Capillarity and active cell movement at mesendoderm translocation in the Xenopus gastrula.

During Xenopus gastrulation, leading edge mesendoderm (LEM) advances animally as a wedge-shaped cell mass over the vegetally moving blastocoel roof (BCR). We show that close contact across the BCR-LEM interface correlates with attenuated net advance of the LEM, which is pulled forward by tip cells while the remaining LEM frequently separates from the BCR. Nevertheless, lamellipodia persist on the detached LEM surface. They attach to adjacent LEM cells and depend on PDGF-A, cell-surface fibronectin and cadherin. We argue that active cell motility on the LEM surface prevents adverse capillary effects in the liquid LEM tissue as it moves by being pulled. It counters tissue surface-tension effects with oriented cell movement and bulges the LEM surface out to keep it close to the curved BCR without attaching to it. Proximity to the BCR is necessary, in turn, for the maintenance and orientation of lamellipodia that permit mass cell movement with minimal substratum contact. Together with a similar process in epithelial invagination, vertical telescoping, the cell movement at the LEM surface defines a novel type of cell rearrangement: vertical shearing.

Drought-Induced Xylem Embolism Limits the Recovery of Leaf Gas Exchange in Scots Pine.

Climate change increases the occurrence of prolonged drought periods with large implications for forest functioning. Scots pine (Pinus sylvestris) is one of the most abundant conifers worldwide, and evidence is rising that its resilience to severe drought is limited. However, we know little about its ability to recover from drought-induced embolism. To analyze postdrought hydraulic recovery, we investigated stress and recovery dynamics of leaf gas exchange, nonstructural carbohydrates, and hydraulic properties in 2.5-year-old Scots pine seedlings. We quantified the degree of xylem embolism by combining in vivo x-ray microtomography with intrusive techniques including measurements of hydraulic conductivity and dye staining during drought progression and short-term (2 d) and long-term (4 weeks) recovery. Seedlings were grown under controlled conditions, and irrigation was withheld until stomata closed and xylem water potential declined to -3.2 MPa on average, causing a 46% loss of stem hydraulic conductivity. Following drought release, we found a gradual recovery of leaf gas exchange to 50% to 60% of control values. This partial recovery indicates hydraulic limitations due to drought-induced damage. Whereas xylem water potential recovered close to control values within 2 d, both x-ray microtomography and intrusive measurements revealed no recovery of stem hydraulic conductivity. Moreover, we did not find indications for nonstructural carbohydrate reserves limiting hydraulic recovery. Our findings demonstrate that Scots pine is able to survive severe drought and to partially recover, although we assume that xylem development during the next growing season might compensate for some of the hydraulic impairment. Such drought-induced legacy effects are important when considering vegetation responses to extreme events.

Novel multicomponent organic-inorganic WPI/gelatin/CaP hydrogel composites for bone tissue engineering.

The present work focuses on the development of novel multicomponent organic-inorganic hydrogel composites for bone tissue engineering. For the first time, combination of the organic components commonly used in food industry, namely whey protein isolate (WPI) and gelatin from bovine skin, as well as inorganic material commonly used as a major component of hydraulic bone cements, namely α-TCP in various concentrations (0-70 wt%) was proposed. The results showed that α-TCP underwent incomplete transformation to calcium-deficient hydroxyapatite (CDHA) during preparation process of the hydrogels. Microcomputer tomography showed inhomogeneous distribution of the calcium phosphate (CaP) phase in the resulting composites. Nevertheless, hydrogels containing 30-70 wt% α-TCP showed significantly improved mechanical properties. The values of Young's modulus and the stresses corresponding to compression of a sample by 50% increased almost linearly with increasing concentration of ceramic phase. Incomplete transformation of α-TCP to CDHA during preparation process of composites provides them high reactivity in simulated body fluid during 14-day incubation. Preliminary in vitro studies revealed that the WPI/gelatin/CaP composite hydrogels support the adhesion, spreading, and proliferation of human osteoblast-like MG-63 cells. The WPI/gelatin/CaP composite hydrogels obtained in this work showed great potential for the use in bone tissue engineering and regenerative medicine applications.

