Paul Zaslansky

Cooperative deformation of mineral and collagen in bone at the nanoscale

In biomineralized tissues such as bone, the recurring structural motif at the supramolecular level is an anisotropic stiff inorganic component reinforcing the soft organic matrix. The high toughness and defect tolerance of natural biomineralized composites is believed to arise from these nanometer scale structural motifs. Specifically, load transfer in bone has been proposed to occur by a transfer of tensile strains between the stiff inorganic (mineral apatite) particles via shearing in the intervening soft organic (collagen) layers. This raises the question as to how and to what extent do the mineral particles and fibrils deform concurrently in response to tissue deformation. Here we show that both mineral nanoparticles and the enclosing mineralized fibril deform initially elastically, but to different degrees. Using in situ tensile testing with combined high brilliance synchrotron X-ray diffraction and scattering on the same sample, we show that tissue, fibrils, and mineral particles take up successively lower levels of strain, in a ratio of 12:5:2. The maximum strain seen in mineral nanoparticles (≈0.15–0.20%) can reach up to twice the fracture strain calculated for bulk apatite. The results are consistent with a staggered model of load transfer in bone matrix, exemplifying the hierarchical nature of bone deformation. We believe this process results in a mechanism of fibril–matrix decoupling for protecting the brittle mineral phase in bone, while effectively redistributing the strain energy within the bone tissue.

On the mineral in collagen of human crown dentine

Dentine, the main material of mammalian teeth, contains mineral platelets that are embedded in a collagen fiber mesh. These particles entail stiffness and longevity, which is important for human teeth because these organs do not remodel. By means of small angle X-ray scattering, we mapped 2D and 3D variations in mineral particle characteristics in molar crowns. Our results show that the mean mineral-platelet thickness of 3.2 nm decreases to 2.6 nm within the shallow 300 microm beneath the dentin-enamel junction (DEJ), and that these platelets become still thinner albeit moderately in deep dentine surrounding the pulp. The mineral volume fraction in crown dentine is mostly 50% except for a 250 microm layer beneath the DEJ. Most of the mineral particles are randomly orientated, with about 20% having a preferred orientation that is parallel to the plane of the DEJ. Beneath the cusps and close to the margins of enamel, higher co-alignment is found: 40% of the particles reveal orientations that match expected load trajectories that are imposed on teeth during mastication in the general cusp-root direction. This suggests that variations in mineral platelet arrangements help to locally tune dentine anisotropy and stiffness. The serendipitous finding of incipient caries suggests that at least in early stages of pathological destruction, mineral particle thickness and orientation resemble those of the intact tissue.

Enamel-like apatite crown covering amorphous mineral in a crayfish mandible

Carbonated hydroxyapatite is the mineral found in vertebrate bones and teeth, whereas invertebrates utilize calcium carbonate in their mineralized organs. In particular, stable amorphous calcium carbonate is found in many crustaceans. Here we report on an unusual, crystalline enamel-like apatite layer found in the mandibles of the arthropod Cherax quadricarinatus (freshwater crayfish). Despite their very different thermodynamic stabilities, amorphous calcium carbonate, amorphous calcium phosphate, calcite and fluorapatite coexist in well-defined functional layers in close proximity within the mandible. The softer amorphous minerals are found primarily in the bulk of the mandible whereas apatite, the harder and less soluble mineral, forms a wear-resistant, enamel-like coating of the molar tooth. Our findings suggest a unique case of convergent evolution, where similar functional challenges of mastication led to independent developments of structurally and mechanically similar, apatite-based layers in the teeth of genetically remote phyla: vertebrates and crustaceans.

Six-dimensional real and reciprocal space small-angle X-ray scattering tomography

When used in combination with raster scanning, small-angle X-ray scattering (SAXS) has proven to be a valuable imaging technique of the nanoscale, for example of bone, teeth and brain matter. Although two-dimensional projection imaging has been used to characterize various materials successfully, its three-dimensional extension, SAXS computed tomography, poses substantial challenges, which have yet to be overcome. Previous work using SAXS computed tomography was unable to preserve oriented SAXS signals during reconstruction. Here we present a solution to this problem and obtain a complete SAXS computed tomography, which preserves oriented scattering information. By introducing virtual tomography axes, we take advantage of the two-dimensional SAXS information recorded on an area detector and use it to reconstruct the full three-dimensional scattering distribution in reciprocal space for each voxel of the three-dimensional object in real space. The presented method could be of interest for a combined six-dimensional real and reciprocal space characterization of mesoscopic materials with hierarchically structured features with length scales ranging from a few nanometres to a few millimetres—for example, biomaterials such as bone or teeth, or functional materials such as fuel-cell or battery components.

