Nano-computed Tomography Research Articles

Bone-targeted ultrasound-responsive nanobubbles for siRNA delivery to treat osteoporosis in mice

This project aimed to study the efficacy of a bone-targeted ultrasound-responsive nanobubble (NB) platform to deliver gene-silencing cathepsin K (CTSK) siRNA into the bone for osteoporosis treatment using in vitro and in vivo studies. To this end, characterization of CTSK siRNA loaded NB functionalized with alendronate (NB-CTSK siRNA-AL) was performed using transmission electron microscopy (TEM) imaging, and a release profile was obtained through fluorescent spectroscopy. In vitro studies were conducted by culturing NB-CTSK siRNA-AL with osteoclasts to evaluate siRNA uptake, CTSK expression, and the expression of tartrate-resistant acid phosphatase (TRAP). A control group and an NB-CTSK siRNA-AL treated group of ovariectomized (OVX) mice (n = 4) were tested. The OVX group that received treatment underwent weekly sessions for 4 weeks, during which they were exposed to low-intensity pulsed ultrasound (LIPUS) stimulation following administration of NB-CTSK siRNA-AL, prior to being sacrificed. Both groups underwent a series of tests to evaluate the bone targeting, safety, and efficacy of the nanoplatform. These tests included biodistribution studies conducted at 4 h and 24 h post-injection, a 3-point bending test of the femurs, nano-computed tomography analysis, as well as Hematoxylin & Eosin histological staining, Masson's Trichrome staining, and CTSK staining. The biodistribution showed the accumulation of NB-CTSK siRNA-AL in the bone and liver. Results showed that the OVX mice treated with NB-CTSK siRNA-AL had increased distal cortical bone thickness (174.4 ± 5.28 μm vs. 144.3 ± 10.66 μm, p > 0.05)) and bone volume fraction (16.5 ± 3.96 % vs. 6.55 ± 0.13 % (p > 0.05)). A reduced collagen degradation and downregulated CTSK expression were evident in the staining procedures. No adverse effects were recorded within histological assessments on the liver, kidney, and heart post-treatment. Morphology was shown to be normal and healthy within muscle cells post-LIPUS stimulation of NB-CTSK siRNA-AL. From these results, it can be concluded that an ultrasound-mediated NB-CTSK siRNA-AL can serve as a reliable, safe CTSK siRNA carrier to bone-specific targets for in vivo osteoporosis treatment.

Enhancing craniofacial bone tissue engineering strategy: integrating rapid wet chemically synthesised bioactive glass with photopolymerized resins

BackgroundCraniofacial bone regeneration represents a dynamic area within tissue engineering and regenerative medicine. Central to this field, is the continual exploration of new methodologies for template fabrication, leveraging established bio ceramic materials, with the objective of restoring bone integrity and facilitating successful implant placements.MethodsPhotopolymerized templates were prepared using three distinct bio ceramic materials, specifically a wet chemically synthesized bioactive glass and two commercially sourced hydroxyapatite variants. These templates underwent comprehensive characterization to assess their physicochemical and mechanical attributes, employing techniques including Fourier transform infrared spectroscopy, scanning electron microscopy, and nano-computed tomography. Evaluation of their biocompatibility was conducted through interaction with primary human osteoblasts (hOB) and subsequent examination using scanning electron microscopy.ResultsThe results demonstrated that composite showed intramolecular hydrogen bonding interactions with the photopolymer, while computerized tomography unveiled the porous morphology and distribution within the templates. A relatively higher porosity percentage (31.55 ± 8.70%) and compressive strength (1.53 ± 0.11 MPa) was noted for bioactive glass templates. Human osteoblast cultured on bioactive glass showed higher viability compared to other specimens. Scanning micrographs of human osteoblast on templated showed cellular adhesion and the presence of filopodia and lamellipodia.ConclusionIn summary these templates have the potential to be used for alveolar bone regeneration in critical size defect. Photopolymerization of bioceramics may be an interesting technique for scaffolds fabrication for bone tissue engineering application but needs more optimization to overcome existing issues like the ideal ratio of the photopolymer to bioceramics.

