Conical shell X-ray beam tomosynthesis and micro-computed tomography for microarchitectural characterisation.

Bone quality is commonly used to diagnose bone diseases such as osteoporosis, with many studies focusing on microarchitecture for fracture prediction. In this study a bovine distal femur was imaged using both micro-computed tomography (µCT) and tomosynthesis using focal construct geometry (FCG) for comparison of microarchitectural parameters. Six regions of interest (ROIs) were compared between the two imaging modalities, with both global and adaptive methods used to binarize the images. FCG images were downsampled to the same pixel size as the µCT images. Bone morphometrics were determined using BoneJ, for each imaging modality, binarization technique and ROI. Bone area/total area was found to have few significant differences between FCG and µCT (p < 0.05 for two of six ROIs). Fractal Dimension had only one significant difference (p < 0.05 for one of six ROIs) between µCT and downsampled FCG (where pixel size was equalized). Trabecular thickness and trabecular spacing were observed to follow trends as observed for the corresponding µCT images, although many absolute values were significantly different (p < 0.05 for between one and six ROIs depending on image types used). This study demonstrates the utility of tomosynthesis for measurement of microarchitectural morphometrics.

Microcalcification crystallography as a potential marker of DCIS recurrence.

Ductal carcinoma in-situ (DCIS) accounts for 20-25% of all new breast cancer diagnoses. DCIS has an uncertain risk of progression to invasive breast cancer and a lack of predictive biomarkers may result in relatively high levels (~ 75%) of overtreatment. To identify unique prognostic biomarkers of invasive progression, crystallographic and chemical features of DCIS microcalcifications have been explored. Samples from patients with at least 5-years of follow up and no known recurrence (174 calcifications in 67 patients) or ipsilateral invasive breast cancer recurrence (179 microcalcifications in 57 patients) were studied. Significant differences were noted between the two groups including whitlockite relative mass, hydroxyapatite and whitlockite crystal maturity and, elementally, sodium to calcium ion ratio. A preliminary predictive model for DCIS to invasive cancer progression was developed from these parameters with an AUC of 0.797. These results provide insights into the differing DCIS tissue microenvironments, and how these impact microcalcification formation.

Investigating pair distribution function use in analysis of nanocrystalline hydroxyapatite and carbonate-substituted hydroxyapatite.

Hydroxyapatite (HA) is a complex material, which is often nanocrystalline when found within a biological setting. This work has directly compared the structural characteristics derived from data collected using a conventional laboratory-based X-ray diffractometer with those collected from a dedicated pair distribution function (PDF) beamline at Diamond Light Source. In particular, the application of PDF analysis methods to carbonated HA is evaluated. 20 synthetic samples were measured using both X-ray diffraction (XRD) and PDFs. Both Rietveld refinement (of laboratory XRD data) and real-space refinement (of PDF data) were used to analyse all samples. The results of Rietveld and real-space refinements were compared to evaluate their application to crystalline and nanocrystalline hydroxyapatite. Significant relationships were observed between real-space refinement parameters and increasing carbonate substitution. Understanding the local order of synthetic hydroxyapatite can benefit several fields, including both biomedical and clinical settings.

Age related changes of rib cortical bone matrix and the application to forensic age-at-death estimation.

Forensic anthropology includes, amongst other applications, the positive identification of unknown human skeletal remains. The first step in this process is an assessment of the biological profile, that is: sex, age, stature and ancestry. In forensic contexts, age estimation is one of the main challenges in the process of identification. Recently established admissibility criteria are driving researchers towards standardisation of methodological procedures. Despite these changes, experience still plays a central role in anthropological examinations. In order to avoid this issue, age estimation procedures (i) must be presented to the scientific community and published in peer reviewed journals, (ii) accurately explained in terms of procedure and (iii) present clear information about the accuracy of the estimation and possible error rates. In order to fulfil all these requirements, a number of methods based on physiological processes which result in biochemical changes in various tissue structures at the molecular level, such as modifications in DNA-methylation and telomere shortening, racemization of proteins and stable isotopes analysis, have been developed. The current work proposes a new systematic approach in age estimation based on tracing physicochemical and mechanical degeneration of the rib cortical bone matrix. This study used autopsy material from 113 rib specimens. A set of 33 parameters were measured by standard bio-mechanical (nanoindentation and microindentation), physical (TGA/DSC, XRD and FTIR) and histomorphometry (porosity-ImageJ) methods. Stepwise regressions were used to create equations that would produce the best 'estimates of age at death' vs real age of the cadavers. Five equations were produced; in the best of cases an equation counting 7 parameters had an R2 = 0.863 and mean absolute error of 4.64 years. The present method meets all the admissibility criteria previously described. Furthermore, the method is experience-independent and as such can be performed without previous expert knowledge of forensic anthropology and human anatomy.

New insights into the application of pair distribution function studies to biogenic and synthetic hydroxyapatites.

Biogenic and synthetic hydroxyapatites are confounding materials whose properties remain uncertain, even after years of study. Pair distribution function (PDF) analysis was applied to hydroxyapatites in the 1970's and 1980's, but this area of research has not taken full advantage of the relatively recent advances in synchrotron facilities. Here, synchrotron X-ray PDF analysis is compared to techniques commonly used to characterise hydroxyapatite (such as wide angle X-ray scattering, Fourier-transform infrared spectroscopy and thermogravimetric analysis) for a range of biogenic and synthetic hydroxyapatites with a wide range of carbonate substitution. Contributions to the pair distribution function from collagen, carbonate and finite crystallite size were examined through principal component analysis and comparison of PDFs. Noticeable contributions from collagen were observed in biogenic PDFs when compared to synthetic PDFs (namely r < 15 Å), consistent with simulated PDFs of collagen structures. Additionally, changes in local structure were observed for PDFs of synthetic hydroxyapatites with differing carbonate content, notably in features near 4 Å, 8 Å and 19 Å. Regression models were generated to predict carbonate substitution from peak position within the PDFs.

