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A key issue for the fabrication of scaffolds for tissue engineering is the development of processing techniques flexible enough to produce materials with a wide spectrum of solubility (bioresorption rates) and mechanical properties matching those of calcified tissues. These techniques must also have the capability of generating adequate porosity to further serve as a framework for cell penetration, new bone formation, and subsequent remodeling. In this study we show how hybrid organic/inorganic scaffolds with controlled microstructures can be built using robotic assisted deposition at room temperature. Polylactide or polycaprolactone scaffolds with pore sizes ranging between 200-500 microm and hydroxyapatite contents up to 70 wt % were fabricated. Compressive tests revealed an anisotropic behavior of the scaffolds, strongly dependant on their chemical composition. The inclusion of an inorganic component increased their stiffness but they were not brittle and could be easily machined even for ceramic contents up to 70 wt %. The mechanical properties of hybrid scaffolds did not degrade significantly after 20 days in simulated body fluid. However, the stiffness of pure polylactide scaffolds increased drastically due to polymer densification. Scaffolds containing bioactive glasses were also printed. After 20 days in simulated body fluid they developed an apatite layer on their surface., (Copyright (c) 2007 Wiley Periodicals, Inc.)

Objective: Glucocorticoids (GCs) alter bone strength such that patients receiving these medications have a high rate of fragility-related fractures. The purpose of this study was to assess whether concurrent treatment with GCs (prednisolone) and risedronate (an aminobisphosphonate) would prevent the reduction in bone strength induced by GCs, in a mouse model of GC-induced bone loss and in patients enrolled in a clinical study., Methods: We evaluated mice treated with prednisolone pellets alone, GCs plus risedronate, or placebo alone and iliac crest biopsy specimens obtained from patients who were treated with GCs plus placebo or GCs plus risedronate for 1 year. We measured the mass, architecture, and physical and material properties of bone (subject to therapeutic treatments) at nanoscale to macroscopic dimensions, using synchrotron x-ray tomography, elastic modulus mapping, transmission electron microscopy, and small-angle x-ray scattering techniques., Results: GC treatment reduced trabecular bone mass, microarchitecture, and the degree of bone mineralization and elastic modulus within the trabeculae. Concurrent treatment with GCs and risedronate prevented the deterioration of trabecular bone architecture, reduced the degree of mineralization, and preserved elastic modulus within the trabeculae, in both mouse and human bone. In addition, treatment with risedronate plus GCs in mice appeared to preserve bone crystal orientation, compared with treatment with GCs alone., Conclusion: Risedronate prevented the localized changes in mineral and material properties of bone induced by GCs, which may ultimately improve bone strength.

Unlabelled: UV resonance Raman spectroscopy (UVRRS) using 244-nm excitation was used to study the impact of aging on human dentin. The intensity of a spectroscopic feature from the peptide bonds in the collagen increases with tissue age, similar to a finding reported previously for human cortical bone., Introduction: The structural changes that lead to compromised mechanical properties with age in dentin and bone are under intense study. However, in situ analyses of the content and distribution of the mineral phase are more highly developed at present than equivalent probes of the organic phase., Materials and Methods: Thirty-five human molars were divided into three groups: young/normal (23.3 +/- 3.8 years); aged/transparent (74.3 +/- 6.0 years), which had become transparent because of filling of the tubule lumens with mineral deposits; and aged/nontransparent (73.3 +/- 5.7 years). Control experiments were performed by demineralizing normal dentin., Results: Spectral features caused by both the amide backbone and resonance-enhanced side-chain vibrations were observed. This finding contrasts with reported Raman spectra of proteins in solution excited with similar UV wavelengths, where side chain vibrations, but not strong amide features, are observed. The strong intensity of the amide features observed from dentin is attributed to broadening of the resonance profile for the amide pi --> pi* transition caused by the environment of the collagen molecules in dentin. With increasing age, the height of one specific amide vibration (amide I) becomes significantly higher when comparing teeth from donors with an average age of 23 years to those of 73 years (p < 0.001). This trend of increasing amide I peak height with age is similar to that previously reported for human cortical bone. The amide I feature also increased in dentin that had been demineralized and dehydrated., Conclusions: The similar trend of increasing amide I peak height with age in the UVRR spectra of both teeth and bone is surprising, given that only bone undergoes remodeling. However, by considering those observations together with this study of demineralized/dehydrated dentin and our prior work on dentin dehydrated with polar solvents, a consistent relationship between changes in the UVRR spectra and the collagen environment in the tissue can be developed.

