Young's double-slit interference with single hard X-ray photons.

Double-slit interference experiments using monochromatic hard X-rays with the energy of 25 keV are presented. The experiments were performed at a synchrotron source with a distance of 110 m between the interferometer and the detector to produce an interference pattern with a sufficiently broad period that could be adequately sampled by a photon-counting detector with 75 micrometre pixels. In the single-particle version of the experiment, over one million image frames with a single registered photon in each one were collected. The sum of these frames showed a clear presence of the interference pattern with the expected period. Subsequent analysis provided an objective estimation of the minimal number of detected photons required to determine, in accordance with the Rose criterion, the presence of the photon interference. Apart from a general theoretical interest, these investigations were aimed at exploring the possibility of medical X-ray phase-contrast imaging in photon-counting regime at minimal radiation doses.

Propagation-based phase-contrast imaging of the breast: image quality and the effect of X-ray energy and radiation dose.

OBJECTIVES: Propagation-based phase-contrast computed tomography (PB-CT) is a new imaging technique that exploits refractive and absorption properties of X-rays to enhance soft tissue contrast and improve image quality. This study compares image quality of PB-CT and absorption-based CT (AB-CT) for breast imaging while exploring X-ray energy and radiation dose. METHODS: Thirty-nine mastectomy samples were scanned at energy levels of 28-34keV using a flat panel detector at radiation dose levels of 4mGy and 2mGy. Image quality was assessed using signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR), spatial resolution (res) and visibility (vis). Statistical analysis was performed to compare PB-CT images against their corresponding AB-CT images scanned at 32keV and 4mGy. RESULTS: The PB-CT images at 4mGy, across nearly all energy levels, demonstrated superior image quality than AB-CT images at the same dose. At some energy levels, the 2mGy PB-CT images also showed better image quality in terms of CNR/Res and vis compared to the 4mGy AB-CT images. At both investigated doses, SNR and SNR/res were found to have a statistically significant difference across all energy levels. The difference in vis was statistically significant at some energy levels. CONCLUSION: This study demonstrates superior image quality of PB-CT over AB-CT, with X-ray energy playing a crucial role in determining image quality parameters. ADVANCES IN KNOWLEDGE: Our findings reveal that standard dose PB-CT outperforms standard dose AB-CT across all image quality metrics. Additionally, we demonstrate that low dose PB-CT can produce superior images compared to standard dose AB-CT in terms of CNR/Res and vis.

Dysregulation of Grainyhead-like 3 expression causes widespread developmental defects.

BACKGROUND: The gene encoding the transcription factor, Grainyhead-like 3 (Grhl3), plays critical roles in mammalian development and homeostasis. Grhl3-null embryos exhibit thoraco-lumbo-sacral spina bifida and soft-tissue syndactyly. Additional studies reveal that these embryos also exhibit an epidermal proliferation/differentiation imbalance. This manifests as skin barrier defects resulting in peri-natal lethality and defective wound repair. Despite these extensive analyses of Grhl3 loss-of-function models, the consequences of gain-of-function of this gene have been difficult to achieve. RESULTS: In this study, we generated a novel mouse model that expresses Grhl3 from a transgene integrated in the Rosa26 locus on an endogenous Grhl3-null background. Expression of the transgene rescues both the neurulation and skin barrier defects of the knockout mice, allowing survival into adulthood. Despite this, the mice are not normal, exhibiting a range of phenotypes attributable to dysregulated Grhl3 expression. In mice homozygous for the transgene, we observe a severe Shaker-Waltzer phenotype associated with hearing impairment. Micro-CT scanning of the inner ear revealed profound structural alterations underlying these phenotypes. In addition, these mice exhibit other developmental anomalies including hair loss, digit defects, and epidermal dysmorphogenesis. CONCLUSION: Taken together, these findings indicate that diverse developmental processes display low tolerance to dysregulation of Grhl3.

Effect of x-ray energy on the radiological image quality in propagation-based phase-contrast computed tomography of the breast.

