The identification of matrix Gla protein in cartilage

The vitamin K-dependent bone protein matrix gamma-carboxyglutamic acid (Gla) protein (MGP) has been identified by radioimmunoassay in the guanidine extract of rat cartilage. MGP was present in all cartilages tested at levels comparable to the MGP level in bone. Western blot analysis indicated that the molecular weight of cartilage MGP is the same as bone MGP, and Northern blot analysis revealed that MGP mRNA from cartilage is the same size as the MGP mRNA from bone. The structurally related vitamin K-dependent protein bone Gla protein could not be detected in cartilage by radioimmunoassay or by Northern blot analysis. The discovery that MGP is synthesized by growth plate cartilage could provide an explanation for the excessive growth plate mineralization disorder seen in rats treated with the vitamin K antagonist warfarin and the punctate mineralization of the growth plate seen in infants whose mothers received warfarin in the first trimester of pregnancy (the fetal warfarin syndrome). Both disorders appear to be caused by the inactivation of a vitamin K-dependent mineralization inhibitor in cartilage, an inhibitor which we suggest is MGP.     

The action of infrasound oscillations on the properties of water and of a DNA solution

The effect of mechanical vibrations (MV) electrical conductivity of water and optical density of aqueous DNA solution. Distilled water was treated with MV of several frequencies from 3 to 5000 Hz with an intensity of 90 dB for 30 minutes. Different sensitivities of water specific electrical conductivity (SEC) were determined (the value of distilled water SEC was 209 +/- 2 microS/m). The greatest decrease of SEC (by 15.7%) under the influence of MV was observed at the frequency of 4 Hz. There was no effect at frequencies higher than 100 Hz. The treatment of DNA water solutions with MV of frequencies 4 and 10 Hz decreased its optical density by 4.2 +/- 1.1 and 4.8 +/- 1.2% correspondingly in comparison with control. In cases of treatment with frequencies of 20 and 50 Hz no effect was observed. The mechanism of MV effect on water can be connected with the changes of system structural characteristics. It is confirmed by experiments with DNA solution, where the decrease of optical density (at 260 nm) under MV treatment is conditioned with the increase of the probability of hydrogen binding formation between the bases.     

Constrained tibial vibration in mice: a method for studying the effects of vibrational loading of bone

Vibrational loading can stimulate the formation of new trabecular bone or maintain bone mass. Studies investigating vibrational loading have often used whole-body vibration (WBV) as their loading method. However, WBV has limitations in small animal studies because transmissibility of vibration is dependent on posture. In this study, we propose constrained tibial vibration (CTV) as an experimental method for vibrational loading of mice under controlled conditions. In CTV, the lower leg of an anesthetized mouse is subjected to vertical vibrational loading while supporting a mass. The setup approximates a one degree-of-freedom vibrational system. Accelerometers were used to measure transmissibility of vibration through the lower leg in CTV at frequencies from 20 Hz to 150 Hz. First, the frequency response of transmissibility was quantified in vivo, and dissections were performed to remove one component of the mouse leg (the knee joint, foot, or soft tissue) to investigate the contribution of each component to the frequency response of the intact leg. Next, a finite element (FE) model of a mouse tibia-fibula was used to estimate the deformation of the bone during CTV. Finally, strain gages were used to determine the dependence of bone strain on loading frequency. The in vivo mouse leg in the CTV system had a resonant frequency of 60 Hz for +/-0.5 G vibration (1.0 G peak to peak). Removing the foot caused the natural frequency of the system to shift from 60 Hz to 70 Hz, removing the soft tissue caused no change in natural frequency, and removing the knee changed the natural frequency from 60 Hz to 90 Hz. By using the FE model, maximum tensile and compressive strains during CTV were estimated to be on the cranial-medial and caudolateral surfaces of the tibia, respectively, and the peak transmissibility and peak cortical strain occurred at the same frequency. Strain gage data confirmed the relationship between peak transmissibility and peak bone strain indicated by the FE model, and showed that the maximum cyclic tibial strain during CTV of the intact leg was 330+/-82microepsilon and occurred at 60-70 Hz. This study presents a comprehensive mechanical analysis of CTV, a loading method for studying vibrational loading under controlled conditions. This model will be used in future in vivo studies and will potentially become an important tool for understanding the response of bone to vibrational loading.     