Monoclinic phase transformations of zirconia-based dental prostheses, induced by clinically practised surface manipulations

Full-ceramic zirconia crowns and bridges have become very popular with dentists and patients because of their excellent esthetics and mechanical properties. We studied phase transformations within the outermost surface layer of 3 mol.% yttria-stabilized zirconia (Y-TZP) samples of small, clinically relevant thicknesses, manipulated by polishing, grinding and fracture as might be encountered in everyday clinical practice. Stress-induced transformations of the tetragonal phase were studied in three dimensions in order to better understand the organization and extent of the monoclinically transformed phase. By means of laboratory- and synchrotron-based X-ray diffraction measurements, coupled with electron microscopy and multimodal tomography, it was possible for the first time to visualize and quantify the phase distributions non-destructively and in three dimensions. Highly variable degrees of local transformation result in ragged transformed zones of very inhomogeneous thickness. The overall thickness of the transformation layers strongly depends on the severity and rate of loading. Gentle diamond cutting resulted in surprisingly low transformation ratios of less than 0.1%. When Y-TZP constructions are manipulated before bonding, toughness of the outer layers is reduced and they may become brittle with important implications for the stability of the bond: dental practitioners thus need to be cautious when altering the surfaces of these materials after sintering.

3D variations in human crown dentin tubule orientation: a phase-contrast microtomography study.

OBJECTIVES Tubules dominate the microstructure of dentin, and in crowns of human teeth they are surrounded by thick mineralized peritubular cuffs of high stiffness. Here we examine the three-dimensional (3D) arrangement of tubules in relation to enamel on the buccal and lingual aspects of intact premolars and molars. Specifically we investigate the angular orientation of tubules relative to the plane of the junction of dentin with enamel (DEJ) by means of wet, non-destructive and high-resolution phase-contrast (coherent) tomography. METHODS Enamel capped dentin samples (n=16), cut from the buccal and lingual surfaces of upper and lower premolar and molar teeth, were imaged in water by high-resolution synchrotron-based phase-contrast X-ray radiography. Reconstructed 3D virtual images were co-aligned with respect to the DEJ plane. The average tubule orientation was determined at increasing distances from the DEJ, based on integrated projections onto orthogonal virtual planes. The angle and curl of the tubules were determined every 100 microm to a depth of 1.4mm beneath the DEJ. RESULTS Most tubules do not extend at right angles from the DEJ. Even when they do, tubules always change their orientations substantially within the first half-millimeter zone beneath the DEJ, both on the buccal and lingual aspects of premolar and molar teeth. Tubules also tend to curl and twist within this zone. Student t-tests indicate that lower teeth seem to have greater tilts in the tubule orientations relative to the DEJ normal with an average angle of 42 degrees (+/-2.0 degrees), whereas upper teeth exhibit a smaller change of orientation, with an average of 32 degrees (+/-2.1 degrees). SIGNIFICANCE Tubules are a central characteristic of dentin, with important implications on how it is arranged and what the properties are. Knowing about the path that tubules follow is important for various reasons, ranging form improving control over restorative procedures to understanding or simulating the mechanical properties of teeth. At increasing depths of dentin beneath enamel, tubules are significantly tilted relative to the DEJ norm, which may be important to understand clinical challenges such as sensitivity, effectiveness of bonding techniques or prediction of possible paths for bacterial invasion. Our data show dissimilar average tubule angles of upper versus lower teeth with respect to the DEJ which presumably contributes to different shear responses of the tissue under function. The degree to which this may warrant improved restoratives or new adhesive techniques to enhance adhesive restorations merits further investigation.

Phase shifting speckle interferometry for determination of strain and Young's modulus of mineralized biological materials: a study of tooth dentin compression in water

Mineralized biological materials have complex hierarchical graded structures. It is therefore difficult to understand the relations between their structure and mechanical properties. We report the use of electronic speckle pattern-correlation interferometry (ESPI) combined with a mechanical compression apparatus to measure the strain and Youngs modulus of root dentin compressed under water. We describe the optomechanical instrumentation, experimental techniques and procedures needed to measure cubes as small as 1 x 1 x 2 mm. Calibration of the method is performed using aluminum, which shows that the measurements are accurate within 3% of the compression modulus reported for standard aluminum 6061. Our results reveal that the compression moduli of root dentin from the buccal and lingual sides of the root are quite different from the moduli of the interproximal sides. Root dentin from interproximal locations is found to have an average modulus of 21.3 GPa, which is about 40% stiffer than root dentin from the buccal and lingual locations, found to have a modulus of 15.0 GPa. Our approach can be used to map deformations on irregular surfaces, and measure strain on wet samples of varying sizes. This can be extended to the study of other biological materials including bone and synthetic biomaterials.

