Application of sensitive, high-resolution imaging at a commercial lab-based X-ray micro-CT system using propagation-based phase retrieval.

Several dedicated commercial lab-based micro-computed tomography (µCT) systems exist, which provide high-resolution images of samples, with the capability to also deliver in-line phase contrast. X-ray phase contrast is particularly beneficial when visualizing very small features and weakly absorbing samples. The raw measured projections will include both phase and absorption effects. Extending our previous work that addressed the optimization of experimental conditions at the commercial ZEISS Xradia 500 Versa system, single-distance phase-contrast imaging is demonstrated on complex biological and material samples. From data captured at this system, we demonstrate extraction of the phase signal or the correction of the mixed image for the phase shift, and show how this procedure increases the contrast and removes artefacts. These high-quality images, measured without the use of a synchrotron X-ray source, demonstrate that highly sensitive, micrometre-resolution imaging of 3D volumes is widely accessible using commercially advanced laboratory devices.

Two-shot X-ray dark-field imaging.

In this article, we report on a novel acquisition scheme for time- and dose-saving retrieval of dark-field data in grating-based phase-contrast imaging. In comparison to currently available techniques, the proposed approach only requires two phase steps. More importantly, our method is capable of accurately retrieving the dark-field signal where conventional approaches fail, for instance in the case of very low photon statistics. Finally, we successfully extend two-shot dark-field imaging to tomographic investigations, by implementing an iterative reconstruction with appropriate weights. Our results indicate an important progression towards the clinical feasibility of dark-field tomography.

Basis material decomposition in spectral CT using a semi-empirical, polychromatic adaption of the Beer-Lambert model.

Following the development of energy-sensitive photon-counting detectors using high-Z sensor materials, application of spectral x-ray imaging methods to clinical practice comes into reach. However, these detectors require extensive calibration efforts in order to perform spectral imaging tasks like basis material decomposition. In this paper, we report a novel approach to basis material decomposition that utilizes a semi-empirical estimator for the number of photons registered in distinct energy bins in the presence of beam-hardening effects which can be termed as a polychromatic Beer-Lambert model. A maximum-likelihood estimator is applied to the model in order to obtain estimates of the underlying sample composition. Using a Monte-Carlo simulation of a typical clinical CT acquisition, the performance of the proposed estimator was evaluated. The estimator is shown to be unbiased and efficient according to the Cramér-Rao lower bound. In particular, the estimator is capable of operating with a minimum number of calibration measurements. Good results were obtained after calibration using less than 10 samples of known composition in a two-material attenuation basis. This opens up the possibility for fast re-calibration in the clinical routine which is considered an advantage of the proposed method over other implementations reported in the literature.

Helical X-ray phase-contrast computed tomography without phase stepping.

X-ray phase-contrast computed tomography (PCCT) using grating interferometry provides enhanced soft-tissue contrast. The possibility to use standard polychromatic laboratory sources enables an implementation into a clinical setting. Thus, PCCT has gained significant attention in recent years. However, phase-contrast CT scans still require significantly increased measurement times in comparison to conventional attenuation-based CT imaging. This is mainly due to a time-consuming stepping of a grating, which is necessary for an accurate retrieval of the phase information. In this paper, we demonstrate a novel scan technique, which directly allows the determination of the phase signal without a phase-stepping procedure. The presented work is based on moiré fringe scanning, which allows fast data acquisition in radiographic applications such as mammography or in-line product analysis. Here, we demonstrate its extension to tomography enabling a continuous helical sample rotation as routinely performed in clinical CT systems. Compared to standard phase-stepping techniques, the proposed helical fringe-scanning procedure enables faster measurements, an extended field of view and relaxes the stability requirements of the system, since the gratings remain stationary. Finally, our approach exceeds previously introduced methods by not relying on spatial interpolation to acquire the phase-contrast signal.

Bilateral filtering using the full noise covariance matrix applied to x-ray phase-contrast computed tomography.

The purpose of this work is to develop an image-based de-noising algorithm that exploits complementary information and noise statistics from multi-modal images, as they emerge in x-ray tomography techniques, for instance grating-based phase-contrast CT and spectral CT. Among the noise reduction methods, image-based de-noising is one popular approach and the so-called bilateral filter is a well known algorithm for edge-preserving filtering. We developed a generalization of the bilateral filter for the case where the imaging system provides two or more perfectly aligned images. The proposed generalization is statistically motivated and takes the full second order noise statistics of these images into account. In particular, it includes a noise correlation between the images and spatial noise correlation within the same image. The novel generalized three-dimensional bilateral filter is applied to the attenuation and phase images created with filtered backprojection reconstructions from grating-based phase-contrast tomography. In comparison to established bilateral filters, we obtain improved noise reduction and at the same time a better preservation of edges in the images on the examples of a simulated soft-tissue phantom, a human cerebellum and a human artery sample. The applied full noise covariance is determined via cross-correlation of the image noise. The filter results yield an improved feature recovery based on enhanced noise suppression and edge preservation as shown here on the example of attenuation and phase images captured with grating-based phase-contrast computed tomography. This is supported by quantitative image analysis. Without being bound to phase-contrast imaging, this generalized filter is applicable to any kind of noise-afflicted image data with or without noise correlation. Therefore, it can be utilized in various imaging applications and fields.

