Synchrotron-based visualization and segmentation of elastic lamellae in the mouse carotid artery during quasi-static pressure inflation

This dataset contains images that were obtained during quasi-static pressure inflation of mouse carotid arteries. Images were taken with phase propagation imaging at the X02DA TOMCAT beamline of the Swiss Light Source synchrotron at the Paul Scherrer Institute in Villigen, Switzerland. Scans of n=12 left carotid arteries (n-6 Apoe-deficient mice, n=6 wild-type mice, all on a C57Bl6J background) were taken at pressure levels of 0, 10, 20, 30, 40, 50, 70, 90 and 120 mmHg. For analysis we selected 75 images from the center of each stack (starting at the center of the stack, and skipping 2 of every three images in both cranial and caudal axial directions) for each sample and for each pressure level, resulting in a total of 75 x 12 x 9 = 8100 analyzed images from 108 different scans. Segmentation, 3D visualization and geometric analysis is presented in the corresponding manuscript. Files are uploaded in 16bit .tif format and are named: mouseid_pressurelevel_stacknumber, with mouseid consisting of either Apoe (Apoe-deficient) or Bl (wild-type) and the mouse number, pressurelevel varies from P0 to P120 and stacknumber indicates which image from the stack has been uploaded

Micro-structural damage during the early phase of Angiotensin II-induced dissecting aortic aneurysm : the role of aortic biomechanics

Angiotensin II-infused ApoE-/- mice are a popular model for aortic aneurysm and dissection. We have recently demonstrated that the thoraco-abdominal lesions in these mice start with a medial tear near the ostia of celiac and mesenteric arteries. Given the location-specific nature of the disease, we hypothesized that the local mechanical equilibrium may drive disease initiation [1]. In order to investigate this hypothesis we subsequently developed a novel computational approach to evaluate the in-vivo strain field in the abdominal aorta. Combining ex vivo synchrotron images with in vivo micro- CT, we incorporated model features such as non-uniform aortic wall thickness, nonuniform stretch field and the inclusion of small aortic side branches into our computational models and showed how these often overlooked features impact the location of hotspots in the computed strain field [2]. In our current work we validate these simulations with image-guided histology in order to investigate whether regions of high strain collocate with sites of micro-structural. N=10 ApoE-/- mice were infused with Angiotensin-II for 3 days and subsequently underwent a contrast-enhanced micro- CT scan prior to euthanasia. The aorta was imaged ex-vivo using high-resolution Phasecontrast X-Ray Microscopy (PCXTM) at 6.5 um isotropic resolution. The same protocol was followed for n=6 saline-infused controls. An in-house automated framework was implemented to morph the non-pressurized non-stretched ex-vivo PCXTM geometry onto the pressurized stretched in-vivo micro-CT geometry [2]. For each animal the output was a mouse-specific structural finite element simulation. Contrast agent infiltration in the aortic wall was used to detect the location of micro-ruptures in the tunica media [1] and image-guided histology was performed to validate and quantify the vascular damage. Preliminary results show good agreement between hotspots of early vascular damage and hotspots of computed maximal strain. The highest strain values occurred invariably in the vicinity of the celiac and mesenteric arteries and collocated with intramural micro-ruptures and leukocyte infiltration. Moreover, the intersubject variability of the maximal strain locations (cranial/caudal or right/left of the ostium) corresponded qualitatively to the inter-subject variability of PCXTM-detected contrast agent leakage. We conclude that strain concentrations near side branches may play an important role in disease initiation and could partially explain the focal nature of the disease

Unravelling the aortic microstructure : synchrotron-based quasi-static pressure inflation of the mouse carotid artery

The contribution of the aortic microstructure to the mechanical behavior of the aortic wall is poorly understood. Several high-resolution techniques have been proposed to visualize elastic lamellae or collagen fibers, but most have a limited field of view and are challenging to perform in pressurized conditions. In recent experiments we visualized the micro-structure of mouse aortas using phase propagation imaging – a synchrotron-based technique that yielded 3D images on which separate lamellar layers could be identified (unpublished data, manuscript in preparation). In the experimental study that is presented here we used phase propagation imaging to quantify, for the first time, the unfolding of aortic lamellae during quasi-static pressure inflation experiments. Six wild type and six ApoE-/- mice, all male and on a C57Bl6/J background, were used for this study. The left carotid artery was harvested immediately after sacrifice and mounted on a dedicated synchrotron-compatible pressure inflation device. During the experiment pressure was increased quasi-statically with a syringe pump and maintained at a constant level during each imaging step. After two initial loops of 0-120 mmHg to precondition the vessel, scans were taken at pressure levels of 0, 10, 20, 30, 40, 50, 70, 90 and 120 mmHg while the axial stretch was kept at the in vivo value. Phase propagation was performed at 25m source-to-sample distance, 25 cm sample-todetector distance and at 21 keV. A scientific CMOS detector (pco.Edge 5.5) was used in combination with a 4x magnifying visible-light optics and a 20 μm thick scintillator. The effective pixel size was 1.625 x 1.625 μm2. During post-processing the images were skeletonized and a bi-directional graph was generated in Matlab. Using a modified Dijkstra algorithm in which lower weights were assigned to the edges closest to the center of the vessel, we created a Matlab-based algorithm that allows us to automatically segment the main micro-structural features each of the three lamellar layers in the carotid artery. The algorithm exploits the edge connectivity and the shortest path constraints, and weights of edges belonging to the shortest path are subsequently increased order to allow for the detection of subsequent layers. After filtering and de-trending the signal, the undulation of each layer was quantified from the prominence of the peaks in the signal. Both in ApoE-/- and wild type mice we were able to quantify how the increased straightening of the lamellar layers in response to the increasing pressure related to the change in vessel diameter that is quantified in traditional biomechanical experiments. In future work we intend to use the synchrotroncompatible pressure-inflation device in order to experimentally determine the microstructural material properties of aortic lamellae and the interlamellar space

