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A rate-dependent microcrack-bridging model that can explain the strain rate dependency of cortical bone apparent yield strength.

Although there are empirical correlations between strain rate, cortical and cancellous bone apparent stiffness, apparent yield strength, apparent ultimate strength and cortical bone fracture toughness, a mechanistic description for these phenomena is lacking. Microcracking is a major mechanism in cortical and cancellous bone failure, however, microdamage content alone cannot explain the strain rate dependence of bone strength without considering time-dependent behavior of the crack. Using a rate-dependent model of a fiber-bridged microcrack and data from the literature, we demonstrate that the experimental apparent yield strength of bone can be predicted directly from measurements of apparent moduli of elasticity of bone constituents and failure strain of the collagenous matrix. Yield strength predictions for estrogen depleted bone were made using the model and data from ovariectomized sheep. It was predicted that the yield strength of estrogen-deficient bone is comparable to that of normal bone within strain rates associated with physiological activities. For high strain rates, however, the strength of estrogen-depleted bone was predicted to be much weaker than normals suggesting a higher fracture risk due to impact from falls, for individuals with estrogen-depleted bones such as in post-menopausal osteoporosis.

Bone and Bones↗

Vascular smooth muscle polyploidization--from mitotic checkpoints to hypertension.

Aging and hypertension are accompanied by an increase in mass and rigidity of arterial walls. At capacitance arteries, the enlargement and stiffness of the medial smooth muscle layer promote systolic hypertension and contribute to left ventricular hypertrophy and cardiovascular morbidity. Morphological studies have demonstrated that vascular smooth muscle cell (VSMC) hypertrophy, with minimal hyperplasia, causes the enlargement of vascular smooth muscle at capacitance arteries, and that VSMC hypertrophy is strongly associated with VSMC polyploidization. Recent studies demonstrate that hypertrophic signals, such as those elicited by Angiotensin II, abrogate the mechanisms of control of M phase in VSMC and induce cell cycle re-entry and polyploidization. These polyploid VSMC have a lower replicative rate, but a higher mass, protein content and matrix production than their diploid counterparts. Both, the protein kinase Aktl and the cyclin kinase-associated protein CKsl, have been implicated in the mechanism of VSMC polyploidization during hypertension. Here, we review the function of these proteins at the mitotic spindle cell cycle checkpoint and their role in the process of VSMC polyploidization.

Adaptor Proteins, Signal Transducing↗

Dynamic deformational loading results in selective application of mechanical stimulation in a layered, tissue-engineered cartilage construct.

The application of dynamic physiologic loading to a bilayered chondrocyte-seeded agarose construct with a 2% (wt/vol) top layer and 3% (wt/vol) bottom layer was hypothesized to (1) improve overall construct properties and (2) result in a tissue that mimics the mechanical inhomogeneity of native cartilage. Dynamic loading over the 28 day culture period was found to significantly increase bulk mechanical and biochemical properties versus free-swelling culture. The initial depth-distribution of the compressive Young's modulus (EY) reflected the intrinsic properties of the gel in each layer and a similar trend to the native tissue, with a softer 2% gel layer and a much stiffer 3% gel layer. After 28 days in culture, free-swelling conditions maintained this general trend while loaded constructs possessed a reverse profile, with significant increases in EY observed only in the 2% gel. Histological analysis revealed preferential matrix formation in the 2% agarose layer, with matrix localized more pericellularly in the 3% agarose layer. Finite element modeling revealed that, prior to significant matrix elaboration, the 2% layer experiences increased mechanical stimuli (fluid flow and compressive strain) during loading that may enhance chondrocyte stimulation and nutrient transport in that layer, consistent with experimental observations. From these results, we conclude that due to the limitations in 3% agarose, the use of this type of bilayered construct to construct depth-dependent inhomogeneity similar to the native tissue is not likely to be successful under long-term culture conditions. Our study underscores the importance of other physical properties of the scaffold that may have a greater influence on interconnected tissue formation than intrinsic scaffold stiffness.

Animals↗

Brittle bones in spontaneously diabetic female rats cannot be predicted by bone mineral measurements: studies in diabetic and ovariectomized rats.