Laboratory x-ray nano-computed tomography for biomedical research

High-resolution x-ray tomography is a common technique for biomedical research using synchrotron sources. With advancements in laboratory x-ray sources, an increasing number of experiments can be performed in the lab. In this paper, the design, implementation, and verification of a laboratory setup for x-ray nano-computed tomography is presented using a nano-focus x-ray source and high geometric magnification not requiring any optical elements. Comparing a scintillator-based detector to a photon counting detector shows a clear benefit of using photon counting detectors for these applications, where the flux of the x-ray source is limited and samples have low contrast. Sample contrast is enhanced using propagation-based phase contrast. The resolution of the system is verified using 2D resolution charts and using Fourier Ring Correlation on reconstructed CT slices. Evaluating noise and contrast highlights the benefits of photon counting detectors and the contrast improvement through phase contrast. The implemented setup is capable of reaching sub-micron resolution and satisfying contrast in biological samples, like paraffin embedded tissue.

Failure Analysis of Ni-8YSZ Electrode under Reoxidation Based on the Real Microstructure.

During the operation of solid oxide fuel cells (SOFCs), the Ni-8YSZ anodes are subjected to thermal mismatch and reoxidation, accompanied by the risk of damage and failure. These damages and failures are generally induced by small defects at the microscopic level, leading to the degradation of the structural bearing capacity. Therefore, the distribution and quantification of the stresses in the real microstructure of Ni-8YSZ electrodes is essential. In this study, the real Ni-8YSZ microstructure was reconstructed based on nano-computed tomography, and the stress distribution of the real microstructure was analyzed based on the finite element method under reoxidation and different operating temperatures. The failure probability of 8YSZ at different degrees of reoxidation was evaluated according to the Weibull method, and the amount of damaged 8YSZ elements was statistically counted. The study results indicate a high level of stress in the thin necks and relatively sharp areas of the microstructure. The 8YSZ has a high failure probability at a reoxidation extent of 5-10%.

Geometric modelling of corrosion inhibitor pigments in active protective coatings based on SR-nano-CT images

This paper reports on an essential step towards accelerating the development of new, environmentally friendly active protective coatings by structure optimization. The complex microstructure of the pigment particles within a coating have been observed non-destructively in 3D by nano-computed tomography using synchrotron radiation. For the first time, a stochastic geometry model is fitted based on spatial geometric features of the particles observed in the 3D images. The typical cell of a random Gibbs-Laguerre tessellation is used to model the particles' polyhedral shapes as well as the observed joint size and aspect ratio distributions.

Utilizing nano-computed tomography to characterize the structural nature of industrial minerals

Abstract X-ray computed tomography (XRT) is a nondestructive and thus reproducible examination method allowing the three-dimensional (3D) investigation of the internal and external structures of objects regardless of their material and geometry. In the present study, XRT was used to investigate the influence of hydrochloric acid leaching on the composition and constitution of iron-rich bauxite grains. Reducing the iron content in raw bauxites by acid leaching is a promising method for the beneficiation of iron-rich bauxites for subsequent use in the refractory industry. Not only the effect of the leaching process on the chemical composition of the bauxites, but also aspects such as the acid’s influence on the mineralogical composition and the resulting porosity of the individual grains have hardly been taken into account so far. To address these issues, various bauxites were examined before and after acid leaching using XRT analysis and specifically characterized with regard to their constitution.

Enhancing PEMFC Performance: Optimization of Hot-Pressed Decal Transfer Conditions for MEA Fabrication

Proton exchange membrane fuel cells (PEMFCs) hold significant promise in curbing greenhouse gas emissions within the transportation sector. While historically more expensive than conventional internal combustion engines, the cost per kW is poised to significantly drop as production volumes increase and the adoption of fuel cell electric vehicles progresses [1]. In heavy-duty vehicle applications, predictions suggest that hydrogen fuel cells will achieve a cost equivalence with diesel, potentially becoming the most cost-effective option when evaluating the total ownership expenses [2]. Membrane-electrode-assemblies (MEAs) serve as the core of fuel cells, where all reactions—proton and electron generation, distribution, and consumption occur. Their efficiency dictates both cell performance and lifespan. Various methods for fabricating MEAs have been developed, including decal transfer, catalyst-coated membrane (CCM), and catalyst-coated gas diffusion layer (CCG) techniques. Among these, the decal transfer method is widely regarded as the most appropriate for large-scale production of MEAs [3]. This study delves into an examination of diverse hot-pressed decal transfer conditions for the preparation of MEAs tailored for PEMFC application. Our investigation involves fabricating MEAs through hot-press decal transfer at temperatures spanning 115°C to 145°C and pressures ranging from 200 psi to 400 psi, each held for a 5-minute duration. Electrochemical characterizations were performed within a single-cell setup to assess MEA performance. Moreover, an accelerated stress test (AST) was carried out to scrutinize the stability of the fabricated MEAs concerning their decal transfer conditions and morphological variations. Our study extensively explored catalyst layers transferred under diverse hot-pressed decal transfer conditions, focusing on their fuel cell performance and transport properties. To comprehensively understand the influence of morphological alterations on electrochemical characteristics, additional morphological characterizations, including white light interferometer (WLI) imaging, field emission scanning electron microscopy (FESEM), Brunauer-Emmett-Teller (BET) surface area analysis, and nano-computed tomography (Nano-CT) were employed for MEA evaluation. Our findings indicate that the MEA produced at 145°C and 200 psi exhibited superior performance in both pre- and post-AST testing. Comprehensive experimental results and insights will be elaborated upon during the conference presentation.

