3D Microstructure Effects in Ni-YSZ Anodes: Prediction of Effective Transport Properties and Optimization of Redox Stability.

This study investigates the influence of microstructure on the effective ionic and electrical conductivities of Ni-YSZ (yttria-stabilized zirconia) anodes. Fine, medium, and coarse microstructures are exposed to redox cycling at 950 °C. FIB (focused ion beam)-tomography and image analysis are used to quantify the effective (connected) volume fraction (Φeff), constriction factor (ß), and tortuosity (τ). The effective conductivity (σeff) is described as the product of intrinsic conductivity (σ0) and the so-called microstructure-factor (M): σeff= σ0*M. Two different methods are used to evaluate the M-factor: (1) by prediction using a recently established relationship, Mpred= ε߰.36/τ5.17, and (2) by numerical simulation that provides conductivity, from which the simulated M-factor can be deduced (Msim). Both methods give complementary and consistent information about the effective transport properties and the redox degradation mechanism. The initial microstructure has a strong influence on effective conductivities and their degradation. Finer anodes have higher initial conductivities but undergo more intensive Ni coarsening. Coarser anodes have a more stable Ni phase but exhibit lower YSZ stability due to lower sintering activity. Consequently, in order to improve redox stability, it is proposed to use mixtures of fine and coarse powders in different proportions for functional anode and current collector layers.

Correlative 3D imaging: CLSM and FIB-SEM tomography using high-pressure frozen, freeze-substituted biological samples.

Correlative light and electron microscopy aims at combining data from different imaging modalities, ideally from the same area of the one sample, in order to achieve a more holistic view of the hierarchical structural organization of cells and tissues. Modern 3D imaging techniques opened up new possibilities to expand morphological studies into the third dimension at the nanometer scale. Here we present an approach to correlate 3D light microscopy data with volume data from focused ion beam-scanning electron microscopy. An adapted sample preparation method based on high-pressure freezing for structure preservation, followed by freeze-substitution for multimodal en bloc imaging, is described. It is based on including fluorescent labeling during freeze-substitution, which enables histological context description of the structure of interest by confocal laser scanning microscopy prior to high-resolution electron microscopy. This information can be employed to relocate the respective structure in the electron microscope. This approach is most suitable for targeted small 3D volume correlation and has the potential to extract statistically relevant data of structural details for systems biology.

Bridging microscopes: 3D correlative light and scanning electron microscopy of complex biological structures.

The rationale of correlative light and electron microscopy (CLEM) is to collect data on different information levels--ideally from an identical area on the same sample--with the aim of combining datasets at different levels of resolution to achieve a more holistic view of the hierarchical structural organization of cells and tissues. Modern three-dimensional (3D) imaging techniques in light and electron microscopy opened up new possibilities to expand morphological studies into the third dimension at the nanometer scale and over various volume dimensions. Here, we present two alternative approaches to correlate 3D light microscopy (LM) data with scanning electron microscopy (SEM) volume data. An adapted sample preparation method based on high-pressure freezing for structure preservation, followed by freeze-substitution for multimodal en-bloc imaging or serial-section imaging is described. The advantages and potential applications are exemplarily shown on various biological samples, such as cells, individual organisms, human tissue, as well as plant tissue. The two CLEM approaches presented here are per se not mutually exclusive, but have their distinct advantages. Confocal laser scanning microscopy (CLSM) and focused ion beam-SEM (FIB-SEM) is most suitable for targeted 3D correlation of small volumes, whereas serial-section LM and SEM imaging has its strength in large-area or -volume screening and correlation. The second method can be combined with immunocytochemical methods. Both methods, however, have the potential to extract statistically relevant data of structural details for systems biology.

Part 2. Comparison of emergency washing solutions in 70% hydrofluoric acid-burned human skin in an established ex vivo explants model.