Spontaneously diabetic BB rats were sham operated (SO) or ovariectomized (OVX) within days after onset and studied after 4, 8, and 12 weeks. Analyses included histomorphometry of proximal tibial metaphyses, biochemical analyses of humeri, DXA analyses, and biomechanical testing of femora. In SO diabetic rats, no osteoblasts, osteoid tissue, or osteoclasts were present on the trabecular bone surface, but trabecular bone volume (TBV) remained normal compared with control BB rats. The concentration of IGF-I per dry weight of humerus was decreased after 12 weeks of diabetes, whereas the concentrations of calcium and osteocalcin did not change. DXA analysis showed normal bone mineral density (BMD) at both diaphyseal and metaphyseal femoral areas. On biomechanical testing, angular deformation, energy absorption, and torsional strength of the femora were decreased after 8-12 weeks of diabetes, but stiffness was normal. Ovariectomy in diabetic rats caused a decrease in femoral BMD especially at the metaphysis, and there was a trend toward decreased TBV in the tibial metaphysis; TBV loss was less marked than in control OVX rats, however. The increase in BMD at the femoral diaphysis, measured after 12 weeks of OVX in control rats, was absent in diabetic rats. Multiple-regression analysis indicated that the presence of diabetes but not ovariectomy, weight, and mineral content correlated with decreased energy absorption, angular deformation, and strength of the femora. The data infer that the (near) absence of unmineralized bone matrix in severely diabetic rats alters bone microarchitecture and ultimately results in brittle bones, which is not predicted by BMC or BMD measurements.

Absorptiometry, Photon↗

Evaluation of compliance and stiffness of decellularized tissues as scaffolds for tissue-engineered small caliber vascular grafts using intravascular ultrasound.

This study evaluated the compliance and stiffness of decellularized canine common carotid artery as well as decellularized canine ureter and compared it with that of polytetrafluoroethylene, elastin gel combined with polylactic acid tube, and canine common carotid artery. To calculate the compliance and stiffness, internal diameters and cross-sectional areas were measured according to changes in the intraluminal pressures using intravascular ultrasound in a closed circuit system equipped with a syringe pump. The pressure-area curve, stiffness parameter beta, and diameter compliance were evaluated. Canine common carotid artery and decellularized canine common carotid artery, as well as decellularized canine ureter, showed a compliant response, a J-shaped curve. However, the latter evidenced different characteristics in the low pressure range. Although the cross-sectional area of the elastin gel combined with polylactic acid tube showed some changes, it did not present a J-shape curve. Polytetrafluoroethylene exhibited a noncompliant response.The results in this study have shown that the compliance in the decellularized matrices was maintained after cell extraction, which demonstrated the importance of the remaining matrix structure in the mechanical properties of decellularized tissue. A clear difference between the decellularized matrices and synthetic materials was noted in terms of the compliance, even in materials composed of relatively elastic materials.

Animals↗

[An immunosorbent for interferon purification].

Immunoaffinity chromatography was shown to be the method achieving the most complete elimination of antigenic admixtures from leukocyte interferon preparations without the loss of the preparation activity. An affinity sorbent has been developed on the basis of covalently linked polyvinyl alcohol (PVA). The immobilization of the antigen-specific rabbit globulin in the preparation of the sorbent is achieved by reaction of protein amino groups with the activated matrix. The proposed sorbent achieved the elimination of the antigenic admixtures from interferon preparations as effectively as those prepared on the basis of sepharose 4B, the productivity of the purification process being at least 5 times higher. The proposed sorbent is stable at the limit values of rH, is not destroyed by detergents, is sterilized in the process of preparation. Owing to the strong linkage of the immobilized immunoglobulin with the PVA-carrier, immunoaffinity chromatography on this sorbent does not involve contamination of the preparations with rabbit globulin allergenic for man. The combination of a large pore structure, wetting ability, stiffness, mechanical and chemical stability allows the proposed sorbent to be recommended for use in modern large-scale biotechnological production.

Animals↗

A composites theory predicts the dependence of stiffness of cartilage culture tissues on collagen volume fraction.

