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The application of secondary electron (SE) imaging, backscattered electron imaging (BSE) and electron backscattered diffraction (EBSD) was investigated in this work to study the bacterial adhesion and proliferation on a commercially pure titanium (cp Ti) and a Ti6Al4V alloy (Ti 64) with respect to substrate microstructure and chemical composition. Adherence of Gram-positive Staphylococcus epidermidis 11047 and Streptococcus sanguinis GW2, and Gram-negative Serratia sp. NCIMB 40259 and Escherichia coli 10418 was compared on cp Ti, Ti 64, pure aluminium (Al) and vanadium (V). The substrate microstructure and the bacterial distribution on these metals were characterised using SE, BSE and EBSD imaging. It was observed that titanium alloy-phase structure, grain boundaries and grain orientation did not influence bacterial adherence or proliferation at microscale. Adherence of all four strains was similar on cp Ti and Ti 64 surfaces whilst inhibited on pure Al. This work establishes a nondestructive and straight-forward statistical method to analyse the relationship between microbial distribution and metal alloy structure., (© 2017 The Authors Journal of Microscopy © 2017 Royal Microscopical Society.)

The Odontogenic keratocyst (OKC) is a cystic lesion of the jaws, which has high growth and recurrence rates compared to other cysts of the jaws (for instance, radicular cyst, which is the most common jaw cyst type). For this reason OKCs are considered by some to be benign neoplasms. There exist two sub-types of OKCs (sporadic and syndromic) and the ability to discriminate between these sub-types, as well as other jaw cysts, is an important task in terms of disease diagnosis and prognosis. With the development of digital pathology, computational algorithms have become central to addressing this type of problem. Considering that only basic feature-based methods have been investigated in this problem before, we propose to use a different approach (the Bouligand-Minkowski descriptors) to assess the success rates achieved on the classification of a database of histological images of the epithelial lining of these cysts. This does not require the level of abstraction necessary to extract histologically-relevant features and therefore has the potential of being more robust than previous approaches. The descriptors were obtained by mapping pixel intensities into a three dimensional cloud of points in discrete space and applying morphological dilations with spheres of increasing radii. The descriptors were computed from the volume of the dilated set and submitted to a machine learning algorithm to classify the samples into diagnostic groups. This approach was capable of discriminating between OKCs and radicular cysts in 98% of images (100% of cases) and between the two sub-types of OKCs in 68% of images (71% of cases). These results improve over previously reported classification rates reported elsewhere and suggest that Bouligand-Minkowski descriptors are useful features to be used in histopathological images of these cysts., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Human dental tissue is a hydrated biological mineral composite. In terms of volume and mass, a human tooth mainly consists of dentine and enamel. Human dental tissues have a hierarchical structure and versatile mechanical properties. The dentine enamel junction (DEJ) is an important biological interface that provides a durable bond between enamel and dentine that is a life-long success story: while intact and free from disease, this interface does not fail despite the harsh thermo-mechanical loading in the oral cavity. The underlying reasons for such remarkable strength and durability are still not fully clear from the structural and mechanical perspectives. One possibility is that, in an example of residual stress engineering, evolution has led to the formation of a layer of inelastic strain adjacent to the DEJ during odontogenesis (tooth formation). However, due to significant experimental and interpretational challenges, no meaningful quantification of residual stress in the vicinity of the DEJ at the appropriate spatial resolution has been reported to date. In this study, we applied a recently developed flexible and versatile method for measuring the residual elastic strain at (sub)micron-scale utilising focused ion beam (FIB) milling with digital image correlation (DIC). We report the results that span the transition from human dentine to enamel, and incorporate the material lying at and in the vicinity of the DEJ. The capability of observing the association between internal architecture and the residual elastic strain state at the micrometre scale is useful for understanding the remarkable performance of the DEJ and may help the creation of improved biomimetic materials for clinical and engineering applications., Statement of Significance: We studied the micron-scale residual stresses that exist within human teeth, between enamel (outer tooth shell, hardest substance in the human body) and dentine (soft bone-like vascularised tooth core). The dentine-enamel junction (DEJ) is an extremely interesting example of nature's design in terms of hierarchical structuring and residual stress management. Key developments reported are systematic focused ion beam (FIB) milling and digital image correlation (DIC) micrometre scale residual strain evaluation, and the determination of principal strain direction near DEJ, correlated with internal architecture responsible for remarkable strength. This work helps understanding DEJ performance and improving biomimetic materials design for clinical and engineering applications., (Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Objectives: The objective of this study was to assess local vascular architecture in atrophic-erosive oral lichen planus (OLP)., Materials and Methods: We investigated the capillary structure of the oral mucosa in 31 patients with OLP and 32 healthy controls. Capillaries images were captured in vivo through a capillaroscope. We applied fractal analysis to quantify the microvasculature morphometric changes in the oral mucosa of atrophic-erosive OLP patients in terms of their fractal dimension (D)., Results: The oral vascular networks of atrophic-erosive OLP lesions had a significantly higher D, both in buccal mucosae (D=1.167, P=.019) and in tongue (D=1.196, P=.038), compared with the control population (1.123 for both locations, respectively)., Conclusions: The present study confirms previous literature data on a close relationship between abnormal vascular architecture and atrophic-erosive OLP. Fractal analysis provided a quantitative descriptor of the complexity of the vascular patterns, which increases in the OLP samples. These data may provide new information on the OLP pathogenesis, as well as serve as morphologic quantifiers for monitoring treatment strategies., (Copyright © 2015 Elsevier Inc. All rights reserved.)

