with polarization-sensitive optical coherence tomography (PS-OCT). Here we demonstrate local OAx imaging in healthy human skin in vivo. The images reveal dense, weaving patterns that are imperceptible in OCT intensity tomograms or conventional PS-OCT metrics and that suggest a mesh-like tissue organization, consistent with the morphology of dermal collagen. Using co-registered polarization-sensitive optical coherence microscopy, we furthermore investigated the impact of spatial resolution on the recovered OAx patterns and confirmed their consistency. OAx orientation as a contrast mechanism merits further exploration for applications in dermatology.

Stem cell migration and mechanotransduction on linear stiffness gradient hydrogels.

The spatial presentation of mechanical information is a key parameter for cell behavior. We have developed a method of polymerization control in which the differential diffusion distance of unreacted cross-linker and monomer into a prepolymerized hydrogel sink results in a tunable stiffness gradient at the cell-matrix interface. This simple, low-cost, robust method was used to produce polyacrylamide hydrogels with stiffness gradients of 0.5, 1.7, 2.9, 4.5, 6.8, and 8.2 kPa/mm, spanning the in vivo physiological and pathological mechanical landscape. Importantly, three of these gradients were found to be nondurotactic for human adipose-derived stem cells (hASCs), allowing the presentation of a continuous range of stiffnesses in a single well without the confounding effect of differential cell migration. Using these nondurotactic gradient gels, stiffness-dependent hASC morphology, migration, and differentiation were studied. Finally, the mechanosensitive proteins YAP, Lamin A/C, Lamin B, MRTF-A, and MRTF-B were analyzed on these gradients, providing higher-resolution data on stiffness-dependent expression and localization.

Ultrahigh-Resolution Optical Coherence Elastography Images Cellular-Scale Stiffness of Mouse Aorta.

Cellular-scale imaging of the mechanical properties of tissue has helped to reveal the origins of disease; however, cellular-scale resolution is not readily achievable in intact tissue volumes. Here, we demonstrate volumetric imaging of Young's modulus using ultrahigh-resolution optical coherence elastography, and apply it to characterizing the stiffness of mouse aortas. We achieve isotropic resolution of better than 15 µm over a 1-mm lateral field of view through the entire depth of an intact aortic wall. We employ a method of quasi-static compression elastography that measures volumetric axial strain and uses a compliant, transparent layer to measure surface axial stress. This combination is used to estimate Young's modulus throughout the volume. We demonstrate differentiation by stiffness of individual elastic lamellae and vascular smooth muscle. We observe stiffening of the aorta in regulator of G protein signaling 5-deficient mice, a model that is linked to vascular remodeling and fibrosis. We observe increased stiffness with proximity to the heart, as well as regions with micro-structural and micro-mechanical signatures characteristic of fibrous and lipid-rich tissue. High-resolution imaging of Young's modulus with optical coherence elastography may become an important tool in vascular biology and in other fields concerned with understanding the role of mechanics within the complex three-dimensional architecture of tissue.

Depth-encoded optical coherence elastography for simultaneous volumetric imaging of two tissue faces.

Depth-encoded optical coherence elastography (OCE) enables simultaneous acquisition of two three-dimensional (3D) elastograms from opposite sides of a sample. By the choice of suitable path-length differences in each of two interferometers, the detected carrier frequencies are separated, allowing depth-ranging from each interferometer to be performed simultaneously using a single spectrometer. We demonstrate depth-encoded OCE on a silicone phantom and a freshly excised sample of mouse liver. This technique minimizes the required spectral detection hardware and halves the total scan time. Depth-encoded OCE may expedite clinical translation in time-sensitive applications requiring rapid 3D imaging of multiple tissue surfaces, such as tumor margin assessment in breast-conserving surgery.

Ultrahigh-resolution optical coherence elastography.

Visualizing stiffness within the local tissue environment at the cellular and subcellular level promises to provide insight into the genesis and progression of disease. In this Letter, we propose ultrahigh-resolution optical coherence elastography (UHROCE), and demonstrate 3D imaging of local axial strain of tissues undergoing compressive loading. We combine optical coherence microscopy (OCM) and phase-sensitive detection of local tissue displacement to produce strain elastograms with resolution (x,y,z) of 2×2×15 µm. We demonstrate this performance on a freshly excised mouse aorta and reveal the mechanical heterogeneity of vascular smooth muscle cells and elastin sheets, otherwise unresolved in a typical, lower resolution optical coherence elastography (OCE) system.