Grating-based X-ray Dark-field Computed Tomography of Living Mice.

Changes in x-ray attenuating tissue caused by lung disorders like emphysema or fibrosis are subtle and thus only resolved by high-resolution computed tomography (CT). The structural reorganization, however, is of strong influence for lung function. Dark-field CT (DFCT), based on small-angle scattering of x-rays, reveals such structural changes even at resolutions coarser than the pulmonary network and thus provides access to their anatomical distribution. In this proof-of-concept study we present x-ray in vivo DFCTs of lungs of a healthy, an emphysematous and a fibrotic mouse. The tomographies show excellent depiction of the distribution of structural - and thus indirectly functional - changes in lung parenchyma, on single-modality slices in dark field as well as on multimodal fusion images. Therefore, we anticipate numerous applications of DFCT in diagnostic lung imaging. We introduce a scatter-based Hounsfield Unit (sHU) scale to facilitate comparability of scans. In this newly defined sHU scale, the pathophysiological changes by emphysema and fibrosis cause a shift towards lower numbers, compared to healthy lung tissue.

Preferential binding of Staphylococcus aureus to skin sites of Th2-mediated inflammation in a murine model.

Staphylococcus aureus is found on over 90% of atopic dermatitis skin lesions and is thought to contribute to skin inflammation via the production of potent exotoxins. In contrast, less than 5% of normal subjects harbor S. aureus. This suggests that an atopic immune response itself may play a role in preferential binding of S. aureus to the skin. To examine this issue more directly, we analyzed the S. aureus binding characteristics of skin in mice undergoing different T helper type 1 cell versus T helper type 2 cell inflammatory responses using a novel in vitro bacterial binding assay. BALB/C female mice were first sensitized to ovalbumin with alum or ovalbumin with complete Freund's adjuvant to induce T helper type 2 or T helper type 1 responses, respectively. Mice were then challenged intradermally with either saline (control) or ovalbumin. Forty-eight hours later, skin specimens were obtained from the challenge sites, and the number of S. aureus binding to each skin section was quantitated. Bacterial binding was found to be significantly greater at skin sites of BALB/C mice that had been ovalbumin/alum sensitized compared with ovalbumin/complete Freund's adjuvant sensitized (p < or = 0.01). When compared to the ovalbumin sensitized/challenged skin of wild type BALB/C mice or interferon-gamma gene knockout mice, interleukin-4, but not interferon-gamma, gene knockout mice had significantly less S. aureus binding at their ovalbumin sensitized/challenged skin sites. Mutant S. aureus strains that lacked either fibronectin- or fibrinogen-binding protein expression showed significantly reduced S. aureus binding compared with the parent wild type strain (p < 0.005). Moreover, preincubation of the wild type bacteria with fibronectin or fibrinogen, but not collagen, resulted in significantly less skin binding of S. aureus (p < 0.01). Incubation of skin with interleukin-4, and less so with interferon-gamma, led to more binding of wild type S. aureus but not of an S. aureus mutant deficient in fibronectin binding protein expression. After interleukin-4 incubation, but not interferon-gamma, epidermal immunoreactivity for fibronectin was observed in murine skin explants. These results show that a T helper type 2 inflammatory environment can promote skin binding by S. aureus and that this binding is mediated by fibronectin and fibrinogen.

Staphylococcus aureus isolated from Kawasaki disease patients hyper-releases extracellular protein A.

S. aureus isolates from patients with Kawasaki disease (KD) release high levels of extracellular protein A (SpA), as compared to S. aureus in other diseases. The molecular weight of this released protein A is about 70 kDa. Extracellular KD SpA purified by affinity chromatography possessed the same amino acid sequence at the NH2-terminal IgG binding region and the same antigenic specificity as recombinant and cell-wall-bound SpA preparations. The size of DNA fragments containing the spa gene from S. aureus KD strains was 160-165 kb. All of these DNA fragments contained the igb portion encoding the IgG-binding region of KD SpA. Significantly higher molecular size of the SpA molecules hyper-released in the stationary-phase culture and the lack of production of other exo-proteins allow us to speculate that S. aureus isolated from patients with KD have mutations occurring in the agr locus.