The tensile stiffness of tissue grown from chondrocyte culture was both measured experimentally and predicted using a composites model theory relating tissue microstructure to macroscopic material stiffness. The tissue was altered by several treatment protocols to provide a wide range of collagen fibril volume fraction (0.015-0.15). The rate of change of tissue modulus with change in collagen volume fraction predicted by the theory was within 14% of the slope of the linear fit through the experimental data, without the use of fitting parameters for the theoretical value of the slope. Use of the model to simulate cytokine mediated tissue digestion suggests that the action of IL-1beta and retinoic acid is mainly removal of proteoglycans and some removal of collagen. The model also indicates that the matrix and collagen remaining in the tissue has the same elastic properties as the untreated tissue, and is not damaged due to the alteration. Young's modulus of the collagen fibrils is predicted to be 120 MPa, a value in the range of previous studies. This value is dependent mainly on the matrix modulus and collagen fibril volume fraction and not on Poisson's ratio of either matrix or fibril. Poisson's ratio of the tissue depends primarily on the Poisson's ratio of the matrix.

Animals↗

In vivo effects of stromelysin on the composition and physical properties of rabbit articular cartilage in the presence and absence of a synthetic inhibitor.

OBJECTIVE: To characterize the effects of intraarticular injection of recombinant human stromelysin (SLN) on the matrix composition and physical properties of cartilage from lapine stifle joints and the modulation of these effects by the systemic administration of an N-carboxyalkyl synthetic matrix metalloproteinase inhibitor, L-696,418. METHODS: Female 6-8-week-old New Zealand white rabbits received an intraarticular injection of 100 micrograms activated SLN in 1 stifle joint and buffer in the contralateral control knee; these animals were killed after 1 hour. A separate group of animals received an intravenous injection of either 30 mg/kg L-696,418 or buffer prior to intraarticular injection of SLN. Joints were dissected and analyzed for proteoglycan (PG) loss into joint fluid, tissue biochemical composition, and histology by toluidine blue or anti-VDIPEN antibody staining, or were frozen for physical property analysis. Disks of femoropatellar groove cartilage were harvested from the stifle joint and tested in uniaxially confined compression for determination of electromechanical and mechanical properties. RESULTS: Lapine stifle joints that received injection of SLN without systemic administration of L-696,418 showed a 13-fold increase in loss of PG into synovial fluid. Cartilage from these joints showed significant decreases in streaming potential at 1 Hz and electrokinetic coupling coefficient, but no change in equilibrium modulus, dynamic stiffness, or hydraulic permeability. Systemic treatment with L-696,418 resulted in a significant decrease in loss of PG into joint fluid and elimination of changes in cartilage high-frequency streaming potential and coupling coefficient in joints that were injected with SLN. CONCLUSION: The 1-hour exposure to SLN in vivo resulted in loss of PG and exposure of the VDIPEN epitope of the aggrecan core protein in the superficial region of the tissue near the articular surface. This highly localized degradation resulted in electromechanical behavior changes, but little or no change occurred in mechanical properties. Systemic administration of L-696,418 significantly decreased loss of PG from cartilage and prevented the highly localized tissue degradation and the resultant changes in electromechanical behavior caused by intraarticular SLN injection.

Animals↗

Manipulation of remodeling pathways to enhance the mechanical properties of a tissue engineered blood vessel.

There is a current need for a small diameter vascular graft due to the limited supply of autogenous grafts and the failure of synthetic grafts due to thrombosis and/or intimal hyperplasia. The use of living cells and tissues to fabricate a small diameter graft (i.e., tissue engineered blood vessel, TEBV) could be useful given the endothelialization potential and biocompatibility benefits of such a graft. However, while sufficient strength has been attained in a TEBV, coordinate compliance has yet to be fine-tuned. In this study we investigate the effects of biological response modifiers, retinoic acid (RA) and ascorbic acid (AA) on TEBV biomechanics as a function of time and subsequently correlate observed RA/AA induced changes in TEBV mechanics with alterations in smooth muscle cell (SMC) biochemistry. TEBVs were constructed using a fibrillar type I collagen network populated by human aortic smooth muscle cells (AoSMC). Following construction this TEBV was treated with 0.3 mM AA and 0.1 mM RA (concentrations found to induce changes in VSMC phenotype). Ultimate tensile stress (UTS), rate of relaxation (RR) and elastic efficiency (EE) of RA/AA treated and untreated TEBVs were measured following 1, 7, 15, 30, 45, and 60 days of treatment. At corresponding time points, the effect of these treatments on collagen and elastin protein synthesis and mRNA expression was examined. RA/AA treated TEBV strength increased and stiffness decreased compared to controls as a function of time. Relative collagen synthesis in treated TEBVs exceeded control levels by nearly two-fold at 15 and 30 days of incubation. RA/AA treated collagen gene expression followed a similar trend. Relative elastin synthesis was also greater in treated TEBVs as compared to untreated TEBVs at 15 and 30 days of incubation and correspondingly elastin mRNA expression was significantly elevated at 15 days of incubation. These data provide evidence that RA/AA treated TEBVs exhibit mechanical properties which more closely mimic those of a native vessel than their untreated counterparts and that changes in extracellular matrix composition and matrix gene expression in the presence of RA/AA treatment may play an important role in the development of said mechanical properties.