The use of µCT and fractal dimension for fracture prediction in osteoporotic individuals.

Osteoporosis (OP) is a widespread condition with commonly associated fracture sites at the hip, vertebra and wrist. This study examines the effects of age and osteoporosis on bone quality by comparing the efficacy of using parameters which indicate bone quality (both traditional clinical parameters such as bone mineral density (BMD), as well as apparent Young's modulus determined by finite element analysis, among others) to predict fracture. Non-fracture samples were collected from the femoral heads of 83 donors (44 males, 39 females), and fracture samples were obtained from the femoral heads of 17 donors (female). Microarchitectural parameters (Bone Volume/Total Volume [BV/TV], Bone Surface/Bone Volume [BS/BV], Tissue Mineral Density [TMD, etc.]) were measured from µCT of each sample as well as 2D and 3D fractal dimension (D2D and D3D respectively). A cube was cropped from µCT images and an isotropic hexahedral element was assigned to each voxel. Finite element analysis was used to calculate the Young's modulus for each sample. Overall, values for microarchitectural characteristics, fractal dimension measurements and Young's Modulus were consistent with values within literature. Significant correlations are observed between age and BV/TV for non-fracture males and females, as well as between age and volumetric BMD (vBMD) for the same groups. Significant differences are present between age-matched non-fracture and fracture females for BV/TV, BS/BV, vBMD, TMD, D2D, D3D, (p < 0.01 for all). Properties which are not age dependent are significantly different between age-matched non-fracture and fracture specimens, indicating OP is a disease, and not just an accelerated aging process.

Inter-species variation in bone mineral behavior upon heating.

The characterization of inter-species variation in bone mineral (b-HAP) is of relevance to forensic science and archaeology, but has not previously been widely explored. Results of an investigation into unheated bone mineral and behavior of bone upon heating for 12 animal species (including human) demonstrate that b-HAP characteristics, quantitatively measured using X-ray diffraction (XRD) analysis, exhibit significant inter-species variation. Human bone was found to be significantly different to all other species in terms of b-HAP lattice parameter values from unheated and heated bone and in terms of recrystallization behavior of b-HAP upon heating bone to 600°C. The amounts of b-HAP thermal decomposition products were also significantly different for human bone heated to 1400°C compared to those obtained for most other species. Therefore, there is potential for the development of an XRD-based method of species identification, particularly one that distinguishes human from non-human bone.

Composition of uroliths in small domestic animals in the United Kingdom.

The mineral composition of 7819 small animal uroliths in the UK was determined by semi-quantitative X-ray diffraction over a period of 90 months from 2002 to 2010. Canine and feline uroliths constituted 97% of the study population and the mineral phase detected most frequently was struvite (43%), followed by calcium oxalate (41%). Uroliths from crossbreeds, Dalmatians, Yorkshire terriers and Shih Tzus accounted for almost 30% of all canine uroliths, with the highest frequency in Dalmatians, which had a predominance of urate uroliths. The average ages of dogs and cats with uroliths were 7.0 years and 7.4 years, respectively. The ratio of the number of dogs presenting with struvite compared to oxalate phases reached a maximum at 3years of age.

High throughput X-ray diffraction analysis of combinatorial polycrystalline thin film libraries.

With the increasing demand for new materials, analytical techniques which are able to rapidly characterize a large number of samples are becoming indispensable. Thin film technology has the potential to improve the amount of information contained on as-deposited samples by creating compositionally graded libraries. Conventionally, raster scan methods are used to interrogate such libraries but, in this paper, a different approach is presented to provide a method of high-throughput data collection and analysis using an X-ray diffraction (XRD) probe. An extended X-ray beam was used to illuminate the libraries, and a large area detector was used to collect the data. A new algorithm "Bandit" has been employed to analyze the collected data and extract the crystallographic information. The results of the technique have been compared with the raster scans showing that the algorithm provides reliable data at a significantly increased data acquisition speed.

Small angle scatter imaging from wide beam diffraction patterns.

In this paper we report on the extension of the technique of mapping small angle x-ray scatter (SAXS) across a soft material specimen several millimetres square. In the conventional SAXS mapping technique a pencil beam of x-rays is raster scanned over the specimen with the scatter pattern recorded from each point in the raster. In our technique a wide, parallel beam is used, speeding up the data collection time considerably. An image processing algorithm is used to separate the scatter pattern features from individual points along the line of the beam. To test the efficacy of the technique a phantom was constructed using gelatin and rat tail tendon collagen. Collagen fibres in the phantom were arranged in quarters horizontally, diagonally and vertically leaving one quarter with just gelatin. The phantom was used to collect both raster scanned sets of SAXS patterns spaced at 0.25 mm horizontally and vertically and also a wide beam data set. The width of the beam in this case was approximately 7 mm. Using the third-order diffraction of rat tail tendon intensity data were gathered from each SAXS pattern and used to construct a map. Data from the raster scan image and that from the wide beam are compared. Finally using a phantom made from dehydrated rat tail tendon and paraffin wax a tomographic slice constructed using data from SAXS patterns is shown.