Purpose: Breast cancer is the most common cancer in women in developing and developed countries and is responsible for 15% of women's cancer deaths worldwide. Conventional absorption-based breast imaging techniques lack sufficient contrast for comprehensive diagnosis. Propagation-based phase-contrast computed tomography (PB-CT) is a developing technique that exploits a more contrast-sensitive property of x-rays: x-ray refraction. X-ray absorption, refraction, and contrast-to-noise in the corresponding images depend on the x-ray energy used, for the same/fixed radiation dose. The aim of this paper is to explore the relationship between x-ray energy and radiological image quality in PB-CT imaging. Approach: Thirty-nine mastectomy samples were scanned at the imaging and medical beamline at the Australian Synchrotron. Samples were scanned at various x-ray energies of 26, 28, 30, 32, 34, and 60 keV using a Hamamatsu Flat Panel detector at the same object-to-detector distance of 6 m and mean glandular dose of 4 mGy. A total of 132 image sets were produced for analysis. Seven observers rated PB-CT images against absorption-based CT (AB-CT) images of the same samples on a five-point scale. A visual grading characteristics (VGC) study was used to determine the difference in image quality. Results: PB-CT images produced at 28, 30, 32, and 34 keV x-ray energies demonstrated statistically significant higher image quality than reference AB-CT images. The optimum x-ray energy, 30 keV, displayed the largest area under the curve ( AUC VGC ) of 0.754 ( p = 0.009 ). This was followed by 32 keV ( AUC VGC = 0.731 , p ≤ 0.001 ), 34 keV ( AUC VGC = 0.723 , p ≤ 0.001 ), and 28 keV ( AUC VGC = 0.654 , p = 0.015 ). Conclusions: An optimum energy range (around 30 keV) in the PB-CT technique allows for higher image quality at a dose comparable to conventional mammographic techniques. This results in improved radiological image quality compared with conventional techniques, which may ultimately lead to higher diagnostic efficacy and a reduction in breast cancer mortalities.

Propagation-Based Phase-Contrast CT of the Breast Demonstrates Higher Quality Than Conventional Absorption-Based CT Even at Lower Radiation Dose.

RATIONALE AND OBJECTIVES: Propagation-based phase-contrast CT (PB-CT) is an advanced X-ray imaging technology that exploits both refraction and absorption of the transmitted X-ray beam. This study was aimed at optimizing the experimental conditions of PB-CT for breast cancer imaging and examined its performance relative to conventional absorption-based CT (AB-CT) in terms of image quality and radiation dose. MATERIALS AND METHODS: Surgically excised breast mastectomy specimens (n = 12) were scanned using both PB-CT and AB-CT techniques under varying imaging conditions. To evaluate the radiological image quality, visual grading characteristics (VGC) analysis was used in which 11 breast specialist radiologists compared the overall image quality of PB-CT images with respect to the corresponding AB-CT images. The area under the VGC curve was calculated to measure the differences between PB-CT and AB-CT images. RESULTS: The highest radiological quality was obtained for PB-CT images using a 32 keV energy X-ray beam and by applying the Homogeneous Transport of Intensity Equation phase retrieval with the value of its parameter γ set to one-half of the theoretically optimal value for the given materials. Using these optimized conditions, the image quality of PB-CT images obtained at 4 mGy and 2 mGy mean glandular dose was significantly higher than AB-CT images at 4 mGy (AUCVGC = 0.901, p = 0.001 and AUCVGC = 0.819, p = 0.011, respectively). CONCLUSION: PB-CT achieves a higher radiological image quality compared to AB-CT even at a considerably lower mean glandular dose. Successful translation of the PB-CT technique for breast cancer imaging can potentially result in improved breast cancer diagnosis.

Failure Evaluation of a SiC/SiC Ceramic Matrix Composite During In-Situ Loading Using Micro X-ray Computed Tomography.

