Browsing by Author "Bouzy, Pascaline"

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    Anisotropy visualisation from X-ray diffraction of biological apatite in mixed phase calcified tissue samples
    (Springer, 2025-02-14) Scott, Robert; Lyburn, Iain D.; Cornford, Eleanor; Bouzy, Pascaline; Stone, Nicholas; Greenwood, Charlene; Gosling, Sarah; Arnold, Emily L.; Bouybayoune, Ihsanne; Pinder, Sarah E.; Rogers, Keith
    X-ray diffraction is widely used to characterise the mineral component of calcified tissue. Broadening of the diffraction peaks yields valuable information on the size of coherently diffracting domains, sometimes loosely described as crystallite size or crystallinity. These domains are markedly anisotropic, hence a single number describing their size is misleading. We present a novel variation on a method for visualising crystallographic anisotropy in X-ray diffraction data. This provides an intuitively interpretable depiction of crystalline domain size and anisotropy. The new method involves creating a polar plot of calculated domain thickness for peaks in a diffractogram versus crystallographic direction. Points with the least error are emphasised. Anisotropic domain dimensions are calculated by refining an ellipsoidal model in a whole pattern fit. These dimensions are then used to overlay an ellipse on the peak broadening plot. This is illustrated by application of the method to calcifications in breast tissue with suspected cancer, which frequently contain whitlockite as well as nanocrystalline apatite. Like most biogenic apatite, this exhibits markedly anisotropic peak broadening. The nature of this anisotropy offers potentially useful information on normal function and pathology of calcified tissue and is a frequently neglected crystallographic feature of these materials.
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    Breast calcification micromorphology classification
    (British Institute of Radiology, 2022-07-25) Robert Scott, Robert Scott; Iain Lyburn, Iain Lyburn; Cornford, Eleanor; Bouzy, Pascaline; Stone, Nicholas; Greenwood, Charlene; Bouybayoune, Ihsanne; Pinder, Sarah; Rogers, Keith
    Objectives: The importance of consistent terminology in describing the appearance of breast calcifications in mammography is well recognised. Imaging of calcifications using electron microscopy is a globally growing field of research. We therefore suggest that the time is ripe to develop a lexicon of terms for classifying the micromorphology of breast calcifications. Methods: Calcifications within a wide range of histological sections of breast tissue, both benign and malignant, were imaged by Scanning Electron Microscopy (SEM). These images were examined, and the micromorphology of calcifications present was grouped to create a classification system. Results: Based on the appearance of the calcifications observed, we propose five main categories for classification of the micromorphology of breast calcifications, namely Dense Homogenous, Punctulate, Banded, Spongy, and Aggregate. Conclusions: Use of the descriptive categories outlined here will help to ensure consistency and comparability of published observations on the micromorphology of breast calcifications. Advances in knowledge: This is the first time a lexicon and classification system has been proposed for the micromorphology of breast calcifications, as observed by scanning electron microscopy of histological sections. This will facilitate comparability of observed relationships between micromorphology, mammographic appearance, chemistry, and pathology.
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    Calcification microstructure reflects breast tissue microenvironment
    (Springer, 2019-12-05) Gosling, Sarah; Scott, Robert; Greenwood, Charlene; Bouzy, Pascaline; Nallala, Jayakrupakar; Lyburn, Iain Douglas; Stone, Nicholas; Rogers, Keith
    Microcalcifications are important diagnostic indicators of disease in breast tissue. Tissue microenvironments differ in many aspects between normal and cancerous cells, notably extracellular pH and glycolytic respiration. Hydroxyapatite microcalcification microstructure is also found to differ between tissue pathologies, including differential ion substitutions and the presence of additional crystallographic phases. Distinguishing between tissue pathologies at an early stage is essential to improve patient experience and diagnostic accuracy, leading to better disease outcome. This study explores the hypothesis that microenvironment features may become immortalised within calcification crystallite characteristics thus becoming indicators of tissue pathology. In total, 55 breast calcifications incorporating 3 tissue pathologies (benign – B2, ductal carcinoma in-situ - B5a and invasive malignancy - B5b) from archive formalin-fixed paraffin-embedded core needle breast biopsies were analysed using X-ray diffraction. Crystallite size and strain were determined from 548 diffractograms using Williamson-Hall analysis. There was an increased crystallinity of hydroxyapatite with tissue malignancy compared to benign tissue. Coherence length was significantly correlated with pathology grade in all basis crystallographic directions (P < 0.01), with a greater difference between benign and in situ disease compared to in-situ disease and invasive malignancy. Crystallite size and non-uniform strain contributed to peak broadening in all three pathologies. Furthermore, crystallite size and non-uniform strain normal to the basal planes increased significantly with malignancy (P < 0.05). Our findings support the view that tissue microenvironments can influence differing formation mechanisms of hydroxyapatite through acidic precursors, leading to differential substitution of carbonate into the hydroxide and phosphate sites, causing significant changes in crystallite size and non-uniform strain.