Using Micrometer Scale X-Ray CT and Pore Network Modeling to Assess High-Energy Bi-Layer Li-Ion Battery Cathode Structures

The optimization of cathode pore space is pivotal for enhancing the rate capability of lithium-ion battery electrodes. Previous studies on simulated cathodes have highlighted the importance of achieving an optimal porosity gradient, with a gradual increase from the current collector side to the separator side to accommodate the growing demand for ion transport in that direction [1]–[3]. In this research, we address this requirement by introducing and investigating bi-layer NCM 811 cathodes. These cathodes consist of a slurry-coated dense bottom layer and a spray-coated highly porous top layer with varying relative thickness to approximate the desired porosity gradient. Employing Micrometer-scale X-ray CT characterization and pore network modeling, we evaluate the structural features, demonstrating the effectiveness of the design, and quantify essential parameters such as porosity and tortuosity. Furthermore, the cathodes are integrated into lithium anode half-cells and subjected to electrochemical cycling at different rates to assess ion transport and rate capability. Our findings reveal that to achieve the highest discharge spatial energy density above 1C for cathodes approximately 100μm thick, the compressed bottom layer with 30% porosity should have a thickness of 60μm, while the highly porous top layer with around 47% porosity should be 40μm thick.Reference:[1] A. Shodiev et al., “Deconvoluting the benefits of porosity distribution in layered electrodes on the electrochemical performance of Li-ion batteries,” Energy Storage Mater., vol. 47, pp. 462–471, May 2022, doi: 10.1016/j.ensm.2022.01.058.[2] X. Lu et al., “3D microstructure design of lithium-ion battery electrodes assisted by X-ray nano-computed tomography and modelling,” Nat. Commun., vol. 11, no. 1, p. 2079, Apr. 2020, doi: 10.1038/s41467-020-15811-x.[3] K. Song, C. Zhang, N. Hu, X. Wu, and L. Zhang, “High performance thick cathodes enabled by gradient porosity,” Electrochimica Acta, vol. 377, p. 138105, May 2021, doi: 10.1016/j.electacta.2021.138105.

Quantitative Three-Dimensional Imaging Analysis of HfO2 Nanoparticles in Single Cells via Deep Learning Aided X-ray Nano-Computed Tomography.

It is crucial for understanding mechanisms of drug action to quantify the three-dimensional (3D) drug distribution within a single cell at nanoscale resolution. Yet it remains a great challenge due to limited lateral resolution, detection sensitivities, and reconstruction problems. The preferable method is using X-ray nano-computed tomography (Nano-CT) to observe and analyze drug distribution within cells, but it is time-consuming, requiring specialized expertise, and often subjective, particularly with ultrasmall metal nanoparticles (NPs). Furthermore, the accuracy of batch data analysis through conventional processing methods remains uncertain. In this study, we used radioenhancer ultrasmall HfO2 nanoparticles as a model to develop a modular and automated deep learning aided Nano-CT method for the localization quantitative analysis of ultrasmall metal NPs uptake in cancer cells. We have established an ultrasmall objects segmentation method for 3D Nano-CT images in single cells, which can highly sensitively analyze minute NPs and even ultrasmall NPs in single cells. We also constructed a localization quantitative analysis method, which may accurately segment the intracellularly bioavailable particles from those of the extracellular space and intracellular components and NPs. The high bioavailability of HfO2 NPs in tumor cells from deeper penetration in tumor tissue and higher tumor intracellular uptake provide mechanistic insight into HfO2 NPs as advanced radioenhancers in the combination of quantitative subcellular image analysis with the therapeutic effects of NPs on 3D tumor spheroids and breast cancer. Our findings unveil the substantial uptake rate and subcellular quantification of HfO2 NPs by the human breast cancer cell line (MCF-7). This revelation explicates the notable efficacy and safety profile of HfO2 NPs in tumor treatment. These findings demonstrate that this 3D imaging technique promoted by the deep learning algorithm has the potential to provide localization quantitative information about the 3D distributions of specific molecules at the nanoscale level. This study provides an approach for exploring the subcellular quantitative analysis of NPs in single cells, offering a valuable quantitative imaging tool for minute amounts or ultrasmall NPs.