Healthy dentin, the mineralized tissue that makes up the bulk of the tooth, is naturally hydrated in vivo; however, it is known that various chemical reagents, including acetone and ethanol, can induce dehydration and thereby affect its properties. Here, we sought to investigate this in light of the effect of alcohol on the mechanical properties of dentin, specifically by measuring the stiffness, strength, and toughness of dentin in simulated body fluid and Scotch whisky. Results indicated that chemical dehydration induced by the whisky had a significant beneficial effect on the elastic modulus, strength, and fracture toughness of dentin. Although this made teeth more resistant to fracture, the change in properties was fully reversible upon rehydration. This effect is considered to be associated with increased cross-linking of the collagen molecules from intermolecular hydrogen-bonding, where water is replaced with weaker hydrogen-bond-forming solvents such as alcohol.

Basic fibroblast growth factor (bFGF), a potent mitogen, has been found to restore trabecular bone mass and connectivity in osteopenic rats. The purpose of this study was to determine how sequential treatment of ovariectomized (OVXed) mice with bFGF followed by risedronate would restore trabecular microarchitecture and improve bone strength through alterations in bone mineralization. Six-month old female Swiss-Webster mice were OVXed or sham-operated and left untreated for 4 weeks to develop osteopenia. At week 5, a group of Sham and OVXed mice were treated with vehicle, and 3 other groups of OVXed mice were treated with bFGF (1 mg/kg daily, s.c., 5x/week) for 3 weeks. At week 8, one group of bFGF-treated mice was sacrificed and the other two bFGF-treated groups were treated with vehicle or risedronate (Ris, 5 microg/kg, s.c., 3x/week) for an additional 6 weeks. Study endpoints included trabecular microarchitecture by microCT, histomorphometry, bone turnover, degree of bone mineralization (DBM), and whole bone strength for the lumbar vertebral body. Compared to sham-operated animals, OVXed mice had significant reductions in trabecular bone volume, connectivity density, DBM, and bone biomechanical properties (P < 0.05). Treatment with bFGF resulted in higher trabecular bone structure and bone strength compared to pre-treatment sham control (P < 0.05). Treatment of OVXed mice with bFGF for 3 weeks followed by 6 weeks Ris maintained the trabecular microarchitecture gained by bFGF treatment, and DBM and bone strength were restored to baseline control levels. Also compared to Sham-operated animals, serum TGF-beta1 was transiently increased after OVX, increased an additional 100% after bFGF withdrawal, and decreased by 30% with risedronate. In addition, DBM was the strongest predictor for bone biomechanical properties (R2 > 0.7, P < 0.001). Serum TGF-beta1 correlated with bone turnover (DPD/Cr, osteocalcin) and was negatively correlated to DBM. Thus, in osteopenic mice, sequential treatment with bFGF followed by risedronate increased trabecular bone microarchitecture, DBM, and bone strength. In addition, suppression of the serum TGF-beta1 with risedronate was associated with increased DBM. Therefore, sequential treatment with bFGF and Ris restores trabecular architecture and allows mineralization of bone to increase, which appears to be beneficial to bone strength.

The adverse effects of stress shielding from the use of high-modulus metallic alloy bio-implant materials has led to increased research into developing polymer-ceramic composite materials that match the elastic modulus of human bone. Of particular interest are poly-l-lactic acid- hydroxyapatite (PLA/HA)-based composites which are fully resorbable in vivo. However, their bioresorbability has a deleterious effect on the mechanical properties of the implant. The purpose of this study is to investigate, from a micromechanistic perspective, the in vitro degradation behavior of such composites manufactured using a simple hot-pressing route for two different hydroxyapatite particles: a fine-grained (average particle size ∼5 μm) commercial powder or coarser whiskers (∼25-30 μm long, ∼5 μm in diameter). We observed that composites with ceramic contents ranging between 70 and 85 wt.% have mechanical properties that match reasonably those of human cortical bone. However, the properties deteriorate with immersion in Hanks' Balanced Salt Solution due to the degradation of the polymer phase. The degradation is more pronounced in samples with larger ceramic content due to the dissolution of the smaller amount of polymer between the ceramic particles.