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    Exploration of utility of combined optical photothermal infrared and Raman imaging for investigating the chemical composition of microcalcifications in breast cancer
    (Royal Society of Chemistry, 2023-02-21) Bouzy, Pascaline; Lyburn, Iain Douglas; Pinder, Sarah E.; Scott, Robert; Mansfield, Jessica; Moger, Julian; Greenwood, Charlene; Bouybayoune, Ihssane; Cornford, Eleanor; Rogers, Keith; Stone, Nick
    Microcalcifications play an important role in cancer detection. They are evaluated by their radiological and histological characteristics but it is challenging to find a link between their morphology, their composition and the nature of a specific type of breast lesion. Whilst there are some mammographic features that are either typically benign or typically malignant often the appearances are indeterminate. Here, we explore a large range of vibrational spectroscopic and multiphoton imaging techniques in order to gain more information about the composition of the microcalcifications. For the first time, we validated the presence of carbonate ions in the microcalcifications by O-PTIR and Raman spectroscopy at the same time, the same location and the same high resolution (0.5 μm). Furthermore, the use of multiphoton imaging allowed us to create stimulated Raman histology (SRH) images which mimic histological images with all chemical information. In conclusion, we established a protocol for efficiently analysing the microcalcifications by iteratively refining the area of interest.
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    A time-course Raman spectroscopic analysis of spontaneous in vitro microcalcifications in a breast cancer cell line
    (Nature Publishing Group, 2021-06-11) Bouzy, Pascaline; O’Grady, Shane; Madupalli, Honey; Tecklenburg, Mary; Rogers, Keith; Palombo, Francesca; Morgan, Maria P.; Stone, Nicholas
    Microcalcifications are early markers of breast cancer and can provide valuable prognostic information to support clinical decision-making. Current detection of calcifications in breast tissue is based on X-ray mammography, which involves the use of ionizing radiation with potentially detrimental effects, or MRI scans, which have limited spatial resolution. Additionally, these techniques are not capable of discriminating between microcalcifications from benign and malignant lesions. Several studies show that vibrational spectroscopic techniques are capable of discriminating and classifying breast lesions, with a pathology grade based on the chemical composition of the microcalcifications. However, the occurrence of microcalcifications in the breast and the underlying mineralization process are still not fully understood. Using a previously established model of in vitro mineralization, the MDA-MB-231 human breast cancer cell line was induced using two osteogenic agents, inorganic phosphate (Pi) and β-glycerophosphate (βG), and direct monitoring of the mineralization process was conducted using Raman micro-spectroscopy. MDA-MB-231 cells cultured in a medium supplemented with Pi presented more rapid mineralization (by day 3) than cells exposed to βG (by day 11). A redshift of the phosphate stretching peak for cells supplemented with βG revealed the presence of different precursor phases (octacalcium phosphate) during apatite crystal formation. These results demonstrate that Raman micro-spectroscopy is a powerful tool for nondestructive analysis of mineral species and can provide valuable information for evaluating mineralization dynamics and any associated breast cancer progression, if utilized in pathological samples.
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    Translating microcalcification biomarker information into the laboratory: a preliminary assessment utilizing core biopsies obtained from sites of mammographic calcification
    (Elsevier, 2024-03-12) Lyburn, Iain D.; Scott, Robert; Cornford, Eleanor; Bouzy, Pascaline; Stone, Nicholas; Greenwood, Charlene; Bouybayoune, Ihsanne; Pinder, Sarah E.; Rogers, Keith
    The potential of breast microcalcification chemistry to provide clinically valuable intelligence is being increasingly studied. However, acquisition of crystallographic details has, to date, been limited to high brightness, synchrotron radiation sources. This study, for the first time, evaluates a laboratory-based system that interrogates histological sections containing microcalcifications. The principal objective was to determine the measurement precision of the laboratory system and assess whether this was sufficient to provide potentially clinical valuable information. Materials and methods Sections from 5 histological specimens from breast core biopsies obtained to evaluate mammographic calcification were examined using a synchrotron source and a laboratory-based instrument. The samples were chosen to represent a significant proportion of the known breast tissue, mineralogical landscape. Data were subsequently analysed using conventional methods and microcalcification characteristics such as crystallographic phase, chemical deviation from ideal stoichiometry and microstructure were determined. Results The crystallographic phase of each microcalcification (e.g., hydroxyapatite, whitlockite) was easily determined from the laboratory derived data even when a mixed phase was apparent. Lattice parameter values from the laboratory experiments agreed well with the corresponding synchrotron values and, critically, were determined to precisions that were significantly greater than required for potential clinical exploitation. Conclusion It has been shown that crystallographic characteristics of microcalcifications can be determined in the laboratory with sufficient precision to have potential clinical value. The work will thus enable exploitation acceleration of these latent microcalcification features as current dependence upon access to limited synchrotron resources is minimized.