Zooming X-rays with a single rotation in X-ray prism zoom lenses (XPZL).

Prism arrays arranged to form a slightly open alligator mouth were found to focus incident X-rays, as with increasing distance from the object symmetry axis these rays hit an increasing number of refracting prism tips. Such an object is then formally a refractive lens. Due to the strong energy dependence of the refractive index of material for X-rays a refractive X-ray lens is chromatically focusing. The attractive feature of the alligator lens is the inherent zoomability possible as the mouth can easily be opened or closed. However, the required tolerances for the jaw rotations and the jaw positioning are so stringent, that the routine use of such systems has not been reported yet. This study will show that the related technical problems can be overcome by proper object fabrication. In fact the here presented objects can already be aligned in the production stage. Then the assembly can be made with simple tools. And the zooming is achieved by just a simple rotation. The transmission through the devices was found to be as expected, and thus performance-wise these objects can directly compete with other refractive X-ray focusing systems.

Quanfima: An open source Python package for automated fiber analysis of biomaterials.

Hybrid 3D scaffolds composed of different biomaterials with fibrous structure or enriched with different inclusions (i.e., nano- and microparticles) have already demonstrated their positive effect on cell integration and regeneration. The analysis of fibers in hybrid biomaterials, especially in a 3D space is often difficult due to their various diameters (from micro to nanoscale) and compositions. Though biomaterials processing workflows are implemented, there are no software tools for fiber analysis that can be easily integrated into such workflows. Due to the demand for reproducible science with Jupyter notebooks and the broad use of the Python programming language, we have developed the new Python package quanfima offering a complete analysis of hybrid biomaterials, that include the determination of fiber orientation, fiber and/or particle diameter and porosity. Here, we evaluate the provided tensor-based approach on a range of generated datasets under various noise conditions. Also, we show its application to the X-ray tomography datasets of polycaprolactone fibrous scaffolds pure and containing silicate-substituted hydroxyapatite microparticles, hydrogels enriched with bioglass contained strontium and alpha-tricalcium phosphate microparticles for bone tissue engineering and porous cryogel 3D scaffold for pancreatic cell culturing. The results obtained with the help of the developed package demonstrated high accuracy and performance of orientation, fibers and microparticles diameter and porosity analysis.

Effect of low-temperature plasma treatment of electrospun polycaprolactone fibrous scaffolds on calcium carbonate mineralisation.

This article reports on a study of the mineralisation behaviour of CaCO3 deposited on electrospun poly(ε-caprolactone) (PCL) scaffolds preliminarily treated with low-temperature plasma. This work was aimed at developing an approach that improves the wettability and permeability of PCL scaffolds in order to obtain a superior composite coated with highly porous CaCO3, which is a prerequisite for biomedical scaffolds used for drug delivery. Since PCL is a synthetic polymer that lacks functional groups, plasma processing of PCL scaffolds in O2, NH3, and Ar atmospheres enables introduction of highly reactive chemical groups, which influence the interaction between organic and inorganic phases and govern the nucleation, crystal growth, particle morphology, and phase composition of the CaCO3 coating. Our studies showed that the plasma treatment induced the formation of O- and N-containing polar functional groups on the scaffold surface, which caused an increase in the PCL surface hydrophilicity. Mineralisation of the PCL scaffolds was performed by inducing precipitation of CaCO3 particles on the surface of polymer fibres from a mixture of CaCl2- and Na2CO3-saturated solutions. The presence of highly porous vaterite and nonporous calcite crystal phases in the obtained coating was established. Our findings confirmed that preferential growth of the vaterite phase occurred in the O2-plasma-treated PCL scaffold and that the coating formed on this scaffold was smoother and more homogenous than those formed on the untreated PCL scaffold and the Ar- and NH3-plasma-treated PCL scaffolds. A more detailed three-dimensional assessment of the penetration depth of CaCO3 into the PCL scaffold was performed by high-resolution micro-computed tomography. The assessment revealed that O2-plasma treatment of the PCL scaffold caused CaCO3 to nucleate and precipitate much deeper inside the porous structure. From our findings, we conclude that O2-plasma treatment is preferable for PCL scaffold surface modification from the viewpoint of use of the PCL/CaCO3 composite as a drug delivery platform for tissue engineering.