Optical coherence tomography angiography for longitudinal monitoring of vascular changes in human cutaneous burns.

Assessment of vasculature is an important aspect of monitoring healing of cutaneous burn injuries. Recent advances in optical coherence tomography (OCT) have enabled it to be used to perform high-resolution imaging of the cutaneous vasculature in vivo, with the potential to provide a superior alternative to the conventional assessment of scoring skin color. The goal of this study is to investigate the feasibility of OCT angiography for longitudinal monitoring of vasculature and identification of vascular features in human cutaneous burns. We integrate several OCT imaging protocols and image-processing techniques into a systematic method for longitudinal monitoring and automatic quantification. The demonstration of this method on a partial-thickness burn shows the accurate co-location of longitudinal scans; characteristic vascular features in different healing phases; and eventual decrease of the elevated vasculature area density and vessel diameter to normal levels. Such a method holds promise for longitudinal monitoring of vasculature in burn injures as well as in other cutaneous vascular pathologies and responses to treatment.

Investigation of optical coherence micro-elastography as a method to visualize micro-architecture in human axillary lymph nodes.

BACKGROUND: Evaluation of lymph node involvement is an important factor in detecting metastasis and deciding whether to perform axillary lymph node dissection (ALND) in breast cancer surgery. As ALND is associated with potentially severe long term morbidity, the accuracy of lymph node assessment is imperative in avoiding unnecessary ALND. The mechanical properties of malignant lymph nodes are often distinct from those of normal nodes. A method to image the micro-scale mechanical properties of lymph nodes could, thus, provide diagnostic information to aid in the assessment of lymph node involvement in metastatic cancer. In this study, we scan axillary lymph nodes, freshly excised from breast cancer patients, with optical coherence micro-elastography (OCME), a method of imaging micro-scale mechanical strain, to assess its potential for the intraoperative assessment of lymph node involvement. METHODS: Twenty-six fresh, unstained lymph nodes were imaged from 15 patients undergoing mastectomy or breast-conserving surgery with axillary clearance. Lymph node specimens were bisected to allow imaging of the internal face of each node. Co-located OCME and optical coherence tomography (OCT) scans were taken of each sample, and the results compared to standard post-operative hematoxylin-and-eosin-stained histology. RESULTS: The optical backscattering signal provided by OCT alone may not provide reliable differentiation by inspection between benign and malignant lymphoid tissue. Alternatively, OCME highlights local changes in tissue strain that correspond to malignancy and are distinct from strain patterns in benign lymphoid tissue. The mechanical contrast provided by OCME complements the optical contrast provided by OCT and aids in the differentiation of malignant tumor from uninvolved lymphoid tissue. CONCLUSION: The combination of OCME and OCT images represents a promising method for the identification of malignant lymphoid tissue. This method shows potential to provide intraoperative assessment of lymph node involvement, thus, preventing unnecessary removal of uninvolved tissues and improving patient outcomes.

Optical coherence tomography in respiratory science and medicine: from airways to alveoli.

Optical coherence tomography is a rapidly maturing optical imaging technology, enabling study of the in vivo structure of lung tissue at a scale of tens of micrometers. It has been used to assess the layered structure of airway walls, quantify both airway lumen caliber and compliance, and image individual alveoli. This article provides an overview of the technology and reviews its capability to provide new insights into respiratory disease.

Full wave model of image formation in optical coherence tomography applicable to general samples.

We demonstrate a highly realistic model of optical coherence tomography, based on an existing model of coherent optical microscopes, which employs a full wave description of light. A defining feature of the model is the decoupling of the key functions of an optical coherence tomography system: sample illumination, light-sample interaction and the collection of light scattered by the sample. We show how such a model can be implemented using the finite-difference time-domain method to model light propagation in general samples. The model employs vectorial focussing theory to represent the optical system and, thus, incorporates general illumination beam types and detection optics. To demonstrate its versatility, we model image formation of a stratified medium, a numerical point-spread function phantom and a numerical phantom, based upon a physical three-dimensional structured phantom employed in our laboratory. We show that simulated images compare well with experimental images of a three-dimensional structured phantom. Such a model provides a powerful means to advance all aspects of optical coherence tomography imaging.