In this study, we have examined ceramic matrix composites with silicon carbide fibers in a melt-infiltrated silicon carbide matrix (SiC/SiC). We subjected samples to tensile loads while collecting micro X-ray computed tomography images. The results showed the expected crack slowing mechanisms and lower resistance to crack propagation where the fibers ran parallel and perpendicular to the applied load respectively. Cracking was shown to initiate not only from the surface but also from silicon inclusions. Post heat-treated samples showed longer fiber pull-out than the pristine samples, which was incompatible with previously proposed mechanisms. Evidence for oxidation was identified and new mechanisms based on oxidation or an oxidation assisted boron nitride phase transformation was therefore proposed to explain the long pull-out. The role of oxidation emphasizes the necessity of applying oxidation resistant coatings on SiC/SiC.

An intronic mutation in Chd7 creates a cryptic splice site, causing aberrant splicing in a mouse model of CHARGE syndrome.

Alternate splicing is a critical regulator of gene expression in eukaryotes, however genetic mutations can cause erroneous splicing and disease. Most recorded splicing disorders are caused by mutations of splice donor/acceptor sites, however intronic mutations can affect splicing. Clinical exome analyses largely ignore intronic sequence, limiting the detection of mutations to within coding regions. We describe 'Trooper', a novel mouse model of CHARGE syndrome harbouring a pathogenic point mutation in Chd7. The mutation is 18 nucleotides upstream of exon 10 and creates a cryptic acceptor site, causing exon skipping and partial intron retention. This mutation, though detectable in exome sequence, was initially dismissed by computational filtering due to its intronic location. The Trooper strain exhibited many of the previously described CHARGE-like anomalies of CHD7 deficient mouse lines; including hearing impairment, vestibular hypoplasia and growth retardation. However, more common features such as facial asymmetry and circling were rarely observed. Recognition of these characteristic features prompted manual reexamination of Chd7 sequence and subsequent validation of the intronic mutation, highlighting the importance of phenotyping alongside exome analyses. The Trooper mouse serves as a valuable model of atypical CHARGE syndrome and reveals a molecular mechanism that may underpin milder clinical presentation of the syndrome.

Seeing is believing: Insights from synchrotron infrared mapping for membrane fouling in osmotic membrane bioreactors.

We employed synchrotron infrared (IR) mapping to resolve forward osmosis (FO) membrane fouling in osmotic membrane bioreactor (OMBR). Synchrotron IR mapping offers a unique perspective to elucidate the fouling mechanisms and associated consequences in OMBR operation. We demonstrated the spatial distribution and relative intensity of carbohydrate and protein longitudinally along of the fouled FO membrane at the conclusion of OMBR operation. Both transmission and attenuated total reflection (ATR) modes were used to map the cross-section and surface of the fouled FO membrane. Micro X-ray computed tomography revealed patchy, "sand-dune" features on the membrane surface at the conclusion of OMBR operation. Synchrotron IR-ATR mapping demonstrated that the development of membrane fouling layer in OMBR operation was initiated by polysaccharide-like carbohydrate, followed by layering with protein-like substance, resulting in a characteristic "sand-dune" three dimensional feature. Synchrotron FTIR mapping shed light on foulant occurrence and accumulation in the draw solution. Strong penetration of protein-like substance into membrane matrix was visualised, resulting the detection of protein adsorption in the region of membrane supporting layer.

Lab-on-a-chip device made by autohesion-bonded polymers.

Polymers have the obvious advantages of flexibility in design and cost effectiveness to fabricate a lab-on-a-chip (LOC) device. Polyether ether ketone (PEEK) in particular is very attractive choice as it adds biocompatibility in addition to the possibility of hematic sealing in a 3D design. Hereby, we extend our previous successful technology of autohesive hermetic bonding of medical implants into lab-on-a-chip devices. We explore a conceptual 3D micro channels design with hermetic potential using PEEK and PS sheets. A hermetic and mechanically strong (through tensile test) 3D multilayer device was obtained using plasma treatment with oxygen and methane as precursors followed by pressing at temperature near of Tg + 20 of the polymer with the lowest Tg (PS). This nanotexturing technique is also used to facilitate thermal and mechanical stability of the microchannels for microfluidic applications. X-ray tomography measurements showed that 3D polymer made chips, at certain plasma and press bonding conditions, have structural integrity and no deformation were detected in channels shape post thermal pressing process. The dimension stability of channels and reservoirs and the rigid interfacial region at PEEK-PS make this chip design attractive and feasible for advanced lab-on-a-chip applications.