Silk scaffolds connected with different naturally occurring biomaterials for prostate cancer cell cultivation in 3D.

In the present work, different biopolymer blend scaffolds based on the silk protein fibroin from Bombyx mori (BM) were prepared via freeze-drying method. The chemical, structural, and mechanical properties of the three dimensional (3D) porous silk fibroin (SF) composite scaffolds of gelatin, collagen, and chitosan as well as SF from Antheraea pernyi (AP) and the recombinant spider silk protein spidroin (SSP1) have been systematically investigated, followed by cell culture experiments with epithelial prostate cancer cells (LNCaP) up to 14 days. Compared to the pure SF scaffold of BM, the blend scaffolds differ in porous morphology, elasticity, swelling behavior, and biochemical composition. The new composite scaffold with SSP1 showed an increased swelling degree and soft tissue like elastic properties. Whereas, in vitro cultivation of LNCaP cells demonstrated an increased growth behavior and spheroid formation within chitosan blended scaffolds based on its remarkable porosity, which supports nutrient supply matrix. Results of this study suggest that silk fibroin matrices are sufficient and certain SF composite scaffolds even improve 3D cell cultivation for prostate cancer research compared to matrices based on pure biomaterials or synthetic polymers.

Single-distance phase retrieval algorithm for Bragg Magnifier microscope.

We present an improved, single-distance phase retrieval algorithm applicable for holographic X-ray imaging of biological objects for an in-line germanium Bragg Magnifier Microscope (BMM). The proposed algorithm takes advantage of a modified shrink-wrap algorithm for phase objects, robust unwrapping algorithm as well as other reasonable constraints applied to the wavefield at the object and the detector plane. The performance of the algorithm is analyzed on phantom objects and the results are shown and discussed. We demonstrated the suitability of the algorithm for the phase retrieval on a more complex biological specimen Tardigrade, where we achieved successful phase retrieval from only a single hologram. The spatial resolution obtained by Fourier spectral power method for biological objects is ∼ 300 nm, the same value as obtained from the reconstructed test pattern. Our results achieved using the new algorithm confirmed the potential of BMM for in-vivo, dose-efficient single-shot imaging of biological objects.

Superporous Poly(ethylene glycol) Diacrylate Cryogel with a Defined Elastic Modulus for Prostate Cancer Cell Research.

The physical and mechanical properties of the tumor microenvironment are crucial for the growth, differentiation and migration of cancer cells. However, such microenvironment is not found in the geometric constraints of 2D cell culture systems used in many cancer studies. Prostate cancer research, in particular, suffers from the lack of suitable in vitro models. Here a 3D superporous scaffold is described with thick pore walls in a mechanically stable and robust architecture to support prostate tumor growth. This scaffold is generated from the cryogelation of poly(ethylene glycol) diacrylate to produce a defined elastic modulus for prostate tumor growth. Lymph node carcinoma of the prostate (LNCaP) cells show a linear growth over 21 d as multicellular tumor spheroids in such a scaffold with points of attachments to the walls of the scaffold. These LNCaP cells respond to the growth promoting effects of androgens and demonstrate a characteristic cytoplasmic-nuclear translocation of the androgen receptor and androgen-dependent gene expression. Compared to 2D cell culture, the expression or androgen response of prostate cancer specific genes is greatly enhanced in the LNCaP cells in this system. This scaffold is therefore a powerful tool for prostate cancer studies with unique advantages over 2D cell culture systems.

Comparative thorax morphology of death-feigning flightless cryptorhynchine weevils (Coleoptera: Curculionidae) based on 3D reconstructions.

