Biology subjects

Walsh, C. L.

Publications and source records attributed to Walsh, C. L..

14 recordsLinked to original sources

bioRxiv2026

Revealing multiscale architecture of the intact human spine using hierarchical phase contrast tomography

Background: Computed tomography (CT) and magnetic resonance imaging (MRI) provide detailed clinical assessment of the human spine; however, their millimeter-scale spatial resolution limits direct visualization of tissue microarchitecture and interfaces. Hierarchical phase-contrast tomography (HiP-CT) enables ex vivo non-destructive, micron-resolution imaging of intact human organs without exogenous contrast agents, extending anatomical assessment to spatial scales inaccessible to conventional clinical imaging. Purpose: To evaluate the capability of HiP-CT to visualize macro- to microanatomy of an intact adult human spine ex vivo. Methods: An intact spine (C1-L5) from a 62-year-old male donor without known skeletal disease was formalin-fixed and imaged by HiP-CT at the European Synchrotron. Whole-spine imaging was performed at 16.26 m isotropic voxels, with selected regions of interest imaged using 2.2 m voxels. Ex vivo MRI using clinical sequences was performed for anatomical comparison. Three-dimensional segmentation and quantitative analyses included intervertebral disc (IVD) grading, annulus fibrosus fiber tracing, and vertebral trabecular micro-morphometry. Results: HiP-CT enabled high spatial-resolution, multiscale imaging of the intact human spine, simultaneously visualizing vertebral bone and endplate, IVDs, spinal cord and meninges, nerve roots and ganglia, vasculature, ligaments, and musculature. Three-dimensional segmentation and virtual sectioning mapped neurovascular anatomy and vertebral vascular supply, without exogenous contrast enhancement, while preserving their relationships with surrounding tissues. Whole-spine images enabled quantitative analysis of annulus fibrosus fiber organization and vertebral trabecular microarchitecture, permitting the detection of focal posterolateral fiber disruption at the L4-L5 IVD and regional differences in trabecular architecture between cervical and lumbar vertebral bodies. Local tomography resolved annulus fibrosus lamellae, vertebral endplate Haversian systems, and facet joint hyaline cartilage at micron resolution. Conclusions: HiP-CT enables non-destructive, multiscale three-dimensional imaging of the intact human spine while preserving tissue relationships across spatial scales and hard-soft tissue boundaries. HiP-CT could provide an ex vivo microanatomical reference for future correlation with clinical imaging investigations of spinal disease and in the development and validation of imaging biomarkers.

bioengineering↗

bioRxiv2026

Multiscale mapping of venous remodelling in idiopathic pulmonary fibrosis

The integrity of the pulmonary vasculature is a key determinant of lung health, yet challenges in visualisation and quantification have hindered interrogation of its pathophysiological role in chronic lung disease. Here, we align recent advances in microscale image acquisition with novel computer vision-based vessel segmentation models to demonstrate expansion of the bronchial and pulmonary veins across varying severities of tissue remodelling in idiopathic pulmonary fibrosis (IPF). Relating these imaging findings to molecular data in control, mild, and severe fibrosis, we show that bronchial venous endothelial cells expand beyond their physiological peribronchial niche even in mild disease, acquiring persistent angiogenic, inflammatory, and matrix-remodelling programmes that define a specialised fibrovascular-immune interface. Finally, we relate these microscale and molecular observations to clinical CT imaging, demonstrating that intrapulmonary vein enlargement independently associates with worsened survival across three IPF cohorts. Collectively, our multimodal, multiscale approach connects previously unresolved three-dimensional venous architecture to its molecular endothelial correlate, establishing venous enlargement as a prognostically significant and clinically relevant feature of IPF. Our analytical approach also demonstrates how biological discoveries made in intact ex vivo human organs can translate into measurable phenotypes in living patients, providing a template for other organs and diseases.

pathology↗

bioRxiv2026

Hierarchical X-ray microscopy and mesoscopic diffusion MRI in the same brain reveal the human connectome across scales