The pulsed water jet for selective removal of bone cement during revision arthroplasty]

Conventional tools used in prosthetic revision surgery have a limited range of action within the narrow cement mantle. Water jet cutting technology permits tiny and precisely controlled cuts, and may therefore be an alternative method of bone cement removal. Our study compares the cutting performance on bone cement (PMMA) and bone of a pulsed water jet and a continuous water jet. The aim of the study was to establish whether selective removal of PMMA is possible. 55 bone specimens (bovine femora) and 32 specimens of PMMA were cut with a continuous and a pulsed water jet at different pressures (40 MPa, 60 MPa) and pulse frequencies (0Hz, 50Hz, 250Hz). To ensure comparability of the results, the depths of cut were related to the hydraulic power of that part of the jet actually impinging on the material. While for PMMA the power-related depth of cut increased significantly with the pulse frequency, this did not apply to bone. The cuts produced in bone were sharp-edged. Since PMMA is more brittle than bone, the water jet caused cracks that enlarged further until particles of bone broke away. Although selective removal of PMMA without doing damage to the bone was not possible at the investigated settings of the jet parameters, the results do show that a pulsed water jet can cut bone cement much more effectively than bone. This is an important advantage over conventional non-selective tools for the removal of bone cement.     

The frequency window effect of sinusoidal electromagnetic fields in promoting osteogenic differentiation and bone formation involves extension of osteoblastic primary cilia and activation of protein kinase A

Electromagnetic fields (EMFs) have emerged as a versatile means for osteoporosis treatment and prevention. However, its optimal application parameters are still elusive. Here, we optimized the frequency parameter first by cell culture screening and then by animal experiment validation. Osteoblasts isolated from newborn rats (ROBs) were exposed 90 min/day to 1.8 mT SEMFs at different frequencies (ranging from 10 to 100 Hz, interval of 10 Hz). SEMFs of 1.8 mT inhibited ROB proliferation at 30, 40, 50, 60 Hz, but increased proliferation at 10, 70, 80 Hz. SEMFs of 10, 50, and 70 Hz promoted ROB osteogenic differentiation and mineralization as shown by alkaline phosphatase (ALP) activity, calcium content, and osteogenesis-related molecule expression analyses, with 50 Hz showing greater effects than 10 and 70 Hz. Treatment of young rats with 1.8 mT SEMFs at 10, 50, or 100 Hz for 2 months significantly increased whole-body bone mineral density (BMD) and femur microarchitecture, with the 50 Hz group showing the greatest effect. Furthermore, 1.8 mT SEMFs extended primary cilia lengths of ROBs and increased protein kinase A (PKA) activation also in a frequency-dependent manner, again with 50 Hz SEMFs showing the greatest effect. Pretreatment of ROBs with the PKA inhibitor KT5720 abolished the effects of SEMFs to increase primary cilia length and promote osteogenic differentiation/mineralization. These results indicate that 1.8 mT SEMFs have a frequency window effect in promoting osteogenic differentiation/mineralization in ROBs and bone formation in growing rats, which involve osteoblast primary cilia length extension and PKA activation.     

Collagenolytic activity of cathepsin K is specifically modulated by cartilage-resident chondroitin sulfates

Cathepsin K is the predominant cysteine protease in osteoclast-mediated bone remodeling, and the protease is thought to be involved in the pathogenesis of diseases with excessive bone and cartilage resorption. Osteoclastic matrix degradation occurs in the extracellular resorption lacuna and upon phagocytosis within the cell's lysosomal-endosomal compartment. Since glycosaminoglycans (GAGs) are abundant in extracellular matrixes of cartilage and growing bone, we have analyzed the effect of GAGs on the activity of bone and cartilage-resident cathepsins K and L and MMP-1. GAGs, in particular chondroitin sulfates, specifically and selectively increased the stability of cathepsin K but had no effect on cathepsin L and MMP-1. GAGs strongly enhanced the stability and, to a lesser extent, the catalytic activity of cathepsin K. To combine the activity and stability parameters, we defined a novel kinetic term, named cumulative activity (CA), which reflects the total substrate turnover during the life span of the enzyme. In the presence of chondroitin-4-sulfate (C-4S), the CA value increased 200-fold for cathepsin K but only 25-fold with chondroitin-6-sulfate (C-6S). C-4S dramatically increased the hydrolysis of soluble as well insoluble type I and II collagens, whereas the effects of C-6S and hyaluronic acid were less pronounced. C-4S acts in a concentration-dependent manner but reaches saturation at approximately 0.1%, a concentration similar to that found in the synovial fluid of arthritis patients. C-4S increased the cathepsin K-mediated release of hydroxyproline from insoluble type I collagen 10-fold but had only a less than 2-fold enhancing effect on the hydrolysis of intact cartilage. The relatively small increase in the hydrolysis of cartilage by C-4S was attributed to the endogenous chondroitin sulfate content present in the cartilage. Although C-4S increased the pH stability at neutral pH, a significant increase in the collagenolytic activity of cathepsin K at this pH was not observed, thus suggesting that the unique collagenolytic activity of cathepsin K at acidic pH is mechanistically determined and not by the enzyme's instability at neutral pH. The selective and significant stabilization and activation of cathepsin K activity by C-4S may provide a rationale for a novel mechanism to regulate the enzyme's activity during bone growth and aging, two processes known for significant changes in the GAG content.     