3D quantification of the lacunocanalicular network on human femoral diaphysis through synchrotron radiation-based nanoCT

Osteocytes are the major actors in bone mechanobiology. Within bone matrix, they are trapped close together in a submicrometric interconnected network: the lacunocanalicular network (LCN). The interstitial fluid circulating within the LCN transmits the mechanical information to the osteocytes that convert it into a biochemical signal. Understanding the interstitial fluid dynamics is necessary to better understand the bone mechanobiology. Due to the submicrometric dimensions of the LCN, making it difficult to experimentally investigate fluid dynamics, numerical models appear as a relevant tool for such investigation. To develop such models, there is a need for geometrical and morphological data on the human LCN. This study aims at providing morphological data on the human LCN from measurement of 27 human femoral diaphysis bone samples using synchrotron radiation nano-computed tomography with an isotropic voxel size of 100 nm. Except from the canalicular diameter, the canalicular morphological parameters presented a high variability within one sample. Some differences in terms of both lacunar and canalicular morphology were observed between the male and female populations. But it has to be highlighted that all the canaliculi cannot be detected with a voxel size of 100 nm. Hence, in the current study, only a specific population of large canaliculi that could be characterize. Still, to the authors knowledge, this is the first time such a data set was introduced to the community. Further processing will be achieved in order to provide new insight on the LCN permeability.

An Electrolyte Engineered Homonuclear Copper Complex as Homogeneous Catalyst for Lithium-Sulfur Batteries.

Lithium-sulfur (Li-S) batteries suffer from severe polysulfide shuttle, retarded sulfur conversion kinetics and notorious lithium dendrites, which has curtailed the discharge capacity, cycling lifespan and safety. Engineered catalysts act as a feasible strategy to synchronously manipulate the evolution behaviors of sulfur and lithium species. Herein, a chlorine bridge-enabled binuclear copper complex (Cu-2-T) is in situ synthesized in electrolyte as homogeneous catalyst for rationalizing the Li-S redox reactions. The well-designed Cu-2-T provides completely active sites and sufficient contact for homogeneously guiding the Li2S nucleation/decomposition reactions, and stabilizing the lithium working interface according to the synchrotron radiation X-ray 3D nano-computed tomography, small angle neutron scattering and COMSOL results. Moreover, Cu-2-T with the content of 0.25wt% approaching saturated concentration in electrolyte further boosts the homogeneous optimization function in really operated Li-S batteries. Accordingly, the capacity retention of the Li-S battery is elevated from 51.4% to 86.3% at 0.2 C, and reaches 77.0% at 1.0 C over 400 cycles. Furthermore, the sulfur cathode with the assistance of Cu-2-T realizes the stable cycling under the practical scenarios of soft-packaged pouch cell and high sulfur loading (6.5mg cm-2 with the electrolyte usage of 4.5µL mgS -1).