Data for fracture in human humeral cortical bone are re-analyzed to assess the validity for this material of linear-elastic fracture mechanics (LEFM), which is the standard method of analyzing toughness and one basis for analyzing clinical data relating to bone quality. A nonlinear fracture model, which is based on representing the damage zone in the bone by a cohesive model, is calibrated against a number of sets of test data for normal (not diseased or aged) human cortical bone taken from cadavers. The data consist of load vs. load-point displacement measurements from standard compact-tension fracture tests. Conventional LEFM is unable to account for the shape of the load-displacement curves, but the nonlinear model overcomes this deficiency. Calibration of the nonlinear model against one data curve leads to predictions of the peak load and the displacement to peak load for two other data curves that are, for this limited test set, more accurate than those made using LEFM. Furthermore, prior observations of damage mechanisms in bone are incompatible with the modeling assumption of LEFM that all nonlinearity is confined to a zone much smaller than the specimen and the crack length. The predictions of the cohesive model and the prior observations concur that the length of the nonlinear zone in human cortical bone varies in the range 3-10 mm, which is comparable to or larger than naturally-occurring bones and the specimens used to test them. We infer that LEFM is not an accurate model for cortical bone. The fracture toughness of bone deduced via LEFM from test data will not generally be a material constant, but will take different values for different crack lengths and test configurations. The accuracy of using LEFM or single-parameter fracture toughness for analyzing the significance of data from clinical studies is called into question. The nonlinear cohesive zone model is proposed to be a more accurate model of bone and the traction-displacement or cohesive law is hypothesized to be a material property. The cohesive law contains a more complete representation of the mechanics of material failure than the single-parameter fracture toughness and may therefore provide a superior measure of bone quality, e.g., for assessing the efficacy of therapy for osteoporosis.

Recently published data for fracture in human humeral cortical bone are analyzed using cohesive-zone models to deal with the nonlinear processes of material failure. Such models represent the nonlinear deformation processes involved in fracture by cohesive tractions exerted by the failing material along a fracture process zone, rather than attributing all damage to a process occurring at a single point, as in conventional linear-elastic fracture mechanics (LEFM). The relationship between the tractions and the net displacement discontinuity across the process zone is hypothesized to be a material property for bone. To test this hypothesis, the cohesive law was evaluated by analyzing published load vs. load-point displacement data from one laboratory; the calibrated law was then used to predict similar data taken for a different source of bone using a different specimen geometry in a different laboratory. Further model calculations are presented to illustrate more general characteristics of the nonlinear fracture of bone and to demonstrate in particular that LEFM is not internally consistent for all cases of interest. For example, the fracture toughness of bone deduced via LEFM from test data is not necessarily a material constant, but will take different values for different crack lengths and test configurations. LEFM is valid when the crack is much longer than a certain length scale, representative of the length of the process zone in the cohesive model, which for human cortical bone ranges from 3 to 10mm. Since naturally occurring bones and the specimens used to test them are not much larger than this dimension for most relevant orientations, it is apparent that only nonlinear fracture models can give an internally consistent account of their fracture. The cohesive law is thus a more complete representation of the mechanics of material failure than the single-parameter fracture toughness and may therefore provide a superior measure of bone quality. The analysis of fracture data also requires proper representation of the approximately orthotropic elasticity of the bone specimen; if the specimen is incorrectly assumed to be isotropic, the initial measured compliance cannot be reproduced to within a factor of four and the fracture toughness deduced from the measured work of fracture will be overestimated by approximately 30%.

Unlabelled: This study compares changes in bone microstructure in 6-month-old male GC-treated and female ovariectomized mice to their respective controls. In addition to a reduction in trabecular bone volume, GC treatment reduced bone mineral and elastic modulus of bone adjacent to osteocytes that was not observed in control mice nor estrogen-deficient mice. These microstructural changes in combination with the macrostructural changes could amplify the bone fragility in this metabolic bone disease., Introduction: Patients with glucocorticoid (GC)-induced secondary osteoporosis tend to fracture at higher bone mineral densities than patients with postmenopausal osteoporosis. This suggests that GCs may alter bone material properties in addition to BMD and bone macrostructure., Materials and Methods: Changes in trabecular bone structure, elastic modulus, and mineral to matrix ratio of the fifth lumbar vertebrae was assessed in prednisolone-treated mice and placebo-treated controls for comparison with estrogen-deficient mice and sham-operated controls. Compression testing of the third lumbar vertebrae was performed to assess whole bone strength., Results: Significant reductions in trabecular bone volume and whole bone strength occurred in both prednisolone-treated and estrogen-deficient mice compared with controls after 21 days (p < 0.05). The average elastic modulus over the entire surface of each trabecula was similar in all the experimental groups. However, localized changes within the trabeculae in areas surrounding the osteocyte lacunae were observed only in the prednisolone-treated mice. The size of the osteocyte lacunae was increased, reduced elastic modulus around the lacunae was observed, and a "halo" of hypomineralized bone surrounding the lacunae was observed. This was associated with reduced (nearly 40%) mineral to matrix ratio determined by Raman microspectroscopy. These localized changes in elastic modulus and bone mineral to matrix ratio were not observed in the other three experimental groups., Conclusions: Based on these results, it seems that GCs may have direct effects on osteocytes, resulting in a modification of their microenvironment. These changes, including an enlargement of their lacunar space and the generation of a surrounding sphere of hypomineralized bone, seem to produce highly localized changes in bone material properties that may influence fracture risk.