Low-magnitude mechanical signals that stimulate bone formation in the ovariectomized rat are dependent on the applied frequency but not on the strain magnitude

There is growing evidence that extremely small mechanical signals, if applied at a sufficiently high frequency, can serve as anabolic signals to bone tissue. To determine if the responsiveness of bone to low-magnitude, high-frequency parameters is modulated by endocrine imbalance, ovariectomized (OVX) Sprague-Dawley rats were subjected to whole body vibrations (WBV, 0.15 g) at 45 Hz (n=6) or 90 Hz (n=6) for 10 min/day, and compared to OVX age-matched controls (n=6). Five additional rats were used, in vivo, to establish the induced bone surface strain magnitudes (and strain rates). Following a 28 d protocol, bone formation rates in the metaphysis of the proximal tibia were 159% greater in 90 Hz rats when compared to age-matched controls, but 45 Hz rats were not significantly different from controls. Bone morphology of 90 Hz rats indicated significantly greater trabecular bone volume (22% and 25%) and thicker trabeculae (11% and 12%) over either controls or 45 Hz rats in the epiphysis of the distal femur, respectively. Despite the enhanced sensitivity of the skeleton towards the 90 Hz signal, the strain magnitudes and strain rates induced by this frequency were significantly lower than during 45 Hz vibration, suggesting that factors other than matrix strain are driving the anabolic response. Ideally, such mechanical signals represent a non-pharmacologic means of controlling bone mass and morphology in spite of systemic pressures for bone resorption.     

Proliferation of human-derived osteoblast-like cells depends on the cycle number and frequency of uniaxial strain

We tested the hypothesis whether the number of applied load cycles and the frequency of uniaxial strain have an effect on proliferation of human bone derived osteoblast-like cells. A new approach was developed in order to differentiate between the effects of frequency and the effects of cycle number and strain duration. Monolayers of subconfluently grown cells were stretched in rectangular silicone dishes with cyclic predominantly uniaxial movement along there longitudinal axes. Strain was applied over 2 days varying the number of applied load cycles (4-3600) at a constant frequency (1Hz) or varying the frequency (0.1-30Hz) at a constant number of applied cycles (1800) or at a constant strain duration (5min). At a constant frequency, proliferative response increases (103%) with the number of applied cycles until a cycle number maximum (1800 cycles) was reached. 3600 cycles reduced cell number (43%) in contrast to the maximum. The variation of the frequency of applied strain tended to result in slight differences with regard to cell proliferation when cycle number was left constant. However, combined with an appropriate number of cycles there was an optimal frequency (1Hz) as stimulus for bone cell proliferation (84%). A higher frequency (30Hz) in combination with a high cycle number (9000) reduced cell number to control level (4%). This study demonstrates a frequency and cycle number dependent proliferative response of human osteoblast-like cells. It could be shown that effects of the frequency should not be considered separately from the effects of the cycle number.     

The effects of diphosphonates on the growth and glycolysis of connective-tissue cells in culture.