Local Structural Damage Evaluation of a C/C-SiC Ceramic Matrix Composite.

Ceramic matrix composites (CMCs) are structural materials, which have useful properties that combine high strength at high temperatures with moderate toughness. Carbon fibers within a matrix of carbon and silicon carbide, called C/C-SiC, are a particular class of CMC noted for their high oxidation resistance. Here we use a combination of four-point bending and X-ray radiography, to study the mechanical failure of C/C-SiC CMCs. Correlating X-ray radiographic and load/displacement curve data reveals that the fiber bundles act to slow down crack propagation during four-point bending tests. We attribute this to the fact that strain energy is expended in breaking these fibers and in pulling fiber bundles out of the surrounding matrix material. In addition, we find that the local distribution and concentration of SiC plays an important role in reducing the toughness of the material.

Elemental Contrast X-ray Tomography Using Ross Filter Pairs with a Polychromatic Laboratory Source.

The majority of current laboratory based X-ray sources are polychromatic and are not tuneable. This lack of monochromaticity limits the range of applications for these sources and in particular it reduces the elemental specificity of laboratory based X-ray imaging experiments. Here we present a solution to this problem based on the use of Ross filter pairs. Although such Ross filter arrangements have been applied in proof-of-principle spectroscopy experiments, to date there have been no reports of this approach used for full-field X-ray imaging. Here we report on the experimental demonstration of Ross filter pairs being used for quasi-monochromatic, full-field imaging. This arrangement has several important benefits for laboratory based X-ray imaging including, as we demonstrate, elemental contrast enhancement. The method is demonstrated both for two-dimensional radiography and for three-dimensional X-ray tomography.

Mice lacking the conserved transcription factor Grainyhead-like 3 (Grhl3) display increased apposition of the frontal and parietal bones during embryonic development.

BACKGROUND: Increased apposition of the frontal and parietal bones of the skull during embryogenesis may be a risk factor for the subsequent development of premature skull fusion, or craniosynostosis. Human craniosynostosis is a prevalent, and often serious embryological and neonatal pathology. Other than known mutations in a small number of contributing genes, the aetiology of craniosynostosis is largely unknown. Therefore, the identification of novel genes which contribute to normal skull patterning, morphology and premature suture apposition is imperative, in order to fully understand the genetic regulation of cranial development. RESULTS: Using advanced imaging techniques and quantitative measurement, we show that genetic deletion of the highly-conserved transcription factor Grainyhead-like 3 (Grhl3) in mice (Grhl3 -/- ) leads to decreased skull size, aberrant skull morphology and premature apposition of the coronal sutures during embryogenesis. Furthermore, Grhl3 -/- mice also present with premature collagen deposition and osteoblast alignment at the sutures, and the physical interaction between the developing skull, and outermost covering of the brain (the dura mater), as well as the overlying dermis and subcutaneous tissue, appears compromised in embryos lacking Grhl3. Although Grhl3 -/- mice die at birth, we investigated skull morphology and size in adult animals lacking one Grhl3 allele (heterozygous; Grhl3 +/- ), which are viable and fertile. We found that these adult mice also present with a smaller cranial cavity, suggestive of post-natal haploinsufficiency in the context of cranial development. CONCLUSIONS: Our findings show that our Grhl3 mice present with increased apposition of the frontal and parietal bones, suggesting that Grhl3 may be involved in the developmental pathogenesis of craniosynostosis.

Spontaneous retrotransposon insertion into TNF 3'UTR causes heart valve disease and chronic polyarthritis.