A compact source condition for modelling focused fields using the pseudospectral time-domain method.

The pseudospectral time-domain (PSTD) method greatly extends the physical volume of biological tissue in which light scattering can be calculated, relative to the finite-difference time-domain (FDTD) method. We have developed an analogue of the total-field scattered-field source condition, as employed in FDTD, for introducing focussed illuminations into PSTD simulations. This new source condition requires knowledge of the incident field, and applies update equations, at a single plane in the PSTD grid. Numerical artifacts, usually associated with compact PSTD source conditions, are minimized by using a staggered grid. This source condition's similarity with that used by the FDTD suggests a way in which existing FDTD codes can be easily adapted to PSTD codes.

Energy-efficient low-Fresnel-number Bessel beams and their application in optical coherence tomography.

Bessel beams feature a very large depth-of-focus (DOF) compared to conventional focusing schemes, but their central lobe carries only a small fraction of the total beam power, leading to a strongly reduced peak irradiance. This is problematic for power-limited applications, such as optical coherence tomography (OCT) or optical coherence microscopy, as it can result in a prohibitive reduction of the signal-to-noise ratio (SNR). Using scalar diffraction theory, we show that the trade-off between DOF and peak irradiance of Bessel beams depends solely on the Fresnel number N. We demonstrate the existence of a low-Fresnel-number regime, N<10, in which axicons with Gaussian illumination can generate energy-efficient Bessel beams with a small number of sidelobes. In the context of OCT, this translates into DOF enhancements of up to 13× for a SNR penalty below 20 dB, which is confirmed by our experiments. We expect that these findings will enable improved performance of optical systems with extended DOF.

Optical palpation: optical coherence tomography-based tactile imaging using a compliant sensor.

We present optical palpation, a tactile imaging technique for mapping micrometer- to millimeter-scale mechanical variations in soft tissue. In optical palpation, a stress sensor consisting of translucent, compliant silicone with known stress-strain behavior is placed on the tissue surface and a compressive load is applied. Optical coherence tomography (OCT) is used to measure the local strain in the sensor, from which the local stress at the sample surface is calculated and mapped onto an image. We present results in tissue-mimicking phantoms, demonstrating the detection of a feature embedded 4.7 mm below the sample surface, well beyond the depth range of OCT. We demonstrate the use of optical palpation to delineate the boundary of a region of tumor in freshly excised human breast tissue, validated against histopathology.

Optofluidic needle probe integrating targeted delivery of fluid with optical coherence tomography imaging.

We present an optofluidic optical coherence tomography (OCT) needle probe capable of modifying the local optical properties of tissue to improve needle-probe imaging performance. The side-viewing probe comprises an all-fiber-optic design encased in a hypodermic needle (outer diameter 720 µm) and integrates a coaxial fluid-filled channel, terminated by an outlet adjacent to the imaging window, allowing focal injection of fluid to a target tissue. This is the first fully integrated OCT needle probe design to incorporate fluid injection into the imaging mechanism. The utility of this probe is demonstrated in air-filled sheep lungs, where injection of small quantities of saline is shown, by local refractive index matching, to greatly improve image penetration through multiple layers of alveoli. 3D OCT images are correlated against histology, showing improvement in the capability to image lung structures such as bronchioles and blood vessels.

Towards standardising retinal OCT angiography image analysis with open-source toolbox OCTAVA.

Quantitative assessment of retinal microvasculature in optical coherence tomography angiography (OCTA) images is important for studying, diagnosing, monitoring, and guiding the treatment of ocular and systemic diseases. However, the OCTA user community lacks universal and transparent image analysis tools that can be applied to images from a range of OCTA instruments and provide reliable and consistent microvascular metrics from diverse datasets. We present a retinal extension to the OCTA Vascular Analyser (OCTAVA) that addresses the challenges of providing robust, easy-to-use, and transparent analysis of retinal OCTA images. OCTAVA is a user-friendly, open-source toolbox that can analyse retinal OCTA images from various instruments. The toolbox delivers seven microvascular metrics for the whole image or subregions and six metrics characterising the foveal avascular zone. We validate OCTAVA using images collected by four commercial OCTA instruments demonstrating robust performance across datasets from different instruments acquired at different sites from different study cohorts. We show that OCTAVA delivers values for retinal microvascular metrics comparable to the literature and reduces their variation between studies compared to their commercial equivalents. By making OCTAVA publicly available, we aim to expand standardised research and thereby improve the reproducibility of quantitative analysis of retinal microvascular imaging. Such improvements will help to better identify more reliable and sensitive biomarkers of ocular and systemic diseases.