1. The effects of two diphosphonates (compounds containing a P-C-P bond), disodium dichloromethanediphosphonate and disodium 1-hydroxyethane-1,1-diphosphonate, on the metabolism of cultured rat calvaria cells, rabbit ear cartilage cells and rat skin fibroblasts were investigated. 2. The diphosphonates had no effect on the growth of cartilage cells and on the exponential growth of the calvaria cells and the fibroblasts. However, dichloromethanediphosphonate stopped the growth of the calvaria cells and the fibroblasts after the beginning of confluence, whereas the untreated cells were still growing to a certain extent. This inhibition was dose-dependent. After the drug was withdrawn, the cells recovered slowly. 1-Hydroxyethane-1,1-diphosphonate had no detectable effect on the growth of any of the cell types studied. Both diphosphonates decreased the cloning efficiency of calvaria cells and fibroblasts. 3. The K+ content of cartilage, calvaria and skin cells was diminished only by the highest (0.25 mM) concentration of dichloromethanediphosphonate. 4. Radioactive dichloromethanediphosphonate and 1-hydroxyethane-1,1-diphosphonate were taken up linearly with time for at least 48 h by calvaria cells and fibroblasts. The diphosphonate concentration in the cells depended on its concentration in the medium. 5. Both diphosphonates, in a dose-dependent fashion, markedly inhibited glycolysis, dichloromethanediphosphonate being more effective than 1-hydroxyethane-1,1-diphosphonate, at drug doses that had no effect on cell growth or cellular K+ content. Calvaria cells were much more sensitive than cartilage cells. When cartilage cells were cultured in an N2 atmosphere, these effects on glucose and lactate metabolism disappeared. 6. As increased acid production appears to be associated with resorption of bone, this decrease in lactate may explain why diphosphonates are effective inhibitors of bone resorption in vivo.     

Comparisons of antibody reactivity and enzyme sensitivity between small proteoglycans from bovine tendon, bone, and cartilage

Preparations of small proteoglycans from bovine tendon, bone, and cartilage have been compared for sensitivity to various enzymes and reactivity with different polyclonal antibodies. Chondroitinase ABC digestion of all proteoglycans generated a core protein preparation that migrated similarly in sodium dodecyl sulfate-polyacrylamide electrophoresis as a doublet band with Mr approximately equal to 45,000. The small proteoglycans of cartilage were divided into two populations based upon electrophoretic migration of the intact molecules (Rosenberg, L. C., Choi, H. U., Tank, L-H., Johnson, T. L., Pal, S., Webber, C., Reiner, A., and Poole, A. R. (1985) J. Biol. Chem. 260, 6304-6313). The core preparations of tendon, bone, and the faster-migrating (PG II) proteoglycans of cartilage all interacted in Western blot/enzyme-linked immunosorbent assay analysis with polyclonal antibody raised against either the tendon or bone proteoglycans. The slower-migrating (PG I) proteoglycans of cartilage did not react with these antibodies. Digestion of the tendon small proteoglycan with Staphylococcus aureus V8 protease released glycosaminoglycan chains from the molecule and generated a 40-kDa protein fragment that was resistant to further rapid degradation by the enzyme. This large digestion fragment was also prominent following V8 protease digestion of the faster-migrating (PG II) population of small cartilage proteoglycans, but not the small proteoglycan of bone. The N-terminal amino acid sequence of the tendon (PG II) proteoglycan was determined. These observations provide additional evidence for heterogeneity among the chemically similar small proteoglycans from different tissues.     

Suspension osteopenia in mice: Whole body electromagnetic field effects

Whole-body fields were tested for their efficacy in preventing the osteopenia caused by tail suspension in mice. The fields had fundamental frequencies corresponding to the upper range of predicted endogenous impact-generated frequencies (0.25–2.0 kHz) in the long bones. Three distinct whole-body EMFs were applied for 2 weeks on growing mice. Structural, geometric, and material properties of the femora, tibiae, and humeri of suspended mice were altered compared to controls. Comparison of suspended mice and mice subjected to caloric restriction indicates that the changes in caloric intake do not explain either the suspension or the field-induced effects. In agreement with past studies, rather, unloading appears to cause the suspension effects and to be addressed by the EMFs. The EMF effects on bone properties were apparently frequency dependent, with the lower two fundamental frequencies (260 and 910 Hz) altering, albeit slightly, the suspension-induced bone effects. The fields are not apparently optimized for frequency, etc., with respect to therapeutic potential; however, suspension provides a model system for further study of the in vivo effects of EMFs. © 1995 Wiley-Liss, Inc.     

Methylation of chloroplatinate by methylcobalamin.