Pepsin-digested peanut contains T-cell epitopes but no IgE epitopes.

BACKGROUND: Peanuts are a common cause of food-induced anaphylaxis and fatalities. Previous studies have demonstrated that rush immunotherapy to crude peanut extract reduces clinical symptoms triggered by oral peanut challenges, but the immunotherapy was associated with an unacceptably high incidence of systemic allergic reactions. One approach to reduce the frequency of allergic reactions would be to use a modified peanut antigen with low allergenic properties. OBJECTIVE: We sought to determine the immunologic characteristics of crude intact peanut extract before and after pepsin digestion. METHODS: We used IgE immunoblotting and assessment of T-lymphocyte responses to intact and peptic digests of peanut extracts. RESULTS: Western blot analysis of sera from 5 subjects with peanut allergy showed multiple IgE-reactive proteins in crude intact peanut extract that were eliminated after pepsin treatment of the peanut extract. In contrast, pepsin-digested peanut induced significant T-cell proliferation responses (stimulation index = 30) in vitro in PBMCs from 7 subjects with peanut allergy, albeit at lower levels than that induced by intact peanut (stimulation index = 66). Furthermore, IFN-gamma production was induced by intact peanut and pepsin-digested peanut in a concentration-dependent manner. Importantly, T-cell lines generated in response to intact peanut also reacted to pepsin-digested peanut, indicating cross-reactive T-cell epitopes in intact and pepsin-digested peanut. CONCLUSION: These findings suggest that pepsin-digested peanut may be useful in peanut immunotherapy because pepsin digestion eliminates IgE reactivity but maintains T-cell reactivity.

Staphylococcus aureus isolates from patients with Kawasaki disease express high levels of protein A.

Kawasaki disease (KD) is an acute vasculitis of young children that can be complicated by coronary artery abnormalities. Recent findings suggest that a superantigen(s) may play an important role in stimulating the immune activation associated with the disease, although the origin of this superantigen(s) is unclear. Staphylococcus aureus, isolated from the rectum or pharynx of patients with KD, secretes toxic shock syndrome toxin 1 (TSST-1). The KD isolates express low levels of other exoproteins compared to isolates from patients with toxic shock syndrome (TSS). Thus, it was previously suggested that the KD isolates may be defective in the global regulatory locus agr (for accessory gene regulator), which positively regulates these factors (D. Y. M. Leung et al., Lancet 342:1385-1388, 1993). Here we describe another characteristic of KD isolates. When considered collectively, the KD isolates were found to express higher levels of staphylococcal protein A than the TSS isolates, another characteristic of an agr-defective phenotype. This correlated with a higher level of spa mRNA in these isolates. In contrast, the KD and TSS isolates expressed comparable levels of TSST-1, consistent with previous findings (D. Y. M. Leung et al., Lancet 342:1385-1388, 1993). Analysis of RNAIII transcript levels and nucleotide sequence analysis of the RNAIII-coding region suggested that the KD isolates are not defective in RNAIII, the effector molecule of the agr regulatory system. However, induction of RNAIII transcription in the KD isolates did not result in a dramatic decrease in the amount of spa mRNA, as has been reported for other strains (F. Vandenesch, J. Kornblum, and R. P. Novick, J. Bacteriol. 173:6313-6320, 1991).

Association of toxic shock syndrome toxin-secreting and exfoliative toxin-secreting Staphylococcus aureus with Kawasaki syndrome complicated by coronary artery disease.

Kawasaki syndrome (KS) has been reported to be associated with selective expansion of Vbeta2+ T cells and either staphylococcal toxic shock syndrome toxin-1 or streptococcal pyrogenic exotoxin C in uncomplicated cases. However, there have been no previous studies on the role of superantigens in KS associated with coronary artery disease, the major complication of this illness. The present study characterized bacteria isolated from three acute KS patients who developed coronary artery disease. Staphylococcus aureus secreting either TSST-1 (n = 3) or exfoliative toxin A (n = 1), both known to stimulate expansion of Vbeta2+ T cells, were isolated from all three patients. The percent Vbeta2+ T cells was determined in three patients with coronary artery disease. On presentation, one patient demonstrated reduction, whereas the other two showed expansion, of Vbeta2+ T cells. Repeat analyses of the latter two children showed their percent Vbeta2+ T cells to decrease toward normal. These observations suggest that coronary artery disease in KS may result from superantigenic stimulation of Vbeta2+ T cells. This is also the first demonstration of an association of staphylococcal exfoliative toxin with acute KS. The observation that three different bacterial toxins associated with KS are potent activators of Vbeta2+ T cells suggests an important role for this T cell subset in the pathogenesis of this autoimmune disease.