ToScA North America (6 – 8 June 2017, The University of Texas, Austin, TX) Program

ToScA North America will address key areas of science, including Multi-modal Imaging, Geosciences, Forensics, Increasing Contrast, Educational Outreach, Data, Materials Science and Medical and Biological Science.University of Texas High-Resolution X-ray CT Facility (UTCT); Jackson School of Geosciences, The University of Texas at Austin; Natural History Museum (London); Royal Microscopical Society (Oxford, UK)Geological Science

Illuminating the Brain With X-Rays: Contributions and Future Perspectives of High-Resolution Microtomography to Neuroscience

The assessment of three-dimensional (3D) brain cytoarchitecture at a cellular resolution remains a great challenge in the field of neuroscience and constant development of imaging techniques has become crucial, particularly when it comes to offering direct and clear obtention of data from macro to nano scales. Magnetic resonance imaging (MRI) and electron or optical microscopy, although valuable, still face some issues such as the lack of contrast and extensive sample preparation protocols. In this context, x-ray microtomography (μCT) has become a promising non-destructive tool for imaging a broad range of samples, from dense materials to soft biological specimens. It is a new supplemental method to be explored for deciphering the cytoarchitecture and connectivity of the brain. This review aims to bring together published works using x-ray μCT in neurobiology in order to discuss the achievements made so far and the future of this technique for neuroscience

Development and in vitro characterization of three dimensional biodegradable scaffolds for peripheral nerve tissue engineering

Tissue engineering emerges nowadays to seek new solutions to damaged tissues and/or organs by replacing or repairing them with engineered constructs or scaffolds. In nerve tissue engineering, scaffolds for the repair of peripheral nerve injuries should act to support and promote axon growth following implantation. It is believed that substantial progress can be made by creating scaffolds from biomaterials, with growth-promoting molecules and spatially-controlled microstructure. To this end, this research aims to develop three dimensional (3D) scaffolds for peripheral nerve tissue regeneration by focusing on studies on the axon guidance, development and characterization of a novel 3D scaffold, and visualization of scaffolds by means of synchrotron-based diffraction enhanced imaging (DEI). Axon guidance is one of crucial considerations in developing of nerve scaffolds for nerve regeneration. In order to study the axon guidance mechanism, a two dimensional (2D) grid micropatterns were created by dispensing chitosan or laminin-blended chitosan substrate strands oriented in orthogonal directions; and then used in the in vitro dorsal root ganglion (DRG) neuron culture experiments. The results show the effect of the micropatterns on neurite directional growth can preferentially grow upon and follow the laminin-blended chitosan pathways. A novel 3D scaffold was developed for potential applications to peripheral nerve tissue engineering applications. The scaffolds were fabricated from poly L-lactide (PLLA) mixed with chitosan microspheres (CMs) by using a rapid freeze prototyping (RFP) technique, allowing for controllable scaffold microstructure and bioactivities protein release. The scaffold characterization shows that (1) the mechanical properties of the scaffolds depend on the ratio of CMs to PLLA as well as the cryogenic temperature and (2) the protein release can be controlled by adjusting the crosslink degree of the CMs and prolonged after the CMs were embedded into the PLLA scaffolds. Also, the degradation properties of the scaffolds were investigated with the results showing that the addition of CMs to PLLA can decrease the degradation rate as compared to pure PLLA scaffolds. This allows for another means to control the degradation rate. Visualization of polymer scaffolds in soft tissues is challenging, yet essential, to the success of tissue engineering applications. The x-ray diffraction enhanced imaging (DEI) method was explored for the visualization of the PLLA/CMs scaffolds embedded in soft tissues. Among various methods examined, including conventional radiography and in-line phase contrast imaging techniques, the DEI was the only technique able to visualize the scaffolds embedded in unstained muscle tissue as well as the microstructure of muscle tissue. Also, it has been shown that the DEI has the capacity to image the scaffolds in thicker tissue, and reduce the radiation doses to tissues as compared to conventional radiography. The methods and results developed/obtained in this study represent a substantial progress in the development and characterization of 3D scaffolds. This progress forms a basis for the future tests on the scaffolds as applied for peripheral nerve injuries