Limited-sized transplants of culture-expanded autologous or allogeneic bone marrow stromal cells (BMSCs) form cortico-cancellous bone in rodent models. Initiation of clinical studies using autologous BMSC transplantation requires effective bone formation among sizable transplants in a large animal model as well as noninvasive techniques for evaluating transplant success. Here, we obtained bone marrow from the femurs of six dogs and expanded BMSCs in tissue culture. Autologous BMSC-hydroxyapatite/tricalcium phosphate (HA/TCP) transplants were introduced into critical-sized calvarial defects and contralateral control skull defects received HA/TCP vehicle alone. At intervals ranging from 2 to 20 months, transplants were biopsied or harvested for histological and mechanical analysis. Noninvasive studies, including quantitative computed tomography scans and ultrasound, were simultaneously obtained. In all animals, BMSC-containing transplants formed significantly more bone than their control counterparts. BMSC-associated bone possessed mechanical properties similar to the adjacent normal bone, confirmed by both ultrasound and ex vivo analysis. Evaluation by quantitative computed tomography confirmed that the extent of bone formation demonstrated by histology could be discerned through noninvasive means. These results show that autologous cultured BMSC transplantation is a feasible therapy in clinical-sized bone defects and that such transplants can be assessed noninvasively, suggesting that this technique has potential for use in patients with certain bone defects.

Materials that are strong, ultralightweight, and tough are in demand for a range of applications, requiring architectures and components carefully designed from the micrometer down to the nanometer scale. Nacre, a structure found in many molluscan shells, and bone are frequently used as examples for how nature achieves this through hybrid organic-inorganic composites. Unfortunately, it has proven extremely difficult to transcribe nacre-like clever designs into synthetic materials, partly because their intricate structures need to be replicated at several length scales. We demonstrate how the physics of ice formation can be used to develop sophisticated porous and layered-hybrid materials, including artificial bone, ceramic-metal composites, and porous scaffolds for osseous tissue regeneration with strengths up to four times higher than those of materials currently used for implantation.

An understanding of how fatigue cracks grow in bone is of importance as fatigue is thought to be the main cause of clinical stress fractures. This study presents new results on the fatigue-crack growth behavior of small surface cracks (approximately 75-1000 microm in size) in human cortical bone, and compares their growth rates with data from other published studies on the behavior of both surface cracks and many millimeter, through-thickness large cracks. Results are obtained with a cyclically loaded cantilever-beam geometry using optical microscopy to examine for crack growth after every 100-500 cycles. Based on the current and previous results, small fatigue cracks appear to become more resistant to fatigue-crack growth with crack extension, analogous to the way the fracture resistance of cortical bone increases with crack growth. Mechanistically, a theory attributing such behavior to the development of bridges in the wake of the crack with crack growth is presented. The existence of such bridges is directly confirmed using optical microscopy.

The characteristic toughness and strength of bone result from the nature of bone matrix, the mineralized extracellular matrix produced by osteoblasts. The mechanical properties and composition of bone matrix, along with bone mass and architecture, are critical determinants of a bone's ability to resist fracture. Several regulators of bone mass and architecture have been identified, but factors that regulate the mechanical properties and composition of bone matrix are largely unknown. We used a combination of high-resolution approaches, including atomic-force microscopy, x-ray tomography, and Raman microspectroscopy, to assess the properties of bone matrix independently of bone mass and architecture. Properties were evaluated in genetically modified mice with differing levels of TGF-beta signaling. Bone matrix properties correlated with the level of TGF-beta signaling. Smad3+/- mice had increased bone mass and matrix properties, suggesting that the osteopenic Smad3-/- phenotype may be, in part, secondary to systemic effects of Smad3 deletion. Thus, a reduction in TGF-beta signaling, through its effector Smad3, enhanced the mechanical properties and mineral concentration of the bone matrix, as well as the bone mass, enabling the bone to better resist fracture. Our results provide evidence that bone matrix properties are controlled by growth factor signaling.