Animals↗

Prevention of aortic and cardiac fibrosis by spironolactone in old normotensive rats.

OBJECTIVES: Because the synthesis of aldosterone is mainly modulated by angiotensin II through type I receptor stimulation and because converting enzyme inhibition (CEI) does not modify aortic extracellular matrix in old normotensive rats, the aim of the present study was to determine whether inhibition of aldosterone formation was able to prevent aortic fibrosis in old Sprague-Dawley normotensive rats. BACKGROUND: We have previously shown that long-term aldosterone antagonism prevents the age-related increase in aortic collagen accumulation in young spontaneously hypertensive rats, independent of blood pressure changes. In contrast, we reported that the positive effects of CEI in the prevention of aortic collagen accumulation were related to the inhibition of angiotensin II actions on angiotensin II type I receptors. METHODS: For this purpose, we studied the histomorphometric and stiffness (echo-tracking technique) changes of an eight-week treatment with the aldosterone antagonist spironolactone by comparison with placebo. RESULTS: At the end of treatment, spironolactone in conscious animals did not change intra-arterial blood pressure, aortic and carotid wall thickness, and cardiac weight. Cardiac collagen density and, to a lesser extent, carotid collagen and elastin densities and contents were significantly decreased in association with an increase of carotid distensibility. CONCLUSIONS: These results show that in old normotensive rats, spironolactone can markedly prevent cardiac and, to a lesser extent, arterial fibrosis and improve arterial stiffness, despite a lack of hypotensive effect.

Age Factors↗

Maternal dietary lipids alter bone chemical composition, mechanical properties, and histological characteristics of progeny of Japanese quail.

This study evaluated the effects of maternal dietary lipids on chemical components and mechanical and histological properties of tibia in progeny of Japanese quail fed different dietary lipids. Laying hens were fed a basal diet containing either soybean oil (SBO), hydrogenated soybean oil (HSBO), chicken fat (CF), or menhaden fish oil (FO) at 50 g/kg of the diet. The various maternal dietary lipid treatments did not affect growth of progeny at any developmental stage. There were no differences in tibial length, diameter, or collagen content. Tibial percentage ash was significantly higher in newly hatched progeny from hens fed the FO and HSBO diets. The levels of tibial deoxypyridinoline and total pyridinium crosslinks were higher in the FO and HSBO groups at hatch. At 7 d of age, the tibial deoxypyridinoline links remained higher in the FO group compared to the CF and SBO groups. Likewise, progeny from hens consuming the FO or HSBO diet had higher tibial shear force and stiffness at 7 and 14 d of age. There were no pronounced differences in tibial fracture energy and deflection among treatments. Maternal FO or HSBO enlarged the cartilaginous proliferative and hypertrophic zones of the tibial proximal end in newly hatched quail, which was accompanied by a thicker cortical bone in the diaphysis. However, the width of the hypertrophic zones tended to be smaller in these two groups coupled with improvement in trabecular density and cortical thickness in the proximal end and cortical density in the diaphysis at 3 wk of age. These results suggest that maternal dietary lipids altered bone development by influencing organic matrix quality and mineralization in embryos.

Adipose Tissue↗

Mechanisms of stiffening and strengthening in media-equivalents fabricated using glycation.