We present a multimodal pipeline for 3D imaging of cerebral white-matter archi-tecture across scales, from whole-brain axonal projections down to individual myelinated axons. After diffusion MRI, an adult ex vivo human hemisphere undergoes label-free imaging with Hierarchical Phase-Contrast Tomography (HiP-CT) from 20 {micro}m/voxel in the whole hemisphere to 2 {micro}m/voxel in areas of interest, with intrinsic cross-scale alignment. A 4 cm tissue block extracted from the hemisphere is reimaged with HiP-CT at 0.857 {micro}m/voxel, enabling direct visualisation of single myelinated axons. After osmium staining, micro-CT at 0.364 {micro}m/voxel and electron microscopy at 4 nm/voxel are acquired in biop-sies from the tissue block to validate the presence of myelinated axons in the label-free HiP-CT contrast. Spanning three orders of magnitude in resolution, these co-registered multimodal datasets bridge microscopic wiring and macro-scopic brain organisation, providing a foundation for anatomically grounded whole-brain connectomics.

neuroscience↗

bioRxiv2026

Neuroanatomy of the clitoris

The clitoris is one of the least studied organs of the human body. The detailed anatomy of the clitoris is challenging to address through a gross dissection, as most of its parts are embedded internally, surrounded by pubic bone and several pelvic organs. While clinical imaging methods such as magnetic resonance imaging can capture the gross 3D morphology, they lack the spatial resolution required to resolve the detailed structures. In this study, we generated micron-scale computed tomography images of the female pelvises, leveraging a synchrotron radiation X-ray source. This unique data revealed the complex trajectory of the dorsal nerve of the clitoris, the main sensory nerve of the clitoris. Notably, the nerve trunks within the clitoral glans were revealed, with the maximum diameter ranging from 0.2 to 0.7 mm. They showed a tree-like branching pattern projecting towards the surface of the glans. We also revealed that some branches of the dorsal nerve of the clitoris ramify to innervate the clitoral hood and mons pubis. Finally, the posterior labial nerve, a branch of the perineal nerves, was shown to innervate the surroundings of the clitoris and the labial structures. These findings have an immediate impact on operations performed around the vulva area, such as gender-affirmation surgery and reconstruction surgery after genital mutilation.

neuroscience↗

bioRxiv2026

The Extremely Brilliant Brain: An Isotropic Microscale Human Brain Dataset

We present an isotropic 7.72 {micro}m/voxel post-mortem human brain dataset acquired using Hierarchical Phase-Contrast Tomography (HiP-CT) at the ESRF Extremely Brilliant Source, beamline BM18. This fills a critical gap between whole-brain MRI at 100 {micro}m resolution and serial-section histological reconstructions at 20 {micro}m or finer. HiP-CT contrast, derived from X-ray phase shifts, enables rich 3D visualisation of complex neuroanatomy including white-matter bundles, microvasculature, and sub-nuclei. We provide open-source workflows for online data exploration, subvolume download, segmentation, and reintegration of analyses into the full dataset. We demonstrate the potential of this resource by tracing vasculature over long distances, segmenting nuclei, and extracting whitematter orientations with 3D structure-tensor analysis. High-resolution human brain datasets are transformative for quantitative neuroanatomy, circuit mapping, and validation of clinical imaging; this openly available resource is a critical step for global access to next-generation multiscale brain imaging.

neuroscience↗

bioRxiv2025

The Human Organ Atlas

We present the Human Organ Atlas (HOA), an open data repository making accessible multiscale 3D imaging of human organs. The repository also provides software tools and training resources enabling worldwide access, sharing, and analysis of these datasets, facilitating further research and the continued expansion of the HOA. The images are generated using a synchrotron imaging technique - Hierarchical Phase-Contrast Tomography (HiP-CT) that uses the ESRFs Extremely Brilliant Source, spanning whole organ imaging at around 20 m/voxel with local volumes of interest within the intact organs imaged down to [~] 1 m/voxel. This offers a comprehensive exploration of human anatomy, providing unparalleled insights into intricate structures and spatial relationships. The Human Organ Atlas offers researchers, clinicians, and educators a valuable resource for anatomical study, image analysis, medical education, and large-scale data mining.