Evaluation of Volumetric Changes between Pediatric Rotary Files and Manual Files during Canal Preparation of Primary Mandibular Canine: A Nano-CT Analysis

Abstract Objective Pediatric endodontics has become popular due to advancements in cleaning, shaping, and irrigation systems, resulting in faster and effective removal of infected pulp, saving time, and creating a pathogen-free environment. The patented rotary file system, Kedo S, designed for primary teeth, introduced a single file generation for efficient pulp therapy. However, there are currently no studies assessing canal preparation in primary mandibular canine using nano-computed tomography (nano-CT). This study aims to evaluate the efficacy of rotary file systems (Kedo S plus, Kedo SG blue) against traditional hand files in root canal preparation of primary mandibular canines using nano-CT. Materials and Methods This in vitro study was performed in extracted primary mandibular canine based on certain inclusion and exclusion criteria. Samples were prepared and working length was determined before the preoperative scan using a high-resolution nano-CT device (SkyScan 2214, Bruker, Kontich, Belgium). A single well-experienced pediatric dentist prepared the canals using three file systems: Kedo S plus, Kedo SG blue, and hand K-files. A postoperative scan was performed similar to preoperative scan. Image reconstruction was performed with NRecon software for three-dimensional volumetric visualization and analysis of the root canals. Results Kedo S plus displayed significant alterations in volumetric, surface area, and instrumented areas postpreparation compared with Kedo SG blue and hand files. Statistical analysis revealed significant differences in volumetric and surface changes between Kedo S plus and hand files. Conclusion Rotary file systems, especially Kedo S plus, resulted in increased canal volume and surface area with minimal uninstrumented areas, showing promise for primary dentition root canal preparations. Further clinical assessments are warranted to validate these findings.

The Geometric Correction Method for zircon (U–Th) ∕ He chronology: correcting systematic error and assigning uncertainties to alpha-ejection corrections and eU concentrations

Abstract. The conventional zircon (U–Th) / He (ZHe) method typically uses microscopy measurements of the dated grain together with the assumption that the zircon can be appropriately modeled as a geometrically perfect tetragonal or ellipsoidal prism in the calculation of volume (V), alpha-ejection correction (FT), equivalent spherical radius (RFT), effective uranium concentration (eU), and corrected (U–Th) / He date. Here, we develop a set of corrections for systematic error and determine uncertainties to be used in the calculation of the above parameters for zircon, using the same methodology as Zeigler et al. (2023) for apatite. Our approach involved acquiring both “2D” microscopy measurements and high-resolution “3D” nano-computed tomography (CT) data for a suite of 223 zircon grains from nine samples showcasing a wide range of morphology, size, age, and lithological source, calculating the V, FT, and RFT values for the 2D and 3D measurements and comparing the 2D vs. 3D results. We find that the values derived from the 2D microscopy data overestimate the true 3D V, FT, and RFT values for zircon, with one exception (V of ellipsoidal grains). Correction factors for this misestimation determined by regressing the 3D vs. 2D data range from 0.81–1.04 for V, 0.97–1.0 for FT, and 0.92–0.98 for RFT, depending on zircon geometry. Uncertainties (1σ) derived from the scatter of data around the regression line are 13 %–21 % for V, 5 %–1 % for FT, and 8 % for RFT, again depending on zircon morphologies. Like for apatite, the main control on the magnitude of the corrections and uncertainties is grain geometry, with grain size being a secondary control on FT uncertainty. Propagating these uncertainties into a real dataset (N=28 ZHe analyses) generates 1σ uncertainties of 12 %–21 % in eU and 3 %–7 % in the corrected ZHe date when both analytical and geometric uncertainties are included. Accounting for the geometric corrections and uncertainties is important for appropriately reporting, plotting, and interpreting ZHe data. For both zircon and apatite, the Geometric Correction Method is a practical and straightforward approach for calculating more accurate (U–Th) / He data and for including geometric uncertainty in eU and date uncertainties.

Nano-computed tomography analysis of downscaled cement sheath samples: Insights into porosity distribution

Mitigating greenhouse gas emissions from hydrocarbon wells is an industrywide challenge, with microstructural variations in the cement sheaths lining the wells recognized as key factors in forming leakage pathways. To better understand the microstructural variations in cement sheaths and the corresponding mechanisms, downscaled samples simulating rock–cement–casing and casing–cement–casing cement sheaths were prepared and then scanned using nano-computed tomography at resolutions of 11.93 μm and 6.80 μm, respectively. Both the radial and axial porosity of regions of interest were quantified and analyzed. The results showed that radial porosity distributions were quite different between the rock–cement–casing and casing–cement–casing samples. In the rock–cement–casing samples, the radial porosity decreased from the cement–rock to the cement–casing interface due to cement migration, while in the casing–cement–casing samples, the highest radial porosities occurred in both the cement–casing interfaces due to the influence of an interfacial transition zone. The axial porosity distribution was similar in all the rock–cement–casing and casing–cement–casing samples because the porosity decreased from the top to the bottom due to the particle settling phenomenon. These findings enhance our understanding of the factors affecting gas leakage pathways from wells, and the proposed methodology can be used to optimize cement blends to produce effective wellbore barriers.