BACKGROUND: Hydrofluoric acid (HF) is a small and partially dissociated acid (pK(a) 3.2), able to deeply penetrate into human skin in addition to the corrosiveness of the hydrogen ion (H(+)) and the toxicity of the fluoride ion (F(-)). However, there has been a lack of experimental studies to objectively characterize the results of human HF skin exposure decontamination. METHODOLOGY/PRINCIPAL FINDINGS: A previously established experimental method using a human skin explants ex vivo model (Part 1. Experimental 70% hydrofluoric acid (HF) burns: Histological observations in an established human skin explants ex vivo model) described the lesions that appeared following 70% HF penetration. Within 5 min, 70% HF penetrates to the dermis. Using the same experimental conditions, a comparison study of two different washing protocols was performed: water + topical calcium gluconate (CaG) versus Hexafluorine(®). In these conditions, washing for 15 min with running tap water followed by topical CaG ointment only delayed burn onset, while severe tissue damage appeared later. In contrast, after washing with Hexafluorine(®) over 10 min, no histological lesions developed. These results are in accordance with the results of accidental human industrial case reports. CONCLUSION/SIGNIFICANCE: Amphoteric and hypertonic Hexafluorine(®) can deactivate H(+) and chelate F(-) ions. Based on these results, it should be considered as a promising first-aid decontamination solution to prevent or minimize significant local and systemic consequences of concentrated HF skin exposures.

Experimental 70% hydrofluoric acid burns: histological observations in an established human skin explants ex vivo model.

BACKGROUND: Hydrofluoric acid (HF) is particularly dangerous due to the potential for systemic effects and induction of severe skin necrosis through two mechanisms: corrosiveness and local tissue toxicity. In addition, because it is only partially dissociated (pK(a) 3.2), it is capable of penetrating deeply into tissues. There is a lack of experimental studies that objectively characterize the behavior of HF diffusion into human skin, specifically the kinetics of tissue penetration resulting in severe cellular lesions. METHODOLOGY/PRINCIPAL FINDINGS: We describe the cutaneous effects of HF using an established ex vivo human skin model. The diffusion of 70% HF starts within the first minute of contact at the epidermal surface and after 2 min reaches the basal layer. In the subsequent minute, the epidermis is destroyed and lesions appear in the papillary dermis after 4 min. Soon after, damage appears in the upper reticular dermis. Thus, 70% HF needs only 5 min of contact to completely penetrate human skin explants. This experiment is reproducible and corroborates previous studies and clinical effects reported in accidental HF exposures. CONCLUSION/SIGNIFICANCE: This study shows that the management of HF chemical skin exposure is a question of minutes, especially for initial decontamination. These experimental observations could be useful for objectively comparing skin decontamination methods. Further studies should help to confirm these preliminary results.

Restoration of stratum corneum with nacre lipids.

To discover potential new products for the atopic dermatitis treatment, lipids extracted from nacre from the oyster Pinctada margaritifera were tested on artificially dehydrated skin explants. Expression of filaggrin and transglutaminase 1 was investigated after treatment of dehydrated skin with P. margaritifera lipid extracts according to light microscopy after labelling with specific monoclonal antibodies. The lipids were extracted from the nacre with methanol/chloroform mixture at room temperature and the extract composition was determined according to TLC and densitometry measures. Relative to the dry nacre material, a yield of extraction in lipids of 0.54% (w/w) was determined. Fatty acids, triglycerides, cholesterol and ceramides were in low abundance. Then, application of lipid formulations on skin explants previously dehydrated gave after 3 h an overexpression of filaggrin and a decrease of transglutaminase expression as shown by light microscopy. Using immunofluorescence labelling, we showed that lipids extracted from the mother of pearl of P. margaritifera induced a reconstitution of the intercellular cement of the stratum corneum. The signaling properties of the nacre lipids could be used for a development of new active product treatment against the symptoms of the dermatitis.

Preparation of TEM samples of metal-oxide interface by the focused ion beam technique.

This paper describes a procedure to prepare metal-oxide interfaces for transmission electron microscopy by the focused ion beam technique. The advantage of this procedure is to allow the observation of metal-oxide interfaces of irradiated samples with a homogeneous thickness without the need to have an instrument inside laboratories that are specialized for the manipulation of irradiated materials. A transmission electron microscopy sample is prepared by this method and analysed.