Incubation of microM levels of K2PtCl6 and methylcobalamin (MeB-12) results in the complete conversion of MeB-12 to aquoB-12. Demethylation is optimal at pH 2.0 and is accelerated by the addition of K2PtCl4. The reaction is stoichiometric between MeB-12 and K2PtCl6 (1:1). Incubation of 40 microM K2PtCl6 with either 40 microM [Me-14C]MeB-12 or [Me-3H]MeB-12 followed by lyophilization converts 70% of the label to a stable form which is associated with Pt upon subsequent paper chromatography and electrophoresis. There is no preferential loss of 3H relative to 14C in the reaction product. When 50 mumoles each of [Me-14C]MeB-12 and K2PtCl6 were reacted and subjected to column chromatography, a labeled UV-absorbing product was separated with a 14C/Pt ratio of 0.9-1.2. The 14C-Pt product has absorption maxima at 260 nm and 208 nm with a minimum at 240 nm (A240 nm/A260 nm = 0.5). Proton NMR spectroscopy confirmed the presence of an H-C-Pt covalent bonding pattern (J for 1H, 195Pt = 78.2 Hz; tau for 194Pt-Me + 196Pt-Me = 6.956).     

The Concentration of Manganese, Iron and Strontium in Bone of Red Fox Vulpes vulpes (L. 1758)

The aims of the study were to determine manganese (Mn), iron (Fe) and strontium (Sr) concentrations in fox bone samples from north-western Poland and to examine the relationships between the bone Mn, Fe and Sr concentrations and the sex and age of the foxes. In the studied samples of fox cartilage, cartilage with adjacent compact bone, compact bone and spongy bone, the concentrations of the analysed metals had the following descending order: Fe > Sr > Mn. The only exception was in compact bone, in which the concentrations were arranged in the order Sr > Fe > Mn. Manganese concentrations were significantly higher in cartilage, compact bone and cartilage with compact bone than in spongy bone. Iron concentrations were higher in cartilage and spongy bone compared with compact bone. Strontium concentrations were greater in compact bone than in cartilage and spongy bone. The manganese, iron and strontium concentrations in the same type of bone material in many cases correlated with each other, with the strongest correlation (r?>?0.70) between Mn and Fe in almost all types of samples. In addition, concentrations of the same metals in different bone materials were closely correlated for Mn and Fe in cartilage and cartilage with adjacent compact bone, and for Sr in compact bone and cartilage with compact bone. In the fox from NW Poland, there were no statistically significant differences in Mn, Fe and Sr in any of the types of bone material between the sexes and immature and adult foxes.     

Collagen synthesis of articular cartilage explants in response to frequency of cyclic mechanical loading

Articular cartilage in vivo experiences the effects of both cell-regulatory proteins and mechanical forces. This study has addressed the hypothesis that the frequency of intermittently or continuously applied mechanical loads is a critical parameter in the regulation of chondrocyte collagen biosynthesis. Cyclic compressive pressure was applied intermittently to bovine articular cartilage explants by using a sinusoidal waveform of 0.1–1.0 Hz frequency with a peak stress of 0.5 MPa for a period of 5–20 s followed by a load-free period of 10–1,000 s. These loading protocols were repeated for a total duration of 6 days. In separate experiments, cyclic loading was continuously applied by using a sinusoidal waveform of 0.001–0.5 Hz frequency and a peak stress of 1.0 MPa for a period of 3 days. Unloaded cartilage discs of the same condyle were cultured in identically constructed loading chambers and served as controls. We report quantitative data showing that (1) no correlation exists between the relative rate of collagen synthesis expressed as the proportion of newly synthesized collagen among newly made proteins and either the frequency of intermittently or continuously applied loads or the overall time cartilage is actively loaded, and (2) individual protocols of intermittently applied loads can reduce the relative rate of collagen synthesis and increase the water content, whereas (3) continuously applied cyclic loads always suppress the relative rate of collagen synthesis compared with that of unloaded control specimens. The results provide further experimental evidence that collagen metabolism is difficult to manipulate by mechanical stimuli. This is physiologically important for the maintainance of the material properties of collagen in view of the heavy mechanical demands made upon it. Moreover, the unaltered or reduced collagen synthesis of cartilage explants might reflect more closely the metabolism of normal or early human osteoarthritic cartilage.This work was supported by the Federal Ministry of Education and Research (BMBF no. 0311058) and by the foundation S.E.T.     