Tooth–PDL–bone complex: Response to compressive loads encountered during mastication – A review

The components of the tooth-periodontal ligament (PDL)-alveolar bone complex act in a synergistic manner to dissipate the loads incurred during mastication. The complex incorporates a diverse array of structural features for this purpose. These include the non-mineralized and hence soft PDL that absorbs much of the initial loads. The internal structure of the tooth also includes soft interphases that essentially surround the dentine core. These interphases, although stiffer than the PDL, still are more compliant than the dentine core, and are thus key components that allow the tooth itself to deform and hence help dissipate the compressive loads. There is also direct evidence that even under moderate compressive loads, when the tooth moves in the alveolar bone socket, this movement is guided by specific locations where the tooth comes into contact with the bone surface. The combination of all these responses to load is that each tooth type appears to move and deform in a specific manner when loaded. Much, however, still remains to be learned about these three-dimensional responses to load and the factors that control them. Such an understanding will have major implications for dentistry, that include a better understanding of phenomena such as abfraction, the manner in which tooth implants function even in the absence of a PDL-like tissue and the implications to bone remodelling of the movements imposed during orthodontic interventions.

Identification of root filling interfaces by microscopy and tomography methods

AIM To assess differences in observed cross-sectional areas of root canals and filling materials, as imaged by three microscopy and two tomography methods. METHODOLOGY Six roots filled with laterally compacted Gutta-percha and AH26 were scanned with phase-contrast enhanced microtomography in a synchrotron facility. Reconstructed virtual slices were compared with sections of both wet and acrylic-embedded roots, evaluated also by light and electron microscopy (EM) and laboratory-based microtomography (μCT). The different contrasts of Gutta-percha, voids, sealer and root dentine were identified and correlated. Inner canal border, outer Gutta-percha rim and the external margin of a void were manually delineated, and the enclosed areas were repeatedly measured by three observers. Interobserver and interimaging method differences were tested by 2-way anova with Bonferroni adjustments (P < 0.05). Percentages of Gutta-percha-filled canal areas (PGP) were determined. RESULTS Phase-contrast enhanced microtomography revealed internal interfaces and detailed 3D volumes of accentuated voids as well as micrometre-sized particles and gaps within the treated roots. Overestimates in the cross-sectional areas were obtained by light microscopy, whereas underestimates were obtained by μCT and EM. Differences exceeded 40%; however, PGP values by all methods were within 5% for the same slice. Differences between observers were sometimes significant, but they were not method related (<3%). CONCLUSIONS Phase-contrast enhanced microtomography is a powerful non-destructive ex vivo investigation method for studying the interfaces within root canals and filling materials at a micrometre resolution. The method does not require damage-prone sectioning/polishing during sample preparation procedures. Caution should be used when quantifying the extent of Gutta-percha in root fillings by measurements using μCT, light and EM.

Skeletal maturity leads to a reduction in the strain magnitudes induced within the bone: A murine tibia study

Bone adapts to changes in the local mechanical environment (e.g. strains) through formation and resorption processes. However, the bone adaptation response is significantly reduced with increasing age. The mechanical strains induced within the bone by external loading are determined by bone morphology and tissue material properties. Although it is known that changes in bone mass, architecture and bone tissue quality occur with age, to what extent they contribute to the altered bone adaptation response remains to be determined. This study investigated alterations in strains induced in the tibia of different aged female C57Bl/6J mice (young, 10-week-old; adult, 26-week-old; and elderly, 78-week-old) subjected to in vivo compressive loading. Using a combined in vivo/in silico approach, the strains in the bones were assessed by both strain gauging and finite element modeling experiments. In cortical bone, strain magnitudes induced at the mid-diaphysis decreased by 20% from young to adult mice and by 15% from adult to elderly mice. In the cancellous bone (at the proximal metaphysis), induced strains were 70% higher in young compared with adult and elderly mice. Taking into account previous studies showing a reduced bone adaptation response to mechanical loading in adulthood, these results suggest that the diminished adaptive response is in part due to a reduction in the strains induced within the bone.