Human keratinocyte radiosensitivity is linked to redox modulation.

BACKGROUND: Ionising radiation-induced reactive oxygen species (ROS) overproduction induces keratinocyte alterations and constitutes one of the most common effects after therapeutic gamma-irradiation. ROS production is controlled by a complex enzymatic system. OBJECTIVE: The aim of our study is to analyse the role of radiation-induced oxidative stress in keratinocytes death by apoptosis. We hypothesized that keratinocyte capacity to hamper radiation-induced ROS generation may control their radiosensitivity. METHODS: For this purpose, an original human skin explant model was developed and two types of human epidermal cells were used: primary keratinocytes NHEK and spontaneous non-tumourigenic cell line HaCaT. RESULTS: cDNA-arrays analysis was performed 24h after a 20Gy gamma-radiation and revealed down-regulation of genes involved in oxidative stress control and the apoptosis process. This was confirmed by alterations in catalase, GPx and SOD enzymatic activities. This redox modulation was concomitant to the down-regulation of anti-apoptotic genes and up-regulation of some pro-apoptotic genes (caspase 10, ubiquitin C). Interestingly TUNEL labelling revealed an increase in the number of apoptotic cells. We also demonstrated a differential inducibility of the cell antioxidant network in two keratinocyte lines, which results in a differential cellular level of ROS, explaining their different radiosensitivities. CONCLUSION: Keratinocytes apoptosis is partly dependent on ROS production after exposure to gamma-rays. In addition, the differential radiosensitivity of keratinocytes is linked to different oxidative stress responses.

FIB, TEM and LA-ICPMS investigations on melt inclusions in Martian meteorites-Analytical capabilities and geochemical insights.

In order to obtain full information coverage on melt inclusions in Martian meteorites (subgroup nakhlites) complementary micro-analytical techniques were used, i.e. focused ion beam, transmission electron microscopy and laser ablation. Using focused ion beam several lamellae for transmission electron microscopy were prepared and secondary electron images of cross-sections could be acquired. Laser ablation-inductively coupled plasma mass spectrometry analyses were performed on selected inclusions to obtain mass-oriented bulk composition of inclusions at depth. The differences in composition between melt inclusions in olivine and augite crystals would suggest a xenocrystic origin for olivine. Furthermore, electron diffraction patterns clearly indicated that the SiO(2)-rich phase in inclusions from augite in meteorites from Northwest Africa site is re-crystallized, whereas it is still vitreous in the inclusions from Nakhla sampling site. Therefore, different post-entrapment evolutions were active for the two nakhlite meteorite sets, the Nakhla and the NWA817 set. Melt inclusions in Nakhla olivine presented alteration veins, which were presumably produced before their landing on Earth. If this is the case, this would indicate a alteration stage already on Mars with all the consequence in terms of climate history. Melt inclusions in Nakhla augite resulted unaffected by any alteration or modification following the entrapment, and therefore represent the best candidate to indicate the pristine magma composition.

Site-specific specimen preparation by focused ion beam milling for transmission electron microscopy of metal matrix composites.

This work describes the application and usefulness of the focused ion beam (FIB) technique for the preparation of transmission electron microscopy (TEM) samples from metal matrix composite materials. Results on an Aldiamond composite, manufactured by the squeeze casting infiltration process, were chosen for demonstration. It is almost impossible to prepare TEM specimens of this material by any other conventional method owing to the presence of highly inhomogeneous phases and reinforcement diamond particles. The present article gives a detailed account of the salient features of the FIB technique and its operation. One of the big advantages is the possibility to prepare site-specific TEM specimens with high spatial resolution. The artifacts occurring during the specimen preparation, for example, Ga-ion implantation, curtain effects, amorphous layers, bending of the lamella, or different milling behaviors of the materials have been discussed. Furthermore, TEM examination of the specimens prepared revealed an ultrafine amorphous layer of graphite formed at the interface between the Al and diamond particles that may affect the interfacial properties of the composite materials. This may not have been feasible without the successful application of the FIB technique for production of good quality site-specific TEM specimens.