Propagation-based phase-contrast synchrotron imaging of aortic dissection in mice : from individual elastic lamella to 3D analysis

In order to show the advantage and potential of propagation-based phase-contrast synchrotron imaging in vascular pathology research, we analyzed aortic medial ruptures in BAPN/AngII-infused mice, a mouse model for aortic dissection. Ascending and thoraco-abdominal samples from n = 3 control animals and n = 10 BAPN/AngII-infused mice (after 3, 7 and 14 days of infusion, total of 24 samples) were scanned. A steep increase in the number of ruptures was already noted after 3 days of BAPN/AngII-infusion. The largest ruptures were found at the latest time points. 133 ruptures affected only the first lamella while 135 ruptures affected multiple layers. Medial ruptures through all lamellar layers, leading to false channel formation and intramural hematoma, occurred only in the thoraco-abdominal aorta and interlamellar hematoma formation in the ascending aorta could be directly related to ruptures of the innermost lamellae. The advantages of this technique are (i) ultra-high resolution that allows to visualize the individual elastic lamellae in the aorta; (ii) quantitative and qualitative analysis of medial ruptures; (iii) 3D analysis of the complete aorta; (iv) high contrast for qualitative information extraction, reducing the need for histology coupes; (v) earlier detection of (micro-) ruptures

Synchrotron-based phase contrast imaging of cardiovascular tissue in mice-grating interferometry or phase propagation?

Synchrotron-based x-ray phase-contrast imaging allows for detailed 3D insight into the microstructure of soft tissue and is increasingly used to improve our understanding of mouse models of cardiovascular disease. Two techniques dominate the field: grating interferometry, with superior density contrast at mid to lower microscopic resolutions, and propagation-based phase contrast, facilitating high-resolution tissue imaging. The choice between these techniques depends on which features one is interested in visualizing and is thus highly sample-dependent. In this manuscript we systematically evaluate the advantages and disadvantages of grating interferometry and propagation-based phase contrast for the specific application of pre-clinical cardiovascular tissue. We scanned samples obtained from 5 different mouse models of cardiovascular disease, ranging from carotid plaques over ascending and abdominal aortic aneurysms to hypertrophic hearts. Based on our findings we discuss in detail how synchrotron-based imaging can be used to increase our understanding of the anatomy and biomechanics of cardiovascular disease in mice. We also present a flowchart that can help future users to select the best synchrotron-based phase contrast technique for their pre-clinical cardiovascular samples

Co-localization of microstructural damage and excessive mechanical strain at aortic branches in angiotensin-II-infused mice

Animal models of aortic aneurysm and dissection can enhance our limited understanding of the etiology of these lethal conditions particularly because early-stage longitudinal data are scant in humans. Yet, the pathogenesis of often-studied mouse models and the potential contribution of aortic biomechanics therein remain elusive. In this work, we combined micro-CT and synchrotron-based imaging with computational biomechanics to estimate in vivo aortic strains in the abdominal aorta of angiotensin-II-infused ApoE-deficient mice, which were compared with mouse-specific aortic microstructural damage inferred from histopathology. Targeted histology showed that the 3D distribution of micro-CT contrast agent that had been injected in vivo co-localized with precursor vascular damage in the aortic wall at 3 days of hypertension, with damage predominantly near the ostia of the celiac and superior mesenteric arteries. Computations similarly revealed higher mechanical strain in branching relative to non-branching regions, thus resulting in a positive correlation between high strain and vascular damage in branching segments that included the celiac, superior mesenteric, and right renal arteries. These results suggest a mechanically driven initiation of damage at these locations, which was supported by 3D synchrotron imaging of load-induced ex vivo delaminations of angiotensin-II-infused suprarenal abdominal aortas. That is, the major intramural delamination plane in the ex vivo tested aortas was also near side branches and specifically around the celiac artery. Our findings thus support the hypothesis of an early mechanically mediated formation of microstructural defects at aortic branching sites that subsequently propagate into a macroscopic medial tear, giving rise to aortic dissection in angiotensin-II-infused mice