It is known that fractures are more likely to occur in altered teeth, particularly following restoration or endodontic repair; consequently, it is important to understand the structure of altered forms of dentin, the most abundant tissue in the human tooth, in order to better define the increased propensity for such fractures. Transparent (or sclerotic) dentin, wherein the dentinal tubules become occluded with mineral as a natural progressive consequence of aging, is one such altered form. In the present study, high-resolution transmission electron microscopy is used to investigate the effect of aging on the mineral phase of dentin. Such studies revealed that the intertubular mineral crystallites were smaller in transparent dentin, and that the intratubular mineral (larger crystals deposited within the tubules) was chemically similar to the surrounding intertubular mineral. Exit-wave reconstructed lattice-plane images suggested that the intratubular mineral had nanometer-size grains. These observations support a "dissolution and reprecipitation" mechanism for the formation of transparent dentin.

Micromechanical models for fracture initiation that incorporate local failure criteria have been widely developed for metallic and ceramic materials; however, few such micromechanical models have been developed for the fracture of bone. In fact, although the fracture event in "hard" mineralized tissues such as bone is commonly believed to be locally strain-controlled, only recently has there been experimental evidence (using double-notched four-point bend testing) to support this widely held belief. In the present study, we seek to shed further light on the nature of the local cracking events that precede catastrophic fracture in human cortical bone, and to define their relationship to the microstructure. Specifically, numerical computations are reported that demonstrate that the stress and strain states ahead of such a notch are qualitatively similar irrespective of the deformation mechanism (pressure-insensitive plasticity vs. pressure-sensitive microcracking). Furthermore, we use the double-notched test to examine crack-microstructure interactions from a perspective of determining the salient toughening mechanisms in bone and to characterize how these may affect the anisotropy in fracture properties. Based on preliminary micromechanical models of these processes, the relative contributions of various toughening mechanisms are established. In particular, crack deflection and uncracked-ligament bridging are identified as the major mechanisms of toughening in cortical bone.

Many fractures occur in teeth that have been altered, for example restored or endodontically repaired. It is therefore essential to evaluate the structure and mechanical properties of these altered dentins. One such altered form of dentin is transparent (sometimes called sclerotic) dentin, which forms gradually with aging. The present study focuses on differences in the structure and mechanical properties of normal versus transparent dentin. The mineral concentration, as measured by X-ray computed microtomography, was significantly higher in transparent dentin, the elevated concentration being consistent with the closure of the tubule lumens. Crystallite size, as measured by small angle X-ray scattering, was slightly smaller in transparent dentin, although the importance of this finding requires further study. The elastic properties were unchanged by transparency; however, transparent dentin, unlike normal dentin, exhibited almost no yielding before failure. In addition, the fracture toughness was lowered by roughly 20% while the fatigue lifetime was deleteriously affected at high stress levels. These results are discussed in terms of the altered microstructure of transparent dentin.

The age-related deterioration in bone quality and consequent increase in fracture incidence is an obvious health concern that is becoming increasingly significant as the population ages. Raman spectroscopy with deep-ultraviolet excitation (244 nm) is used to measure vibrational spectra from human cortical bone obtained from donors over a wide age range (34-99 years). The UV Raman technique avoids the fluorescence background usually found with visible and near-infrared excitation and, due to resonance Raman effects, is particularly sensitive to the organic component of bone. Spectral changes in the amide I band at 1640 cm(-1) are found to correlate with both donor age and with previously reported fracture toughness data obtained from the same specimens. These results are discussed in the context of possible changes in collagen cross-linking chemistry as a function of age, and are deemed important to further our understanding of the changes in the organic component of the bone matrix with aging., (2005 Society of Photo-Optical Instrumentation Engineers.)

Although fatigue damage in bone induced by cyclic loading has been recognized as a problem of clinical significance, few fracture mechanics based studies have investigated how incipient cracks grow by fatigue in this material. In the present study, in vitro cyclic fatigue experiments were performed in order to quantify fatigue-crack growth behavior in human cortical bone. Crack-growth rates spanning five orders of magnitude were obtained for the extension of macroscopic cracks in the proximal-distal direction; growth-rate data could be well characterized by the linear-elastic stress-intensity range, using a simple (Paris) power law with exponents ranging from 4.4 to 9.5. Mechanistically, to discern whether such behavior results from "true" cyclic fatigue damage or is simply associated with a succession of quasi-static fracture events, cyclic crack-growth rates were compared to those measured under sustained (non-cyclic) loading. Measured fatigue-crack growth rates were found to exceed those "predicted" from the sustained load data at low growth rates ( approximately 3 x 10(-10) to 5 x 10(-7) m/cycle), suggesting that a "true" cyclic fatigue mechanism, such as alternating blunting and re-sharpening of the crack tip, is active in bone. Conversely, at higher growth rates ( approximately 5 x 10(-7) to 3 x 10(-5) m/cycle), the crack-growth data under sustained loads integrated over the loading cycle reasonably predicts the cyclic fatigue data, indicating that quasi-static fracture mechanisms predominate. The results are discussed in light of the occurrence of fatigue-related stress fractures in cortical bone.