Characterization of alginate scaffolds using X-ray imaging techniques

Alginate is a popular biomaterial in tissue engineering. When crosslinked with calcium ions (Ca2+), alginate forms a hydrogel which provides necessary mechanical support as a scaffold. The material properties as well as the biological properties of alginate scaffold are of great importance. In this thesis, the aim is to use traditional methods, such as scanning electron microscopy (SEM) and light microscopy, and emerging X-ray imaging techniques, such as micro-computed tomography (micro-CT) and synchrotron radiation (SR) X-ray imaging, to characterize the alginate scaffolds. Firstly, the material properties of freeze-dried alginate scaffolds were evaluated using micro-CT, as it is a non-destructive and non-invasive imaging method, and can provide three-dimensional information. Alginate scaffolds made with different sodium alginate concentrations and frozen to different temperatures were scanned and analyzed in micro-CT. Results indicated that lower freezing temperature and higher sodium alginate concentration lead to smaller pore size and porosity. Secondly, cell culture experiments were carried out to study the biological properties and the interactions of alginate hydrogel with cells. A Schwann cell line was either blended with alginate solution before crosslinking with calcium chloride (CaCl2) or put around alginate gel in the culture dish. Light microscopy of sectioned slices showed that cells surrounding the alginate gel could not grow into the gel, while cells blended with alginate solution before crosslinking could proliferate inside the hydrogel. Cells grown inside a thin slice of alginate gels appeared to be in better condition and were larger in size and also grew in clusters. Thirdly, in order to image soft tissue buried inside alginate gels, such as brain slices, novel imaging methods based on synchrotron radiation (SR) were applied, such as absorption and phase contrast imaging, diffraction-enhanced imaging (DEI) and also combined with computed tomography (CT). Synchrotron-based monochromatic X-ray imaging proved to be good at distinguish objects of similar density, especially biological soft tissue samples, even without any staining material, such as osmium tetroxide (OsO4). These three pieces of research work show the potential in applying the emerging X-ray imaging in soft tissue engineering

High resolution 3D visualization of the spinal cord in a post-mortem murine model

A crucial issue in the development of therapies to treat pathologies of the central nervous system is represented by the availability of non-invasive methods to study the three-dimensional morphology of spinal cord, with a resolution able to characterize its complex vascular and neuronal organization. X-ray phase contrast micro-tomography enables a high-quality, 3D visualization of both the vascular and neuronal network simultaneously without the need of contrast agents, destructive sample preparations or sectioning. Until now, high resolution investigations of the post-mortem spinal cord in murine models have mostly been performed in spinal cords removed from the spinal canal. We present here post-mortem phase contrast micro-tomography images reconstructed using advanced computational tools to obtain high-resolution and high-contrast 3D images of the fixed spinal cord without removing the bones and preserving the richness of micro-details available when measuring exposed spinal cords. We believe that it represents a significant step toward the in-vivo application

Non-Invasive Microstructure and Morphology Investigation of the Mouse Lung: Qualitative Description and Quantitative Measurement

BACKGROUND: Early detection of lung cancer is known to improve the chances of successful treatment. However, lungs are soft tissues with complex three-dimensional configuration. Conventional X-ray imaging is based purely on absorption resulting in very low contrast when imaging soft tissues without contrast agents. It is difficult to obtain adequate information of lung lesions from conventional X-ray imaging. METHODS: In this study, a recently emerged imaging technique, in-line X-ray phase contrast imaging (IL-XPCI) was used. This powerful technique enabled high-resolution investigations of soft tissues without contrast agents. We applied IL-XPCI to observe the lungs in an intact mouse for the purpose of defining quantitatively the micro-structures in lung. FINDINGS: The three-dimensional model of the lung was successfully established, which provided an excellent view of lung airways. We highlighted the use of IL-XPCI in the visualization and assessment of alveoli which had rarely been studied in three dimensions (3D). The precise view of individual alveolus was achieved. The morphological parameters, such as diameter and alveolar surface area were measured. These parameters were of great importance in the diagnosis of diseases related to alveolus and alveolar scar. CONCLUSION: Our results indicated that IL-XPCI had the ability to represent complex anatomical structures in lung. This offered a new perspective on the diagnosis of respiratory disease and may guide future work in the study of respiratory mechanism on the alveoli level

Medical Imaging of Microrobots: Toward In Vivo Applications

Medical microrobots (MRs) have been demonstrated for a variety of non-invasive biomedical applications, such as tissue engineering, drug delivery, and assisted fertilization, among others. However, most of these demonstrations have been carried out in in vitro settings and under optical microscopy, being significantly different from the clinical practice. Thus, medical imaging techniques are required for localizing and tracking such tiny therapeutic machines when used in medical-relevant applications. This review aims at analyzing the state of the art of microrobots imaging by critically discussing the potentialities and limitations of the techniques employed in this field. Moreover, the physics and the working principle behind each analyzed imaging strategy, the spatiotemporal resolution, and the penetration depth are thoroughly discussed. The paper deals with the suitability of each imaging technique for tracking single or swarms of MRs and discusses the scenarios where contrast or imaging agent's inclusion is required, either to absorb, emit, or reflect a determined physical signal detected by an external system. Finally, the review highlights the existing challenges and perspective solutions which could be promising for future in vivo applications