Comparative evaluations on the canal-centering ability of novel heat-treated retreatment file system using nano-computed tomography

Abstract Introduction: The main goal of instrumentation during retreatment is to maintain the original curvature of the canal during retrieval of gutta-percha. The present study evaluates the canal-centering ability of two different retreatment file systems. Materials and Methods: Forty extracted single-rooted teeth were decoronated and obturated at the standardized root length of 16 mm. Samples were divided into two groups of each 20 single root teeth and they were instrumented using Solite RS3 Retreatment and ProTaper Retreatment files, respectively. Both pre- and postoperative nano-computed tomography scans were taken, and three-dimensional reconstruction was done for analysis. Sections of both the pre- and postoperative samples were taken at 7 mm, 5 mm, and 3 mm from the apex to calculate the canal-centering ratio. Independent t-test was used for statistical analysis between the two groups. Results: Solite RS3 retreatment files showed a significant difference at the 3 levels (P < 0.000) compared to ProTaper retreatment files. Conclusion: From the above results, we can conclude that Solite RS3 caused minimal canal transportation and produced a relatively centered preparation in comparison to the ProTaper Retreatment system.

Porous Ceramic Metal-Based Flow Battery Composite Membrane.

In metal-based flow battery, membranes significantly impact energy conversion efficiency and security. Unfortunately, damages to the membrane occur due to gradual accumulation of metal dendrites, causing short circuits and shortening cycle life. Herein, we developed a rigid hierarchical porous ceramic flow battery composite membrane with a sub-10-nm-thick polyelectrolyte coating to achieve high ion selectivity and conductivity, to restrain dendrite, and to realize long cycle life and high areal capacity. An aqueous zinc-iron flow battery prepared using this membrane achieved an outstanding energy efficiency of >80%, exhibiting excellent long-term stability (over 1000 h) and extremely high areal capacity (260 mAh cm-2). Low-field nuclear magnetic resonance (NMR) spectroscopy, small-angle X-ray scattering, in situ infrared spectroscopy, solid-state NMR analysis, and nano-computed tomography revealed that the rigid hierarchical pore structures and numerous hydrogen bonding networks in the membrane contributed to the stable operation and superior battery performance. This study contributes to the development of next-generation metal-based flow battery membranes for energy and power generation.

Parasite escape mechanisms drive morphological diversification in avian lice.

Organisms that have repeatedly evolved similar morphologies owing to the same selective pressures provide excellent cases in which to examine specific morphological changes and their relevance to the ecology and evolution of taxa. Hosts of permanent parasites act as an independent evolutionary experiment, as parasites on these hosts are thought to be undergoing similar selective pressures. Parasitic feather lice have repeatedly diversified into convergent ecomorphs in different microhabitats on their avian hosts. We quantified specific morphological characters to determine (i) which traits are associated with each ecomorph, (ii) the quantitative differences between these ecomorphs, and (iii) if there is evidence of displacement among co-occurring lice as might be expected under louse-louse competition on the host. We used nano-computed tomography scan data of 89 specimens, belonging to four repeatedly evolved ecomorphs, to examine their mandibular muscle volume, limb length and three-dimensional head shape data. Here, we find evidence that lice repeatedly evolve similar morphologies as a mechanism to escape host defences, but also diverge into different ecomorphs related to the way they escape these defences. Lice that co-occur with other genera on a host exhibit greater morphological divergence, indicating a potential role of competition in evolutionary divergence.

Impact of crystalline domains on long-term stability and mechanical performance of anisotropic silk fibroin sponges.