bioinformatics↗

bioRxiv2025

Multiscale Segmentation using Hierarchical Phase-contrast Tomography and Deep Learning

Biomedical systems span multiple spatial scales, encompassing tiny functional units to entire organs. Interpreting these systems through image segmentation requires the effective propagation and integration of information across different scales. However, most existing segmentation methods are optimised for single-scale imaging modalities, limiting their ability to capture and analyse small functional units throughout complete human organs. To facilitate multiscale biomedical image segmentation, we utilised Hierarchical Phase-Contrast Tomography (HiP-CT), an advanced imaging modality that can generate 3D multiscale datasets from high-resolution volumes of interest (VOIs) at ca. 1 {micro}m/voxel to whole-organ scans at ca. 20 {micro}m/voxel. Building on these hierarchical multiscale datasets, we developed a deep learning-based segmentation pipeline that is initially trained on manually annotated high-resolution HiP-CT data and then extended to lower-resolution whole-organ scans using pseudo-labels generated from high-resolution predictions and multiscale image registration. As a case study, we focused on glomeruli in human kidneys, benchmarking four 3D deep learning models for biomedical image segmentation on a manually annotated high-resolution dataset extracted from VOIs, at 2.58 to ca. 5 {micro}m/voxel, of four human kidneys. Among them, nnUNet demonstrated the best performance, achieving an average test Dice score of 0.906, and was subsequently used as the baseline model for multiscale segmentation in the pipeline. Applying this pipeline to two low-resolution full-organ data at ca. 25 {micro}m/voxel, the model identified 1,019,890 and 231,179 glomeruli in a 62-year-old donor without kidney diseases and a 94-year-old hypertensive donor, enabling comprehensive morphological analyses, including cortical spatial statistics and glomerular distributions, which aligned well with previous anatomical studies. Our results highlight the effectiveness of the proposed pipeline for segmenting small functional units in multiscale bioimaging datasets and suggest its broader applicability to other organ systems.

bioinformatics↗

bioRxiv2024

Vasculature segmentation in 3D hierarchical phase-contrast tomography images of human kidneys

Efficient algorithms are needed to segment vasculature in new three-dimensional (3D) medical imaging datasets at scale for a wide range of research and clinical applications. Manual segmentation of vessels in images is time-consuming and expensive. Computational approaches are more scalable but have limitations in accuracy. We organized a global machine learning competition, engaging 1,401 participants, to help develop new deep learning methods for 3D blood vessel segmentation. This paper presents a detailed analysis of the top-performing solutions using manually curated 3D Hierarchical Phase-Contrast Tomography datasets of the human kidney, focusing on the segmentation accuracy and morphological analysis, thereby establishing a benchmark for future studies in blood vessel segmentation within phase-contrast tomography imaging.

bioinformatics↗

bioRxiv2024

A unique subset of pericystic endothelium associates with aberrant microvascular remodelling and impaired blood perfusion early in polycystic kidney disease

Hallmarks of autosomal dominant polycystic kidney disease (ADPKD), the most common hereditary kidney anomaly, include expanding fluid-filled epithelial cysts, inflammation, and fibrosis. Despite previous work showing the potential of vascular-based therapies, renal microvascular alterations in ADPKD, and their timing, are poorly understood. Using single-cell transcriptomics of human kidney microvasculature, we identify a population of endothelial cells adjacent to cysts in ADPKD. This pericystic endothelium, distinguishable by its expression of osteopontin (SPP1), has a distinct molecular profile compared to the common endothelial cell injury signature in other kidney diseases. SPP1+ pericystic endothelium was also present in an orthologous mouse model of ADPKD before overt kidney functional decline. By interrogating geometric, topological and fractal properties from three-dimensional imaging of early ADPKD mouse kidneys, we show that pericystic endothelium associates with disorganisation and non-uniformity of the renal cortical microvasculature. Concurrently, we detected region-specific reductions in cortical blood flow within ADPKD murine kidneys using arterial spin labelling. We conclude that ADPKD kidneys contain a unique subset of endothelium manifesting with aberrant remodelling and impaired blood perfusion. Its detection, prior to renal functional decline, advocates the vasculature as a therapeutic target to modulate or preserve renal function in early ADPKD.