Synchrotron imaging of bovine and human ovaries ex vivo

Background and Rationale: Reproductive dysfunction affects more than 15% of Canadian women; however, the underlying causes remain largely unknown. Ultrasonography is the most commonly used research and diagnostic tool for imaging the ovaries and uterus. However, current ultrasonographic techniques allow the detection of ovarian structures (eg. follicles, corpora lutea) at diameters of only ≥2 mm. The increased effectiveness of synchrotron technology for imaging ovaries in comparison to conventional imaging methods is currently unknown. Overall Objective: The overall objective of this research was to determine the effectiveness of synchrotron techniques for imaging ovaries. We hypothesized that synchrotron techniques would provide greater contrast for visualizing structural details of follicles, corpora lutea (CL), and cumulus oocyte complexes (COC), compared to conventional ultrasonography. Materials and Methods: Three studies were conducted to evaluate phase-contrast based synchrotron imaging methods. The first study involved Diffraction Enhanced Imaging (DEI) of bovine ovaries (n=6). The second study involved Propagation-Based Computed Tomography (PB-CT) imaging of bovine (n=4) and human ovaries (n=4). A third, preliminary study was conducted to explore the use of Talbot Grating Interferometry (TGI-CT) imaging of bovine (n=1) and human ovaries (n=1). Fresh and formalin-fixed bovine and human ovaries were imaged without or with contrast injection into the ovarian artery. Following synchrotron imaging, all ovarian samples were evaluated using diagnostic ultrasonography and histology. Images obtained using synchrotron techniques, ultrasonography and histology were qualitative and quantitatively compared. Results: DEI allowed the identification of 71% of follicles ≥2 mm and 67% of CL detected using ultrasonography. Mean follicle diameter was similar between DEI (9.6 ± 2.4 mm), ultrasonography (9.0 ± 2.6 mm), and histology (6.9 ± 1.9 mm) for fresh ovaries without contrast (P = 0.70). Likewise, no difference in CL diameter was detected between DEI (11.64 ± 1.67 mm), ultrasonography (9.34 ± 0.35 mm), and histology (9.6 ± 0.4 mm), (P = 0.34). Antral Follicle Count (AFC; ≥2mm) was similar between ultrasonography (6.5 ± 0.7 mm, fresh with no contrast; 6.5 ± 2.5 mm, preserved with no contrast) and DEI ( 4.5 ± 0.5 mm, fresh with no contrast; 6.5 ± 0.50 mm, preserved with no contrast) (P > 0.05). However, the contrast resolution for differentiating follicles and CL was inferior with DEI compared to ultrasonography. Small antral follicles <2mm, cell layers comprising the follicle wall and COC were not detected using either DEI or ultrasonography. PB-CT imaging enabled the visualization of 100% of follicles ≥2 mm and 100% of CL that were detected with ultrasonography. CL containing a central cystic cavity were identified using PB-CT; however, CL without a central cystic cavity were not well-visualized. Mean follicle and luteal diameters did not differ among PB-CT, ultrasonography and histology (P>0.05). PB-CT was superior to ultrasonography for detecting small antral follicles <2 mm in bovine ovaries (P = 0.04), and the granulosa and theca cell layers of the follicle wall in bovine and human ovaries (P < 0.0001). However, TGI-CT images exhibited greater contrast resolution for visualizing small and large antral follicles, CL, and the cell layers of the follicle wall compared to both PB-CT and ultrasonography. High contrast structures resembling COC were detected with both PB-CT and TGI-CT, but not with ultrasonography. Only TGI-CT permitted the visualization of the oocyte within the COC in fresh and preserved ovaries. Conclusions: DEI was inferior to ultrasonography for detecting ovarian follicles and CL. PB-CT was superior to ultrasonography for visualizing follicles <2 mm, COC, and the cell layers of the follicle wall. However, PB-CT was as effective as ultrasonography for detecting and measuring follicles ≥2 mm and cystic CL. Preliminary findings suggest that TGI-CT provides the greatest contrast for imaging both ovarian macro- and microanatomy compared to PB-CT, DEI, and ultrasonography