We have recently reported that glycation can be exploited to increase the circumferential tensile stiffness and ultimate tensile strength of media-equivalents (MEs) and increase their resistance to collagenolytic degradation, all without loss of cell viability (Girton et al., 1999). The glycated MEs were fabricated by entrapping high passage adult rat aorta SMCs in collagen gel made from pepsin-digested bovine dermal collagen, and incubated for up to 10 weeks in complete medium with 30 mM ribose added. We report here on experiments showing that ME compaction due to traction exerted by the SMCs with consequent alignment of collagen fibrils was necessary to realize the glycation-mediated stiffening and strengthening, but that synthesis of extracellular matrix constituents by these cells likely contributed little, even when 50 micrograms/ml ascorbate was added to the medium. These glycated MEs exhibited a compliance similar to arteries, but possessed less tensile strength and much less burst strength. MEs fabricated with low rather than high passage adult rat aorta SMCs possessed almost ten times greater tensile strength, suggesting that alternative SMCs sources and biopolymer gels may yield sufficient strength by compositional remodeling prior to implantation in addition to the structural remodeling (i.e., circumferential alignment) already obtained.

Animals↗

Accumulation of in-vivo fatigue microdamage and its relation to biomechanical properties in ageing human cortical bone.

Bone matrix accumulates microdamage in the form of microcracks as a result of everyday cyclic loading activities. In two very recent studies, which used conventional histological stains and light microscopy techniques, the amount of this in-vivo microdamage in the cortices of long bones has been shown to increase with age. These articles have suggested that in-vivo microcracks may have an effect on the material properties of the tissue. However, a precise quantitative relationship between the number of microcracks and the mechanical properties of these same bones has not been produced before, and in particular the way the microcracks may affect the stiffness, the strength or possibly the toughness of the tissue. This article presents an examination of the in-vivo microdamage in human bones by the use of laser scanning confocal microscopy, which offers better discrimination and allows examination of the cracks in-situ. Quantification of in-vivo fatigue microcracks was performed by counting the microcrack numerical density and surface density in specimens for which we have previously derived a full set of mechanical properties as a function of age. It is shown that bone microdamage relates more to the toughness (measured by three different measures) of ageing bone tissue than to its stiffness and strength. The result allows us (i) to re-evaluate the fragility of ageing human bone and put more emphasis on its energy-related resistance to fracture than perhaps on its stiffness or strength and also (ii) to understand more fully the causal relationship and interactions between microcracks and tissue toughness.

Journal Article↗

Accumulation of in-vivo fatigue microdamage and its relation to biomechanical properties in ageing human cortical bone.

Bone matrix accumulates microdamage in the form of microcracks as a result of everyday cyclic loading activities. In two very recent studies, which used conventional histological stains and light microscopy techniques, the amount of this in-vivo microdamage in the cortices of long bones has been shown to increase with age. These articles have suggested that in-vivo microcracks may have an effect on the material properties of the tissue. However, a precise quantitative relationship between the number of microcracks and the mechanical properties of these same bones has not been produced before, and in particular the way the microcracks may affect the stiffness, the strength or possibly the toughness of the tissue. This article presents an examination of the in-vivo microdamage in human bones by the use of laser scanning confocal microscopy, which offers better discrimination and allows examination of the cracks in-situ. Quantification of in-vivo fatigue microcracks was performed by counting the microcrack numerical density and surface density in specimens for which we have previously derived a full set of mechanical properties as a function of age. It is shown that bone microdamage relates more to the toughness (measured by three different measures) of ageing bone tissue than to its stiffness and strength. The result allows us (i) to re-evaluate the fragility of ageing human bone and put more emphasis on its energy-related resistance to fracture than perhaps on its stiffness or strength and also (ii) to understand more fully the causal relationship and interactions between microcracks and tissue toughness.

Adult↗

The use of polylactic acid matrix and periosteal grafts for the reconstruction of rabbit knee articular defects.