pathology↗

bioRxiv2023

Multidimensional Analysis of the Adult Human Heart in Health and Disease using Hierarchical Phase-Contrast Tomography (HiP-CT)

Cardiovascular diseases (CVDs) are a leading cause of death worldwide. Current clinical imaging modalities provide resolution adequate for diagnosis but are unable to provide detail of structural changes in the heart, across length-scales, necessary for understanding underlying pathophysiology of disease. Hierarchical Phase-Contrast Tomography (HiP-CT), using new (4th) generation synchrotron sources, potentially overcomes this limitation, allowing micron resolution imaging of intact adult organs with unprecedented detail. In this proof of principle study (n=2), we show the utility of HiP-CT to image whole adult human hearts ex-vivo: one control without known cardiac disease and one with multiple known cardiopulmonary pathologies. The resulting multiscale imaging was able to demonstrate exemplars of anatomy in each cardiac segment along with novel findings in the cardiac conduction system, from gross (20 um/voxel) to cellular scale (2.2 um/voxel), non-destructively, thereby bridging the gap between macroscopic and microscopic investigations. We propose that the technique represents a significant step in virtual autopsy methods for studying structural heart disease, facilitating research into abnormalities across scales and age-groups. It opens up possibilities for understanding and treating disease; and provides a cardiac blueprint with potential for in-silico simulation, device design, virtual surgical training, and bioengineered heart in the future.

pathology↗

bioRxiv2023

Micro to macro scale analysis of the intact human renal arterial tree with Synchrotron Tomography

The architecture of the kidney vasculature is essential for its function. Although structural profiling of the intact rodent kidney vasculature has been performed, it is challenging to map vascular architecture of larger human organs. We hypothesised that hierarchical phase-contrast tomography (HiP-CT) would enable quantitative analysis of the entire human kidney vasculature. Combining label-free HiP-CT imaging of an intact kidney from a 63-year-old male with topology network analysis, we quantitated vasculature architecture in the human kidney down to the scale of arterioles. Although human and rat kidney vascular topologies are comparable, vascular radius decreases at a significantly faster rate in humans as vessels branch from artery towards the cortex. At branching points of large vessels, radii are theoretically optimised to minimise flow resistance, an observation not found for smaller arterioles. Structural differences in the vasculature were found in different spatial zones of the kidney reflecting their unique functional roles. Overall, this represents the first time the entire arterial vasculature of a human kidney has been mapped providing essential inputs for computational models of kidney vascular flow and synthetic vascular architectures, with implications for understanding how the structure of individual blood vessels collectively scales to facilitate organ function.

bioengineering↗

bioRxiv2023

A closer look at high-energy X-ray-induced bubble formation during soft tissue imaging