Although the propagation of fatigue cracks has been recognized as a problem of clinical significance in dentin, there have been few fracture mechanics-based studies that have investigated this issue. In the present study, in vitro cyclic fatigue experiments were conducted over a range of cyclic frequencies (1-50 Hz) on elephant dentin in order to quantify fatigue-crack growth behavior from the perspective of understanding the mechanism of fatigue in dentin. Specifically, results obtained for crack extension rates along a direction parallel to the dentinal tubules were found to be well described by the stress-intensity range, DeltaK, using a simple Paris power-law approach with exponents ranging from 12 to 32. Furthermore, a frequency dependence was observed for the crack-growth rates, with higher growth rates associated with lower frequencies. By using crack-growth experiments involving alternate cyclic and static loading, such fatigue-crack propagation was mechanistically determined to be the result of a "true" cyclic fatigue mechanism, and not simply a succession of static fracture events. Furthermore, based on the observed frequency dependence of fatigue-crack growth in dentin and observations of time-dependent crack blunting, a cyclic fatigue mechanism involving crack-tip blunting and re-sharpening is proposed. These results are deemed to be of importance for an improved understanding of fatigue-related failures in teeth.

Exposed root surfaces frequently exhibit non-carious notches representing material loss by abrasion, erosion, and/or abfraction. Although a contribution from mechanical stress is often mentioned, no definitive proof exists of a cause-effect relationship. To address this, we examined dimensional changes in dentin subjected to cyclic fatigue in two different pH environments. Human dentin cantilever-beams were fatigued under load control in pH = 6 (n = 13) or pH = 7 (n = 13) buffer, with a load ratio (R = minimum load/maximum load) of 0.1 and frequency of 2 Hz, and stresses between 5.5 and 55 MPa. Material loss was measured at high- and low-stress locations before and after cycling. Of the 23 beams, 7 withstood 1,000,000 cycles; others cracked earlier. Mean material loss in high-stress areas was greater than in low-stress areas, and losses were greater at pH = 6 than at pH = 7, suggesting that mechanical stress and lower pH both accelerate erosion of dentin surfaces.

An understanding of the evolution of toughness is essential for the mechanistic interpretation of the fracture of cortical bone. In the present study, in vitro fracture experiments were conducted on human cortical bone in order to identify and quantitatively assess the salient toughening mechanisms. The fracture toughness was found to rise linearly with crack extension (i.e., rising resistance- or R-curve behavior) with a mean crack-initiation toughness, K0 of approximately 2 MPa square root m for crack growth in the proximal-distal direction. Uncracked ligament bridging, which was observed in the wake of the crack, was identified as the dominant toughening mechanism responsible for the observed R-curve behavior. The extent and nature of the bridging zone was examined quantitatively using multi-cutting compliance experiments in order to assess the bridging zone length and estimate the bridging stress distribution. Additionally, time-dependent cracking behavior was observed at stress intensities well below those required for overload fracture; specifically, slow crack growth occurred at growth rates of approximately 2 x 10(-9) m/s at stress intensities approximately 35% below the crack-initiation toughness. In an attempt to measure slower growth rates, it was found that the behavior switched to a regime dominated by time-dependent crack blunting, similar to that reported for dentin; however, such blunting was apparent over much slower time scales in bone, which permitted subcritical crack growth to readily take place at higher stress intensities.

Focused ion-beam (FIB) milling is a commonly used technique for transmission electron microscopy (TEM) sample preparation of inorganic materials. In this study, we seek to evaluate the FIB as a TEM preparation tool for human dentin. Two particular problems involving dentin, a structural analog of bone that makes up the bulk of the human tooth, are examined. Firstly, the process of aging is studied through an investigation of the mineralization in 'transparent' dentin, which is formed naturally due to the filling up of dentinal tubules with large mineral crystals. Next, the process of fracture is examined to evaluate incipient events that occur at the collagen fiber level. For both these cases, FIB-milling was able to generate high-quality specimens that could be used for subsequent TEM examination. The changes in the mineralization suggested a simple mechanism of mineral 'dissolution and reprecipitation', while examination of the collagen revealed incipient damage in the form of voids within the collagen fibers. These studies help shed light on the process of aging and fracture of mineralized tissues and are useful steps in developing a framework for understanding such processes.