Rheumatoid arthritis (RA) and ankylosing spondylitis (AS) are chronic inflammatory diseases that together affect 2-3% of the population. RA and AS predominantly involve joints, but heart disease is also a common feature in RA and AS patients. Here we have studied a new spontaneous mutation that causes severe polyarthritis in bone phenotype spontaneous mutation 1 (BPSM1) mice. In addition to joint destruction, mutant mice also develop aortic root aneurism and aorto-mitral valve disease that can be fatal depending on the genetic background. The cause of the disease is the spontaneous insertion of a retrotransposon into the 3' untranslated region (3'UTR) of the tumor necrosis factor (TNF), which triggers its strong overexpression in myeloid cells. We found that several members of a family of RNA-binding, CCCH-containing zinc-finger proteins control TNF expression through its 3'UTR, and we identified a previously unidentified regulatory element in the UTR. The disease in BPSM1 mice is independent of the adaptive immune system and does not appear to involve inflammatory cytokines other than TNF. To our knowledge, this is the first animal model showing both polyarthritis and heart disease as a direct result of TNF deregulation. These results emphasize the therapeutic potential of anti-TNF drugs for the treatment of heart valve disease and identify potential therapeutic targets to control TNF expression and inflammation.

Mice Haploinsufficient for Ets1 and Fli1 Display Middle Ear Abnormalities and Model Aspects of Jacobsen Syndrome.

E26 transformation-specific 1 (ETS1) and friend leukemia integration 1 (FLI1) are members of the ETS family of transcription factors, of which there are 28 in humans. Both genes are hemizygous in Jacobsen syndrome, an 11q contiguous gene deletion disorder involving thrombocytopenia, facial dysmorphism, growth and mental retardation, malformation of the heart and other organs, and hearing impairment associated with recurrent ear infections. To determine whether any of these defects are because of hemizygosity for ETS1 and FLI1, we characterized the phenotype of mice heterozygous for mutant alleles of Ets1 and Fli1. Fli1(+/-) mice displayed mild thrombocytopenia, as did Ets1(+/-)Fli1(+/-) animals. Fli1(+/-) and Ets1(+/-)Fli1(+/-) mice also displayed craniofacial abnormalities, including a small middle ear cavity, short nasal bone, and malformed interface between the nasal bone process and cartilaginous nasal septum. They exhibited hearing impairment, otitis media, fusions of ossicles to the middle ear wall, and deformed stapes. Hearing impairment was more penetrant and stapes malformations were more severe in Ets1(+/-)Fli1(+/-) mice than in Fli1(+/-) mice, indicating partial functional redundancy of these transcription factors during auditory development. Our findings indicate that the short nose, otitis media, and hearing impairment in Jacobsen syndrome are likely because of hemizygosity for ETS1 and FLI1.

Micro X-ray computed tomography of pits and fissures.

BACKGROUND: Most tooth cavities start to develop inside pits and fissures where brushing, saliva, fluoride toothpaste or mouthwashes have no access. Therefore, 3D morphology of pits and fissures is an important subject to be studied in relation to maintaining proper oral hygiene. OBJECTIVE: This study aimed to present high resolution images of pits and fissures in human teeth using a laboratory-based micro X-ray Computed Tomography (mXCT) and also to present the basic structure evaluation that could be obtained. METHODS: Three human wisdom teeth were examined. Two different set-ups were used in this study achieving resolution of 14.59 µm (Field of View of 14.9 mm) and resolution of 4.43 µm (FOV of 4.5 mm) respectively. Automated segmentation was performed for further evaluation to distinguish between empty space (pits and fissures) and the filled space (enamel). RESULTS: The 3D tomography results demonstrate detailed morphology with accurate dimensions and the locations of the pits and fissures, which is important to investigate the relationship with tooth decay that mostly starts deep inside pits and fissures. CONCLUSIONS: Segmentation from the mXCT imaging of pits and fissures provides obvious visual evidence to help in promotion of oral health and to improve personal tooth care in preventive treatment protocols.

Biomedical studies on temporal bones of the first multi-channel cochlear implant patient at the University of Melbourne.