Sponge-like materials made from regenerated silk fibroin biopolymers are a tunable and advantageous platform for in vitro engineered tissue culture and in vivo tissue regeneration. Anisotropic, three-dimensional (3D) silk fibroin sponge-like scaffolds can mimic the architecture of contractile muscle. Herein, we use silk fibroin solution isolated from the cocoons of Bombyx mori silkworms to form aligned sponges via directional ice templating in a custom mold with a slurry of dry ice and ethanol. Hydrated tensile mechanical properties of these aligned sponges were evaluated as a function of silk polymer concentration (3% or 5%), freezing time (50% or 100% ethanol), and post-lyophilization method for inducing crystallinity (autoclaving, water annealing). Hydrated static tensile tests were used to determine Young's modulus and ultimate tensile strength across sponge formulations at two strain rates to evaluate rate dependence in the calculated parameters. Results aligned with previous reports in the literature for isotropic silk fibroin sponge-like scaffolds, where the method by which beta-sheets were formed and level of beta-sheet content (crystallinity) had the greatest impact on static parameters, while polymer concentration and freezing rate did not significantly impact static mechanical properties. We estimated the crystalline organization using molecular dynamics simulations to show that larger crystalline regions may be responsible for strength at low strain amplitudes and brittleness at high strain amplitudes in the autoclaved sponges. Within the parameters evaluated, extensional Young's modulus is tunable in the range of 600-2800 kPa. Dynamic tensile testing revealed the linear viscoelastic region to be between 0% and 10% strain amplitude and 0.2-2 Hz frequencies. Long-term stability was evaluated by hysteresis and fatigue tests. Fatigue tests showed minimal change in the storage and loss modulus of 5% silk fibroin sponges for more than 6000 min of continuous mechanical stimulation in the linear regime at 10% strain amplitude and 1 Hz frequency. Furthermore, we confirmed that these mechanical properties hold when decellularized extracellular matrix is added to the sponges and when the mechanical property assessments were performed in cell culture media. We also used nano-computed tomography (nano-CT) and simulations to explore pore interconnectivity and tortuosity. Overall, these results highlight the potential of anisotropic, sponge-like silk fibroin scaffolds for long-term (>6 weeks) contractile muscle culture with an in vitro bioreactor system that provides routine mechanical stimulation.

Comparative evaluation of volumetric changes following rotary and hand files' canal preparation of primary maxillary canine: an in vitro nano-CT analysis.

To evaluate the volumetric changes of two recently introduced paediatric rotary file systems in comparison with conventional hand file systems in primary maxillary canines using an ultra-high-resolution nano-computed tomography. This in vitro study was performed in extracted primary maxillary canines based on certain inclusion and exclusion criteria. Samples were prepared, and working length was determined after the pre-operative scan using a high-resolution nano-CT device (SkyScan 2214, Bruker, Kontich, Belgium). A single well-experienced paediatric dentist prepared the canals using three file systems: Kedo-S plus, Kedo-SG blue and hand K-files. All samples were subjected to post-operative scans performed similar to pre-operative scans. Image reconstruction was performed with NRecon software for 3D volumetric visualisation and analysis of the root canals. Kedo-SG blue file systems had the highest mean difference in the canal volume (4.05%). Hand K-files had the least difference at (3.71%) of canal volume. Kedo-S plus file system had a moderate mean canal volume difference (3.82%) which is closer to hand K-files. Intergroup comparison between the three groups showed that the mean difference in canal volume was statistically significant between all three file systems (p = 0.000). Within the limitations of the current study, rotary file systems produced a significant enlargement of canals as compared to hand files. Kedo-SG blue created a uniform preparation of the canal cervico-apically. Kedo-S plus files were prepared more coronally with minimal preparation apically as close to the preparation of hand files. Trial registration number: IHEC/SDC/PEDO-2103/22/651, Date of registration: 2022.

An enteric ultrastructural surface atlas of the model insect Manduca sexta

The tobacco hornworm is a laboratory model that is particularly suitable for analyzing gut inflammation, but a physiological reference standard is currently unavailable. Here, we present a surface atlas of the healthy hornworm gut generated by scanning electron microscopy and nano-computed tomography. This comprehensive overview of the gut surface reveals morphological differences between the anterior, middle, and posterior midgut, allowing the screening of aberrant gut phenotypes while accommodating normal physiological variations. We estimated a total resorptive midgut surface of 0.42 m2 for L5d6 larvae, revealing its remarkable size. Our data will support allometric scaling and dose conversion from Manduca sexta to mammals in preclinical research, embracing the 3R principles. We also observed non-uniform gut colonization by enterococci, characterized by dense biofilms in the pyloric cone and downstream of the pylorus associated with pore and spine structures in the hindgut intima, indicating a putative immunosurveillance function in the lepidopteran hindgut.