Although healthy dentin is invariably hydrated in vivo, from a perspective of examining the mechanisms of fracture in dentin, it is interesting to consider the role of water hydration. Furthermore, it is feasible that exposure to certain polar solvents, e.g., those found in clinical adhesives, can induce dehydration. In the present study, in vitro deformation and fracture experiments, the latter involving a resistance-curve (R-curve) approach (i.e., toughness evolution with crack extension), were conducted in order to assess changes in the constitutive and fracture behavior induced by three common solvents-acetone, ethanol and methanol. In addition, nanoindentation-based experiments were performed to evaluate the deformation behavior at the level of individual collagen fibers and ultraviolet Raman spectroscopy to evaluate changes in bonding. The results indicate a reversible effect of chemical dehydration, with increased fracture resistance, strength, and stiffness associated with lower hydrogen bonding ability of the solvent. These results are analyzed both in terms of intrinsic and extrinsic toughening phenomena to further understand the micromechanisms of fracture in dentin and the specific role of water hydration.

Age-related deterioration of the fracture properties of bone, coupled with increased life expectancy, is responsible for increasing incidence of bone fracture in the elderly, and hence, an understanding of how its fracture properties degrade with age is essential. The present study describes ex vivo fracture experiments to quantitatively assess the effect of aging on the fracture toughness properties of human cortical bone in the longitudinal direction. Because cortical bone exhibits rising crack-growth resistance with crack extension, unlike most previous studies, the toughness is evaluated in terms of resistance-curve (R-curve) behavior, measured for bone taken from wide range of age groups (34-99 years). Using this approach, both the ex vivo crack-initiation and crack-growth toughness are determined and are found to deteriorate with age; the initiation toughness decreases some 40% over 6 decades from 40 to 100 years, while the growth toughness is effectively eliminated over the same age range. The reduction in crack-growth toughness is considered to be associated primarily with a degradation in the degree of extrinsic toughening, in particular, involving crack bridging in the wake of the crack.

Two major mechanisms that could potentially be responsible for toughening in mineralized tissues, such as bone and dentin, have been identified-microcracking and crack bridging. While evidence has been reported for both mechanisms, there has been no consensus thus far on which mechanism plays the dominant role in toughening these materials. In the present study, we seek to present definitive experimental evidence supporting crack bridging, rather than microcracking, as the most significant mechanism of toughening in cortical bone and dentin. In vitro fracture toughness experiments were conducted to measure the variation of the fracture resistance with crack extension [resistance-curve (R-curve) behavior] for both materials with special attention paid to changes in the sample compliance. Because these two toughening mechanisms induce opposite effects on the sample compliance, such experiments allow for the definitive determination of the dominant toughening mechanism, which in the present study was found to be crack bridging for microstructurally large crack sizes. The results of this work are of relevance from the perspective of developing a micromechanistic framework for understanding fracture behavior of mineralized tissue and in predicting failure in vivo.

Human dentin is susceptible to failure under repetitive cyclic-fatigue loading. This investigation seeks to address the paucity of data that reliably quantify this phenomenon. Specifically, the effect of alternating vs. mean stresses, characterized by the stress- or load-ratio R (ratio of minimum-to-maximum stress), was investigated for three R values (-1, 0.1, and 0.5). Dentin was observed to be prone to fatigue failure under cyclic stresses, with susceptibility varying, depending upon the stress level. The "stress-life" (S/N) data obtained are discussed in the context of constant-life diagrams for fatigue failure. The results provide the first fatigue data for human dentin under tension-compression loading and serve to map out safe and unsafe regimes for failure over a wide range of in vitro fatigue lives (< 10(3) to > 10(6) cycles).

Few studies have focused on a description of the fracture toughness properties of dentin in terms of resistance-curve (R-curve) behavior, i.e., fracture resistance increasing with crack extension, particularly in light of the relevant toughening mechanisms involved. Accordingly, in the present study, fracture mechanics based experiments were conducted on elephant dentin in order to determine such R-curves, to identify the salient toughening mechanisms and to discern how hydration may affect their potency. Crack bridging by uncracked ligaments, observed directly by microscopy and X-ray tomography, was identified as a major toughening mechanism, with further experimental evidence provided by compliance-based experiments. In addition, with hydration, dentin was observed to display significant crack blunting leading to a higher overall fracture resistance than in the dehydrated material. The results of this work are deemed to be of importance from the perspective of modeling the fracture behavior of dentin and in predicting its failure in vivo.