In order to find a material that would improve cartilage repair, we investigated the use of porous polylactic acid matrix (PLA) with and without periosteal grafts in large articular defects in the medial femoral condyles of 18 New Zealand white rabbit knees. The right knee defect was filled with PLA, the left defect was filled with PLA and a periosteal graft. All animals were killed at 12 weeks. PLA allowed for the de novo growth of neocartilage at the articular surface in all specimens and appeared to serve as a scaffolding for cell migration and matrix formation. Histologically, small amounts of PLA remained under the neocartilage with the majority being replaced by bone. PLA was a suitable carrier for periosteal grafts with a high graft survival rate (89%) and proliferation of a neocartilage which was thicker and more closely resembled articular cartilage than PLA alone knees. Biochemically, there was more type II collagen in the grafted knees (83%) than in the PLA alone knees (65%). Biomechanical tests of the neocartilage included equilibrium displacement, aggregate modulus, and apparent permeability. These tests were not statistically different between PLA alone and grafted knees. Comparison to normal cartilage indicated that the neocartilage was less stiff but had similar permeability. A consistent repair of the articular defects was achieved with and without periosteal grafts resulting in a tissue that closely resembled hyaline articular cartilage.

Animals↗

Cell surface receptors transmit sufficient force to bend collagen fibrils.

To better understand the dynamic interaction of cells with their surrounding extracellular matrix, chondrocytes and rat embryo fibroblasts were overlaid with individual collagen fibrils and observed with high-resolution video-enhanced differential interference contrast microscopy. Although the cells had a polygonal shape characteristic of nonmotile cells, they used processes usually associated with cell locomotion to acquire the collagen fibrils. Instead of being transported in a retrograde direction, fibrils on the dorsal cell surface were bent, and regions of the bent fibrils were shifted in diverse directions. A blocking antibody to the beta1 integrin subunit significantly inhibited collagen fibril acquisition and bending. Enhanced actin assembly was only occasionally associated with fibrils undergoing rearrangement. Considering that the relatively stiff collagen fibrils require the application of force to be bent, this study shows that cells with a polygonal morphology (as opposed to a polarized, motile shape) are capable of exerting force through the beta1 integrins on the dorsal surface of the cell. Analysis of the bending patterns indicates that fibril buckling was induced by retrograde force combined with regions held stationary and/or the fibrils were bent by forces acting in opposing directions.

Actins↗

Interstitial stress and fluid pressure within a growing tumor.

A solid tumor is composed of a population of cells that is expanding as a result of cell division. With dense cell packing, the solid matrix of the interstitial tissue is subject to residual stress. In addition, elevated interstitial fluid pressure (IFP) has been reported by researchers for a number of solid tumors. These features were incorporated into a mathematical model that predicts the mechanical response of a solid tumor within its host environment. A theoretical framework accounting for volumetric expansion, transvascular exchange and extravascular transport of fluids was developed using poroelastic theory, and applied to a spherical, vascularized, alymphatic tumor undergoing small growth increments. Simulations of tumor IFP were similar to those predicted by Jain and Baxter, showing elevated IFP that is driven by microvascular fluid pressure. Tumor growth, tissue stiffness, and IFP contribute to the compressive stresses predicted in the solid tumor interior. Tensile and compressive stresses were predicted in adjacent host tissues corresponding to radial and circumferential directions, respectively. An application of this model includes a solid stress-based framework for predicting regions of vascular collapse within the tumor interior.

Blood Pressure↗

Osteocytes and bone lining cells: which are the best candidates for mechano-sensors in cancellous bone?

Previously, we have investigated the possible role of osteocytes as mechano-sensors, and mediators of bone turnover. It was found that the proposed regulatory mechanism produced morphologies of trabecular bone, under particular loading conditions, which were consistent with morphogenesis and adaptation as seen in reality. The main objective of this study was to discern whether lining cells or osteoblasts could possibly play a similar role as effectively with regard to their capacity for self-optimization of the trabecular architecture, in terms of a low apparent mass to stiffness ratio. For that purpose the earlier analyses with osteocytes as mechano-sensors, distributed throughout the bone, were repeated for mechano-sensors located at bone surfaces only. Compared to the osteocyte model, the surface cell remodeling algorithm was reluctant to change its architecture, which implies that it is less sensitive to changes in the loading pattern. This resulted in less efficient bone adaptation, which was reflected by a considerably higher relative mass for a similar apparent stiffness in the loading direction. In other words, more mass is needed to obtain an equally stiff structure, at the apparent level, with respect to the externally applied loads. Furthermore, stresses and strains at the tissue level vary across a much wider range, relative to the osteocyte model, where the higher incidence of elevated strains indicates an increased failure risk. Therefore, we conclude that mechanical information at the bone surface may not be sufficient to adequately regulate functional bone adaptation.

Animals↗