The knowledge of the mechanical properties of dental materials related to their hierarchical structure is essential for understanding and predicting the effect of microstructural alterations on the performance of dental tissues in the context of forensic and archaeological investigation as well as laser irradiation treatment of caries. So far, few studies have focused on the nano-scale structure-mechanical function relations of human teeth altered by chemical or thermal treatment. The response of dental tissues to thermal treatment is thought to be strongly affected by the mineral crystallite size, their spatial arrangement and preferred orientation. In this study, synchrotron-based small and wide angle X-ray scattering (SAXS/WAXS) techniques were used to investigate the micro-structural alterations (mean crystalline thickness, crystal perfection and degree of alignment) of heat-affected dentine and enamel in human dental teeth. Additionally, nanoindentation mapping was applied to detect the spatial and temperature-dependent nano-mechanical properties variation. The SAXS/WAXS results revealed that the mean crystalline thickness distribution in dentine was more uniform compared with that in enamel. Although in general the mean crystalline thickness increased both in dentine and enamel as the temperature increased, the local structural variations gradually reduced. Meanwhile, the hardness and reduced modulus in enamel decreased as the temperature increased, while for dentine, the tendency reversed at high temperature. The analysis of the correlation between the ultrastructure and mechanical properties coupled with the effect of temperature demonstrates the effect of mean thickness and orientation on the local variation of mechanical property. This structural-mechanical property alteration is likely to be due to changes of HAp crystallites, thus dentine and enamel exhibit different responses at different temperatures. Our results enable an improved understanding of the mechanical properties correlation in hierarchical biological materials, and human dental tissue in particular., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Human enamel is a typical hierarchical mineralized tissue with a two-level composite structure. To date, few studies have focused on how the mechanical behaviour of this tissue is affected by both the rod orientation at the microscale and the preferred orientation of mineral crystallites at the nanoscale. In this study, wide-angle X-ray scattering was used to determine the internal lattice strain response of human enamel samples (with differing rod directions) as a function of in situ uniaxial compressive loading. Quantitative stress distribution evaluation in the birefringent mounting epoxy was performed in parallel using photoelastic techniques. The resulting experimental data was analysed using an advanced multiscale Eshelby inclusion model that takes into account the two-level hierarchical structure of human enamel, and reflects the differing rod directions and orientation distributions of hydroxyapatite crystals. The achieved satisfactory agreement between the model and the experimental data, in terms of the values of multidirectional strain components under the action of differently orientated loads, suggests that the multiscale approach captures reasonably successfully the structure-property relationship between the hierarchical architecture of human enamel and its response to the applied forces. This novel and systematic approach can be used to improve the interpretation of the mechanical properties of enamel, as well as of the textured hierarchical biomaterials in general., (Copyright © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Human dentine is a hierarchical mineralized tissue with a two-level composite structure, with tubules being the prominent structural feature at a microlevel, and collagen fibres decorated with hydroxyapatite (HAp) crystallite platelets dominating the nanoscale. Few studies have focused on this two-level structure of human dentine, where the response to mechanical loading is thought to be affected not only by the tubule volume fraction at the microscale, but also by the shape and orientation distribution of mineral crystallites, and their nanoscale spatial arrangement and alignment. In this paper, in situ elastic strain evolution within HAp in dentine subjected to uniaxial compressive loading along both longitudinal and transverse directions was characterized simultaneously by two synchrotron X-ray scattering techniques: small- and wide-angle X-ray scattering (SAXS and WAXS, respectively). WAXS allows the evaluation of the apparent modulus linking the external load to the internal HAp crystallite strain, while the nanoscale HAp distribution and arrangement can be quantified by SAXS. We proposed an improved multiscale Eshelby inclusion model that takes into account the two-level hierarchical structure, and validated it with a multidirectional experimental strain evaluation. The agreement between the simulation and measurement indicates that the multiscale hierarchical model developed here accurately reflects the structural arrangement and mechanical response of human dentine. This study benefits the comprehensive understanding of the mechanical behaviour of hierarchical biomaterials. The knowledge of the mechanical properties related to the hierarchical structure is essential for the understanding and predicting the effects of structural alterations that may occur due to disease or treatment on the performance of dental tissues and their artificial replacements., (Copyright © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