Despite substantial clinical interest in the fracture resistance of human dentin, there is little mechanistic information in archival literature that can be usefully used to model such fracture. In fact, although the fracture event in dentin, akin to other mineralized tissues like bone, is widely believed to be locally strain-controlled, there has never been any scientific proof to support this belief. The present study seeks to address this issue through the use of a novel set of in vitro experiments in Hanks' balanced salt solution involving a double-notched bend test geometry, which is designed to discern whether the critical failure events involved in the onset of fracture are locally stress- or strain-controlled. Such experiments are further used to characterize the notion of "plasticity" in dentin and the interaction of cracks with the salient microstructural features. It is observed that fracture in dentin is indeed locally strain-controlled and that the presence of dentinal tubules does not substantially affect this process of crack initiation and growth. The results presented are believed to be critical steps in the development of a micromechanical model for the fracture of human dentin that takes into consideration the influence of both the microstructure and the local failure mode., (Copyright 2003 Wiley Periodicals, Inc.)

Toughening mechanisms based on the presence of collagen fibrils have long been proposed for mineralized biological tissues like bone and dentin; however, no direct evidence for their precise role has ever been provided. Furthermore, although the anisotropy of mechanical properties of dentin with respect to orientation has been suggested in the literature, accurate measurements to support the effect of orientation on the fracture toughness of dentin are not available. To address these issues, the in vitro fracture toughness of dentin, extracted from elephant tusk, has been characterized using fatigue-precracked compact-tension specimens tested in Hank's balanced salt solution at ambient temperature, with fracture paths perpendicular and parallel to the tubule orientations (and orientations in between) specifically being evaluated. It was found that the fracture toughness was lower where cracking occurred in the plane of the collagen fibers, as compared to crack paths perpendicular to the fibers. The origins of this effect on the toughness of dentin are discussed primarily in terms of the salient toughening mechanisms active in this material; specifically, the role of crack bridging, both from uncracked ligaments and by individual collagen fibrils, is considered. Estimates for the contributions from each of these mechanisms are provided from theoretical models available in the literature.

Although human dentin is known to be susceptible to failure under repetitive cyclic fatigue loading, there are few reports in the literature that reliably quantify this phenomenon. This study seeks to address the paucity of fatigue data through a systematic investigation of the effects of prolonged cyclical loading on human dentin in an environment of ambient temperature Hank's balanced salt solution (HBSS) at cyclic frequencies of 2 and 20 Hz. The "stress-life" (S/N) data thus obtained are discussed in the context of possible mechanisms of fatigue damage and failure in this material. In addition, stiffness loss data collected in situ during the S/N tests are used to deduce crack velocities and the thresholds for such cracking. These results are presented in a fracture mechanics context as plots of fatigue-crack propagation rates (da/dN) as a function of the stress-intensity range (Delta K). Such S/N and da/dN-Delta K data are discussed in light of the development of a framework for a fracture-mechanics-based methodology for the prediction of the fatigue life of teeth. It is concluded that the presence of small (on the order of 250 microm) incipient flaws in human teeth will not radically affect their useful life., (Copyright 2003 Wiley Periodicals, Inc.)

The in vitro fracture toughness of human dentin has been reported to be of the order of 3 MPa (square root)m. This result, however, is based on a single study for a single orientation, and furthermore involves notched, rather than fatigue precracked, test samples. The present study seeks to obtain an improved, lower-bound, value of the toughness, and to show that previously reported values may be erroneously high because of the absence of a sharp crack as the stress concentrator. Specifically, the average measured critical stress intensity, K(c), for the onset of unstable fracture along an orientation perpendicular to the long axis of the tubules in dentin is found to be 1.8 MPa (square root)m in simulated body fluid (Hanks' balanced salt solution), when tested in a three-point bending specimen containing a (nominally) atomically sharp precrack generated during prior fatigue cycling. This is to be compared with a value of 2.7 MPa (square root)m measured under identical experimental conditions except that the bend specimen contained a sharp machined notch (of root radius 30-50 microm). The effect of acuity of the precrack on the fracture toughness of human dentin is discussed in the context of these data., (Copyright 2003 Wiley Periodicals, Inc.)

A mechanistic understanding of fracture in human bone is critical to predicting fracture risk associated with age and disease. Despite extensive work, a mechanistic framework for describing how the microstructure affects the failure of bone is lacking. Although micromechanical models incorporating local failure criteria have been developed for metallic and ceramic materials, few such models exist for biological materials. In fact, there is no proof to support the widely held belief that fracture in bone is locally strain-controlled, as for example has been shown for ductile fracture in metallic materials. In the present study, we provide such evidence through a novel series of experiments involving a double-notch-bend geometry, designed to shed light on the nature of the critical failure events in bone. We examine how the propagating crack interacts with the bone microstructure to provide some mechanistic understanding of fracture and to define how properties vary with orientation. It was found that fracture in human cortical bone is consistent with strain-controlled failure, and the influence of microstructure can be described in terms of several toughening mechanisms. We provide estimates of the relative importance of these mechanisms, such as uncracked-ligament bridging.