Improving the scalability of tissue imaging throughput with bright, coherent X-rays requires identifying and mitigating artifacts resulting from the interactions between X-rays and matter. At synchrotron sources, long-term imaging of soft tissues in solution can result in gas bubble formation or cavitation, which dramatically compromises image quality and integrity of the samples. By combining in-line phase-contrast cineradiography with operando gas chromatography, we were able to track the onset and evolution of high-energy X-ray-induced gas bubbles in ethanol-embedded soft tissue samples for tens of minutes (2 to 3 times the typical scan times). We demonstrate quantitatively that vacuum degassing of the sample during preparation can significantly delay bubble formation, offering up to a twofold improvement in dose tolerance, depending on the tissue type. However, once nucleated, bubble growth is faster in degassed than undegassed samples, indicating their distinct metastable states at bubble onset. Gas chromatography analysis shows increased solvent vaporization concurrent with bubble formation, yet the quantities of dissolved gases remain unchanged. Coupling features extracted from the radiographs with computational analysis of bubble characteristics, we uncover dose-controlled kinetics and nucleation site-specific growth. These hallmark signatures provide quantitative constraints on the driving mechanisms of bubble formation and growth. Overall, the observations highlight bubble formation as a critical, yet often overlooked hurdle in upscaling X-ray imaging for biological tissues and soft materials and we offer an empirical foundation for their understanding and imaging protocol optimization. More importantly, our approaches establish a top-down scheme to decipher the complex, multiscale radiation-matter interactions in these applications. Significance statementBetter probing the X-ray radiation dose limit of bubble formation in biological tissue and developing mitigation methods is essential for improving imaging techniques involving X-ray, such as synchrotron X-ray tomography or crystallography. Here, we combined operando gas chromatography with in-line X-ray phase-contrast radiography on human lung and brain tissue to investigate bubble formation under high-energy X-ray irradiation. We demonstrate that vacuum degassing delays bubble nucleation up to a factor two, depending on the tissue type. Gas chromatography analysis showed increased solvent vaporization during bubble formation; however, the quantities of dissolved gases remained unchanged. Moreover, depending on the nucleation site, bubble growth can be geometrically constrained by sample microstructure, which influence its dynamics.

biochemistry↗

bioRxiv2022

Preparation of large biological samples for high-resolution, hierarchical, multi-modal imaging

Imaging the different scales of biological tissue is essential for understanding healthy organ behavior and pathophysiological changes. X-ray micro-tomography using both laboratory ({micro}CT) and synchrotron sources (sCT) is a promising tool to image the 3D morphology at the macro- and micro-scale of large samples, including intact human organs. Preparation of large samples for high resolution imaging techniques remains a challenge due to limitations with current methods, such as sample shrinkage, insufficient contrast, movement of the sample and bubble formation during mounting or scanning. Here, we describe a protocol to prepare, stabilize, and image large soft-tissue samples with X-ray microtomography. We demonstrate the protocol using intact human organs and Hierarchical Phase-Contrast Tomography (HiP-CT) imaging at the European Synchrotron Radiation Facility, but the protocol is equally applicable to a range of biological samples, including complete organisms, for both laboratory and synchrotron source tomography. Our protocol enhances the contrast of the sample, while preventing sample motion during the scan, even in case of different sample orientations. Bubbles trapped during mounting and those formed during scanning (in case of synchrotron X-ray imaging) are mitigated by multiple degassing steps. The sample preparation is also compatible with magnetic resonance imaging (MRI), CT, and histological observation. We describe a protocol for sample preparation and mounting which requires 25 to 34 days for a large organ such as an intact human brain or heart. This preparation time varies depending on the composition, size, and fragility of the tissue. Use of the protocol enables scanning of intact organs with a diameter of 150 mm with a local pixel size of one micron using HiP-CT.

physiology↗

bioRxiv2021

A multiscale X-ray phase-contrast tomography dataset of whole human left lung

Technological advancements in X-ray imaging using bright and coherent synchrotron sources now allows to decouple sample size and resolution, while maintaining high sensitivity to the microstructure of soft, partially dehydrated tissues. The recently developed imaging technique, hierarchical phase-contrast tomography, is a comprehensive approach to address the challenge of organ-scale (up to tens of centimeters) soft tissue imaging with resolution and sensitivity down to the cellular level. Using this technique, we imaged ex vivo an entire human left lung at an isotropic voxel size of 25.08 m along with local zooms down to 6.05 - 6.5 m and 2.45 - 2.5 m in voxel size. The high tissue contrast offered by the fourth-generation synchrotron source at the European Synchrotron Radiation Facility reveals complex multiscale anatomical constitution of the human lung from the macroscopic (centimeter) down to the microscopic (micrometer) scale. The dataset provides complete organ-scale 3D information of the secondary pulmonary lobules and delineates the microstructure of lung nodules with unprecedented detail.

physiology↗