The Prospective Association between Early Life Growth and Breast Density in Young Adult Women.

Breast density is a strong intermediate endpoint to investigate the association between early-life exposures and breast cancer risk. This study investigates the association between early-life growth and breast density in young adult women measured using Optical Breast Spectroscopy (OBS) and Dual X-ray Absorptiometry (DXA). OBS measurements were obtained for 536 female Raine Cohort Study participants at ages 27-28, with 268 completing DXA measurements. Participants with three or more height and weight measurements from ages 8 to 22 were used to generate linear growth curves for height, weight and body mass index (BMI) using SITAR modelling. Three growth parameters (size, velocity and timing) were examined for association with breast density measures, adjusting for potential confounders. Women who reached their peak height rapidly (velocity) and later in adolescence (timing) had lower OBS-breast density. Overall, women who were taller (size) had higher OBS-breast density. For weight, women who grew quickly (velocity) and later in adolescence (timing) had higher absolute DXA-breast density. Overall, weight (size) was also inversely associated with absolute DXA-breast density, as was BMI. These findings provide new evidence that adolescent growth is associated with breast density measures in young adult women, suggesting potential mediation pathways for breast cancer risk in later life.

Dual-modality needle probe for combined fluorescence imaging and three-dimensional optical coherence tomography.

To the best of our knowledge, we present the first needle probe for combined optical coherence tomography (OCT), and fluorescence imaging. The probe uses double-clad fiber (DCF) that guides the OCT signal and fluorescence excitation light in the core and collects and guides the returning fluorescence in the large-diameter multimode inner cladding. It is interfaced to a 1310 nm swept-source OCT system that has been modified to enable simultaneous 488 nm fluorescence excitation and >500 nm emission detection by using a DCF coupler to extract the returning fluorescence signal in the inner cladding with high efficiency. We present imaging results from an excised sheep lung with fluorescein solution infused through the vasculature. We were able to identify alveoli, bronchioles, and blood vessels. The results demonstrate that the combined OCT plus fluorescence needle images provide improved tissue differentiation over OCT alone.

Pilot study of optical coherence tomography angiography-derived microvascular metrics in hands and feet of healthy and diabetic people.

Optical coherence tomography angiography (OCTA) is a non-invasive, high-resolution imaging modality with growing application in dermatology and microvascular assessment. Accepted reference values for OCTA-derived microvascular parameters in skin do not yet exist but need to be established to drive OCTA into the clinic. In this pilot study, we assess a range of OCTA microvascular metrics at rest and after post-occlusive reactive hyperaemia (PORH) in the hands and feet of 52 healthy people and 11 people with well-controlled type 2 diabetes mellitus (T2DM). We calculate each metric, measure test-retest repeatability, and evaluate correlation with demographic risk factors. Our study delivers extremity-specific, age-dependent reference values and coefficients of repeatability of nine microvascular metrics at baseline and at the maximum of PORH. Significant differences are not seen for age-dependent microvascular metrics in hand, but they are present for several metrics in the foot. Significant differences are observed between hand and foot, both at baseline and maximum PORH, for most of the microvascular metrics with generally higher values in the hand. Despite a large variability over a range of individuals, as is expected based on heterogeneous ageing phenotypes of the population, the test-retest repeatability is 3.5% to 18% of the mean value for all metrics, which highlights the opportunities for OCTA-based studies in larger cohorts, for longitudinal monitoring, and for assessing the efficacy of interventions. Additionally, branchpoint density in the hand and foot and changes in vessel diameter in response to PORH stood out as good discriminators between healthy and T2DM groups, which indicates their potential value as biomarkers. This study, building on our previous work, represents a further step towards standardised OCTA in clinical practice and research.