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Abstract: Additive manufacturing (AM) platforms allow the production of patient tissue engineering scaffolds with desirable architectures. Although AM platforms offer exceptional control on architecture, post-processing methods such as sintering and freeze-drying often deform the printed scaffold structure. In-situ 4D imaging can be used to analyze changes that occur during post-processing. Visualization and analysis of changes in selected volumes of interests (VOIs) over time are essential to understand the underlining mechanisms of scaffold deformations. Yet, automated detection and tracking of VOIs in the 3D printed scaffold over time using 4D image data is currently an unsolved image processing task. This paper proposes a new image processing technique to segment, detect and track volumes of interest in 3D printed tissue engineering scaffolds. The method is validated using a 4D synchrotron sourced microCT image data captured during the sintering of bioactive glass scaffolds in-situ. The proposed method will contribute to the development of scaffolds with controllable designs and optimum properties for the development of patient-specific scaffolds.

Open Access

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Abstract: Multi-scale structural assessment of biological soft tissue is challenging but essential to gain insight into structure–function relationships of tissue/organ. Using the human placenta as an example, this study brings together sophisticated sample preparation protocols, advanced imaging and robust, validated machine-learning segmentation techniques to provide the first massively multi-scale and multi-domain information that enables detailed morphological and functional analyses of both maternal and fetal placental domains. Finally, we quantify the scale-dependent error in morphological metrics of heterogeneous placental tissue, estimating the minimal tissue scale needed in extracting meaningful biological data. The developed protocol is beneficial for high-throughput investigation of structure–function relationships in both normal and diseased placentas, allowing us to optimize therapeutic approaches for pathological pregnancies. In addition, the methodology presented is applicable in the characterization of tissue architecture and physiological behaviours of other complex organs with similarity to the placenta, where an exchange barrier possesses circulating vascular and avascular fluid spaces.

Open Access

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Abstract: Lymph nodes (LN) are crucial for immune function, and comprise an important interface between the blood and lymphatic systems. Blood vessels (BV) in LN are highly specialized, featuring high endothelial venules across which most of the resident lymphocytes crossed. Previous measurements of overall lymph and BV flow rates demonstrated that fluid also crosses BV walls, and that this is important for immune function. However, the spatial distribution of the BV in LN has not been quantified to the degree necessary to analyse the distribution of transmural fluid movement. In this study, we seek to quantify the spatial localization of LNBV, and to predict fluid movement across BV walls. MicroCT imaging of murine popliteal LN showed that capillaries were responsible for approximately 75% of the BV wall surface area, and that this was mostly distributed around the periphery of the node. We then modelled blood flow through the BV to obtain spatially resolved hydrostatic pressures, which were then combined with Starling’s law to predict transmural flow. Much of the total 10 nL/min transmural flow (under normal conditions) was concentrated in the periphery, corresponding closely with surface area distribution. These results provide important insights into the inner workings of LN, and provide a basis for further exploration of the role of LN flow patterns in normal and pathological functions.

Open Access

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Abstract: Bioglass® was the first material to form a stable chemical bond with human tissue. Since its discovery, a key goal was to produce three-dimensional (3D) porous scaffolds which can host and guide tissue repair, in particular, regeneration of long bone defects resulting from trauma or disease. Producing 3D scaffolds from bioactive glasses is challenging because of crystallization events that occur while the glass particles densify at high temperatures. Bioactive glasses such as the 13–93 composition can be sintered by viscous flow sintering at temperatures above the glass transition onset (Tg) and below the crystallization temperature (Tc). There is, however, very little literature on viscous flow sintering of bioactive glasses, and none of which focuses on the viscous flow sintering of glass scaffolds in four dimensions (4D) (3D + time). Here, high-resolution synchrotron-sourced X-ray computed tomography (sCT) was used to capture and quantify viscous flow sintering of an additively manufactured bioactive glass scaffold in 4D. In situ sCT allowed the simultaneous quantification of individual particle (local) structural changes and the scaffold's (global) dimensional changes during the sintering cycle. Densification, calculated as change in surface area, occurred in three distinct stages, confirming classical sintering theory. Importantly, our observations show for the first time that the local and global contributions to densification are significantly different at each of these stages: local sintering dominates stages 1 and 2, while global sintering is more prevalent in stage 3. During stage 1, small particles coalesced to larger particles because of their higher driving force for viscous flow at lower temperatures, while large angular particles became less faceted (angular regions had a local small radius of curvature). A transition in the rate of sintering was then observed in which significant viscous flow occurred, resulting in large reduction of surface area, total strut volume, and interparticle porosity because the majority of the printed particles coalesced to become continuous struts (stage 2). Transition from stage 2 to stage 3 was distinctly obvious when interparticle pores became isolated and closed, while the sintering rate significantly reduced. During stage 3, at the local scale, isolated pores either became more spherical or reduced in size and disappeared depending on their initial morphology. During stage 3, sintering of the scaffolds continued at the strut level, with interstrut porosity reducing, while globally the strut diameter increased in size, suggesting overall shrinkage of the scaffold with the flow of material via the strut contacts. This study provides novel insights into viscous flow in a complex non-idealized construct, where, locally, particles are not spherical and are of a range of sizes, leading to a random distribution of interparticle porosity, while globally, predesigned porosity between the struts exists to allow the construct to support tissue growth. This is the first time that the three stages of densification have been captured at the local and global scales simultaneously. The insights provided here should accelerate the development of 3D bioactive glass scaffolds.

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Abstract: Screw-assisted material extrusion technique is developed for tissue engineering applications to produce scaffolds with well-defined multiscale microstructural features and tailorable mechanical properties. In this study, in situ time-resolved synchrotron diffraction is employed to probe extrusion-based 3D printing of polycaprolactone (PCL) filaments. Time-resolved X-ray diffraction measurements reveals the progress of overall crystalline structural evolution of PCL during 3D printing. Particularly, in situ experimental observations provide strong evidence for the development of strong directionality of PCL crystals during the extrusion driven process. Results also show the evidence for the realization of anisotropic structural features through the melt extrusion-based 3D printing, which is a key development toward mimicking the anisotropic properties and hierarchical structures of biological materials in nature, such as human tissues.