Objective: To explore tissue organization based on the geometry of cell neighborhoods in histologic preparations., Study Design: Local complexity of solid tissues was measured in images of discrete tissue compartments. Exclusive areas associated with cell nuclei (v-cells) were computed using a watershed transform of the nuclear staining intensity. Mathematical morphology was used to define neighborhood membership, distances and identify complete nested neighborhoods. Neighborhood complexity was estimated as the scaling of the number of neighbors relative to reference v-cells., Results: The methodology applied to hematoxylin-eosin-stained sections from normal, dysplastic and neoplastic oral epithelium revealed that the scaling exponent, over a finite range of neighborhood levels, is nonunique and fractional. While scaling values overlapped across classes, the average was marginally higher in neoplastic than in dysplastic and normal epithelia. The best classificatory power of the exponent was 58% correct classification into 3 diagnostic classes (11 levels) and 83% between dysplastic and neoplastic classes (13 levels)., Conclusion: V-cell architecture retains features of the original tissue classes and demonstrates an increase in tissue disorder in neoplasia. This methodology seems suitable for extracting information from tissues where identification of cell boundaries (and therefore segmentation into individual cells) is unfeasible.

The current study aimed to investigate the accuracy of fit and the reproducibility of inner crown profile for two types of high strength ceramics, IPS Empress and Procera. Procera and Empress crowns with four different morphologies were cemented to dies using zinc phosphate dental cement. Vertical and horizontal sections were made through each of the crown/die preparations and images of the vertical sections were compared for curvature reproduction by alignment using image processing. Measurements were made on horizontal sections to determine cement layer thickness. Alignment of the crowns using image analysis identified quantifiable variations in the inner surface profile compared with the outer surface of the die. The largest differences occurred from the cusp tips to the occlusal adaptation area and differences in surface profile were less pronounced for Procera than Empress crowns. Marginal gap varied independently of ceramic or internal crown shape from 7-529 microm for Procera and 26-548 microm for Empress. IPS Empress has a superior ability to reproduce the inner surface profile of the crown morphologies investigated compared with Procera. The reduced reproduction of surface profile was associated with an increased cement thickness at the occlusal contact area that may inadvertently lead to failure of the crowns functional characteristics.

Objective: To compare the architectural and morphometric features of pseudoepitheliomatous hyperplasia (PEH) associated with oral granular cell tumors (GCT), normal oral mucosa and oral epithelial dysplasia., Study Design: Quantitative comparisons between the diagnostic entities were carried out at the tissue level by estimating the fractal complexity of the epithelial connective tissue interface and at the cellular level by analyzing the morphometric features of algorithmically segmented epithelial cell areas., Results: Casewise multivariate analysis showed that the fractal properties produced a correct discrimination rate of 96.4% between PEH and normal mucosa. Cellular parameters gave a 100% correct discrimination rate between PEH and mild dysplasia. Combining the fractal and cellular properties also showed 100% discrimination between PEH and normal mucosa and between PEH and mild dysplasia., Conclusion: The results show that PEH associated with GCT displays quantifiable morphometric features that make it differentiable from normal oral mucosa and oral epithelial dysplasia.

Objective: To formulate a mathematical model of odontogenic cyst growth and establish the dynamics of cyst enlargement and role of osmotic pressure forces throughout its growth., Study Design: The model assumed a spherical cyst with a semipermeable lining of living cells and a core consisting of degraded cellular material, including generic osmotic material, fed by the continuous death of epithelial cells in the lining. The lining cells were assumed to have both elastic and viscous properties, reflecting the action of physical stresses by the surrounding cyst capsule, composed of fibroblasts and collagen fibers. The model couples the cyst radius and osmotic pressure differences resulting in a system of 2 nonlinear ordinary differential equations., Results: The model predicts that in all parameter regimens the long-time behavior of the cyst is the same and that linear radial expansion results., Conclusion: In the early and intermediate stages of cystic growth, osmotic pressure differences play an important role; however, in very large cysts, this role becomes negligible, and cell birth in the lining dominates growth.

This study aimed at quantifying the complexity of the epithelial-connective tissue interface (ECTI) in human normal mucosa, premalignant, and malignant lesions using fractal geometry. Two approaches were used to describe the complexity of 377 oral mucosa ECTI profiles. The box counting method was used to estimate their global fractal dimension, while local fractal dimensions were estimated using the mass radius relation at various local scales. The ECTI complexity significantly increased from normal through premalignant to malignant profiles in both global and local (over 283 microm) scales. Normal mucosa samples from different sites of the oral cavity also had different degrees of global complexity. Fractal geometry is a useful morphological marker of tissue complexity changes taking place during epithelial malignancy and premalignancy, and we propose it as a quantitative marker of epithelial complexity.