The thorax morphology, especially the muscles and the tracheal system of three flightless species of Cryptorhynchinae is examined by digital 3D reconstructions based on synchrotron X-ray microtomography and compared to other Curculionidae. Wings, metanepisternites, and muscles functional in flight are fully reduced in the species examined: Kyklioacalles roboris (Curtis), Trigonopterus scharfi Riedel and Trigonopterus vandekampi Riedel. All three share the same set of thoracic muscles, but differences exist in the shape and size of muscles. Both Trigonopterus species examined have a conspicuous fan-shaped branch of Musculus mesosterni primus contracting pro- and mesothorax, interpreted as an adaption to their thanatosis defense strategy. Trigonopterus vandekampi furthermore shows a marked increase in the size of two metacoxal muscles, which may be functional in this species' thanatosis blocking mechanisms. The metathoracic spiracle of all Trigonopterus species is located at the side of the metaventrite externally and not in the subelytral space as in other beetles. It is hypothesized that this translocation was triggered by the need to improve oxygen supply during thanatosis, when both the mesothoracic spiracle and the subelytral cavity are tightly sealed from the outside.

Performance of a Medipix3RX spectroscopic pixel detector with a high resistivity gallium arsenide sensor.

High resistivity gallium arsenide is considered a suitable sensor material for spectroscopic X-ray imaging detectors. These sensors typically have thicknesses between a few hundred µm and 1 mm to ensure a high photon detection efficiency. However, for small pixel sizes down to several tens of µm, an effect called charge sharing reduces a detector's spectroscopic performance. The recently developed Medipix3RX readout chip overcomes this limitation by implementing a charge summing circuit, which allows the reconstruction of the full energy information of a photon interaction in a single pixel. In this work, we present the characterization of the first Medipix3RX detector assembly with a 500 µm thick high resistivity, chromium compensated gallium arsenide sensor. We analyze its properties and demonstrate the functionality of the charge summing mode by means of energy response functions recorded at a synchrotron. Furthermore, the imaging properties of the detector, in terms of its modulation transfer functions and signal-to-noise ratios, are investigated. After more than one decade of attempts to establish gallium arsenide as a sensor material for photon counting detectors, our results represent a breakthrough in obtaining detector-grade material. The sensor we introduce is therefore suitable for high resolution X-ray imaging applications.

How spectroscopic x-ray imaging benefits from inter-pixel communication.

Spectroscopic x-ray imaging based on pixellated semiconductor detectors can be sensitive to charge sharing and K-fluorescence, depending on the sensor material used, its thickness and the pixel pitch employed. As a consequence, spectroscopic resolution is partially lost. In this paper, we study a new detector ASIC, the Medipix3RX, that offers a novel feature called charge summing, which is established by making adjacent pixels communicate with each other. Consequently, single photon interactions resulting in multiple hits are almost completely avoided. We investigate this charge summing mode with respect to those of its imaging properties that are of interest in medical physics and benchmark them against the case without charge summing. In particular, we review its influence on spectroscopic resolution and find that the low energy bias normally present when recording energy spectra is dramatically reduced. Furthermore, we show that charge summing provides a modulation transfer function which is almost independent of the energy threshold setting, which is in contrast to approaches common so far. We demonstrate that this property is directly linked to the detective quantum efficiency, which is found to increase by a factor of three or more when the energy threshold approaches the photon energy and when using charge summing. As a consequence, the contrast-to-noise ratio is found to double at elevated threshold levels and the dynamic range increases for a given counter depth. All these effects are shown to lead to an improved ability to perform material discrimination in spectroscopic CT, using iodine and gadolinium contrast agents. Hence, when compared to conventional photon counting detectors, these benefits carry the potential of substantially reducing the imaging dose a patient is exposed to during diagnostic CT examinations.

High-resolution high-efficiency X-ray imaging system based on the in-line Bragg magnifier and the Medipix detector.