OBJECTIVE: To analyse the temporal bones and implant of the first University of Melbourne's (UOM) patient (MC-1) to receive the multi-channel cochlear prosthesis. METHODS: The left cochlea was implanted with the prototype multi-channel cochlear prosthesis on 1 August 1978, and the Cochlear versions CI-22 and CI-24 on 22 June 1983 and 10 November 1998, respectively. MC-1 died in 2007. RESULTS: Plain X-rays of the temporal bones showed that after the CI-22 had been explanted seven electrode bands remained in situ. Micro-CT scans also revealed a partially united fracture transecting the left implanted and right control cochleae. Histology indicated a total loss of the organ of Corti on both sides, and a tear of the left basilar membrane. In addition, there was a dense fibrous capsule with heterotopic bone surrounding one proximal band of the CI-22 array that restricted its removal. This pathology was associated with dark particulate material within macrophages, probably due to the release of platinum from the electrode bands. Scanning electron microscopy (SEM) showed possible corrosion of platinum and surface roughening. Three-dimensional reconstruction of the cochlear histology demonstrated the position of the electrode tracts (C1-22 and CI-24) in relation to the spiral ganglion, which showed 85-90% loss of ganglion cells. DISCUSSION AND CONCLUSIONS: This study confirms our first histopathological findings that our first free-fitting banded electrode array produced moderate trauma to the cochlea when inserted around the scala tympani of the basal turn. The difficulty in extraction was most likely due to one band being surrounded by an unusually large amount of fibrous tissue and bone, with an electrode band caught due to surface irregularities. Some surface corrosion and a small degree of platinum deposition in the tissue may also help explain the outcome for this long-term cochlear implantation.

CHD7 deficiency in "Looper", a new mouse model of CHARGE syndrome, results in ossicle malformation, otosclerosis and hearing impairment.

CHARGE syndrome is a rare human disorder caused by mutations in the gene encoding chromodomain helicase DNA binding protein 7 (CHD7). Characteristics of CHARGE are varied and include developmental ear and hearing anomalies. Here we report a novel mouse model of CHD7 dysfunction, termed Looper. The Looper strain harbours a nonsense mutation (c.5690C>A, p.S1897X) within the Chd7 gene. Looper mice exhibit many of the clinical features of the human syndrome, consistent with previously reported CHARGE models, including growth retardation, facial asymmetry, vestibular defects, eye anomalies, hyperactivity, ossicle malformation, hearing loss and vestibular dysfunction. Looper mice display an otosclerosis-like fusion of the stapes footplate to the cochlear oval window and blepharoconjunctivitis but not coloboma. Looper mice are hyperactive and have vestibular dysfunction but do not display motor impairment.

Whole-cell phase contrast imaging at the nanoscale using Fresnel coherent diffractive imaging tomography.

X-ray tomography can provide structural information of whole cells in close to their native state. Radiation-induced damage, however, imposes a practical limit to image resolution, and as such, a choice between damage, image contrast, and image resolution must be made. New coherent diffractive imaging techniques, such Fresnel Coherent Diffractive Imaging (FCDI), allows quantitative phase information with exceptional dose efficiency, high contrast, and nano-scale resolution. Here we present three-dimensional quantitative images of a whole eukaryotic cell by FCDI at a spatial resolution below 70 nm with sufficient phase contrast to distinguish major cellular components. From our data, we estimate that the minimum dose required for a similar resolution is close to that predicted by the Rose criterion, considerably below accepted estimates of the maximum dose a frozen-hydrated cell can tolerate. Based on the dose efficiency, contrast, and resolution achieved, we expect this technique will find immediate applications in tomographic cellular characterisation.

Polychromatic X-ray tomography: direct quantitative phase reconstruction.

We describe a direct quantitative phase reconstruction approach using an X-ray laboratory-based source. Using a single phase-contrast image from each tomographic projection we show that it is possible to modify the filter term in a filtered back projection reconstruction to take account of the broad spectrum from a laboratory source. The accessibility of conventional X-ray laboratory sources makes this method very useful for quantitative phase imaging of homogeneous and weakly absorbing objects.

Quantitative phase imaging with polychromatic x-ray sources.

We introduce theoretically and demonstrate experimentally a contrast transfer function based phase retrieval algorithm that reconstructs the projected thickness of an homogeneous sample using a polychromatic x-ray source. We show excellent quantitative recovery of test samples in 2D using a synchrotron source with significant harmonic contamination, and in 3D using a laboratory source.