The performance of a recently developed full-field X-ray micro-imaging system based on an in-line Bragg magnifier is reported. The system is composed of quasi-channel-cut crystals in combination with a Medipix single-photon-counting detector. A theoretical and experimental study of the imaging performance of the crystals-detector combination and a comparison with a standard indirect detector typically used in high-resolution X-ray imaging schemes are reported. The spatial resolution attained by our system is about 0.75 µm, limited only by the current magnification. Compared with an indirect detector system, this system features a better efficiency, signal-to-noise ratio and spatial resolution. The optimal working resolution range of this system is between ∼0.4 µm and 1 µm, filling the gap between transmission X-ray microscopes and indirect detectors. Applications for coherent full-field imaging of weakly absorbing samples are shown and discussed.

Imaging properties of small-pixel spectroscopic x-ray detectors based on cadmium telluride sensors.

Spectroscopic x-ray imaging by means of photon counting detectors has received growing interest during the past years. Critical to the image quality of such devices is their pixel pitch and the sensor material employed. This paper describes the imaging properties of Medipix2 MXR multi-chip assemblies bump bonded to 1 mm thick CdTe sensors. Two systems were investigated with pixel pitches of 110 and 165 µm, which are in the order of the mean free path lengths of the characteristic x-rays produced in their sensors. Peak widths were found to be almost constant across the energy range of 10 to 60 keV, with values of 2.3 and 2.2 keV (FWHM) for the two pixel pitches. The average number of pixels responding to a single incoming photon are about 1.85 and 1.45 at 60 keV, amounting to detective quantum efficiencies of 0.77 and 0.84 at a spatial frequency of zero. Energy selective CT acquisitions are presented, and the two pixel pitches' abilities to discriminate between iodine and gadolinium contrast agents are examined. It is shown that the choice of the pixel pitch translates into a minimum contrast agent concentration for which material discrimination is still possible. We finally investigate saturation effects at high x-ray fluxes and conclude with the finding that higher maximum count rates come at the cost of a reduced energy resolution.

In-line Bragg magnifier based on V-shaped germanium crystals.

In this work an X-ray imaging system based on a recently developed in-line two-dimensional Bragg magnifier composed of two monolithic V-shaped crystals made of dislocation-free germanium is presented. The channel-cut crystals were used in one-dimensional and in two-dimensional (crossed) configurations in imaging applications and allowed measurement of phase-contrast radiograms both in the edge-enhanced and in the holographic regimes. The measurement of the phase gradient in two orthogonal directions is demonstrated. The effective pixel size attained was 0.17 µm in the one-dimensional configuration and 0.5 µm in the two-dimensional setting, offering a twofold improvement in spatial resolution over devices based on silicon. These results show the potential for applying Bragg magnifiers to imaging soft matter at high resolution with reduced dose owing to the higher efficiency of Ge compared with Si.

A novel epitaxially grown LSO-based thin-film scintillator for micro-imaging using hard synchrotron radiation.

The efficiency of high-resolution pixel detectors for hard X-rays is nowadays one of the major criteria which drives the feasibility of imaging experiments and in general the performance of an experimental station for synchrotron-based microtomography and radiography. Here the luminescent screen used for the indirect detection is focused on in order to increase the detective quantum efficiency: a novel scintillator based on doped Lu(2)SiO(5) (LSO), epitaxially grown as thin film via the liquid phase epitaxy technique. It is shown that, by using adapted growth and doping parameters as well as a dedicated substrate, the scintillation behaviour of a LSO-based thin crystal together with the high stopping power of the material allows for high-performance indirect X-ray detection. In detail, the conversion efficiency, the radioluminescence spectra, the optical absorption spectra under UV/visible-light and the afterglow are investigated. A set-up to study the effect of the thin-film scintillator's temperature on its conversion efficiency is described as well. It delivers knowledge which is important when working with higher photon flux densities and the corresponding high heat load on the material. Additionally, X-ray imaging systems based on different diffraction-limited visible-light optics and CCD cameras using among others LSO-based thin film are compared. Finally, the performance of the LSO thin film is illustrated by imaging a honey bee leg, demonstrating the value of efficient high-resolution computed tomography for life sciences.