In vivo incidence of apoptosis evaluated with the TdT FragEL DNA fragmentation detection kit in cartilage and bone cells of the rat tibia

Previous studies have shown the occurrence of cell death by apoptosis in cartilage and bone cells, and have suggested a functional relationship between bone growth and remodelling on one hand, and numbers of apoptotic cells on the other. At present, no in vivo studies are available on the frequency of the apoptotic process measured at one time and in one place using the cartilage and bone cells of single specimens. The aim of the present investigation was to measure the in vivo incidence of apoptosis in cartilage and bone cells of the upper epiphysis and secondary ossification metaphyseal bone of the tibia in normal young adult rats. Apoptotic cells were visualized with the terminal deoxynucleotidyl transferase (TdT) FragEL DNA fragmentation detection kit, which is analogous to the TdT-mediated nick end-labelling (TUNEL) method. In the growth cartilage, only a few TUNEL-positive terminal hypertrophic chondrocytes were found; they were 1.32 +/- 0.70% of the total hypertrophic chondrocytes counted along the chondro-osseous junction. There were only a few apoptotic osteoblastic cells and osteocytes (0.22 +/- 0.22% and 0.15 +/- 0.16% of total osteoblasts and osteocytes respectively). TUNEL-positive osteoclasts were 1.03 +/- 0.57% of the total of osteoclastic cells; they usually showed only one or two apoptotic nuclei. The total number of TUNEL-positive bone marrow cells were also counted (56.78 +/- 10.29/mm2 of bone marrow spaces). Our results confirm that apoptosis does occur in hypertrophic chondrocytes and bone cells, and show that its frequency is very low. However, chiefly because of its short lifespan, the frequency of apoptosis in cartilage and bone may be higher than that shown by the TUNEL method. The static estimate that can be obtained with this method might lead to misleading conclusions on the physiological significance of such a dynamic, rapid and asynchronous process, whose precise importance in bone growth and remodelling remains to be determined.     

Plutonium in bone: a high resolution autoradiographic study using plutonium-241.

Plutonium-241 citrate solution at pH 6-5 was injected intravenously into hamsters and an adult rabbit at a dose of 10 kBq g-1 (260 nCi g-1). The hamsters were killed serially at 15 min, 2 hours, 1 day, 10 days, 1 month and 6 months after injection and the rabbit at 1 week. Their knee-joints or femora were examined for plutonium-241 by autoradiography. Few differences were found between the pattern of plutonium distribution in the hamsters and the rabbit. The results showed that although plutonium is initially distributed on bone surfaces, at long periods after injection it becomes deposited throughout the bone matrix. Plutonium uptake by cells in resorbing areas of periosteum, in active osteoblasts, and in chondrocytes in regions of cartilage mineralization was rapid. Plutonium concentrated more slowly on the resting bone surfaces and at sites of low metabolic activity. In addition, some unlabelled sections of skeletal tissues were immersed in a plutonium-241 citrate solution. When autoradiographed, it was found that plutonium was bound by cell nuclei, tooth enamel matrix, dentine, predentine and bone matrix. Plutonium binding to cartilage matrix was weak. The results are discussed with reference to the literature, and a model is proposed to explain the distribution pattern and fate of plutonium deposits in bone.     

Diagnosis of dystrophic lesions of the muscular and ligamentous tendons

The authors presented the results of investigations of the humoral, ulnar, hip, knee joints and calcanei in 4140 patients. Histology has shown that calcifications and ossifications at the sites of tendon fixation are localized in tendons. The substitution of tendinous tissue for a bone results from calcareous dystrophy. Three stages can be singled out: (I) the substitution of a tendon portion for a fibrillar cartilage; (II) the calcification of a cartilage; (III) the substitution of a calcified cartilage for a newly developed spongy bone. According to the preliminary data, similar changes can bep6 observed in ligaments, too. The clinical and x-ray examination have shown the frequency of tendinous and ligamentous lesions (tendinosis and ligamentosis) of different sites. They were shown to be pathology that may not be accompanied by pains. The appearance of the pain syndrome means the development of disease.     

Characteristics of mineral particles in the human bone/cartilage interface

Bone and cartilage consist of different organic matrices, which can both be mineralized by the deposition of nano-sized calcium phosphate particles. We have studied these mineral particles in the mineralized cartilage layer between bone and different types of cartilage (bone/articular cartilage, bone/intervertebral disk, and bone/growth cartilage) of individuals aged 54 years, 12 years, and 6 months. Quantitative backscattered electron imaging and scanning small-angle X-ray scattering at a synchrotron radiation source were combined with light microscopy to determine calcium content, mineral particle size and alignment, and collagen orientation, respectively. Mineralized cartilage revealed a higher calcium content than the adjacent bone (p<0.05 for all samples), whereas the highest values were found in growth cartilage. Surprisingly, we found the mineral platelet width similar for bone and mineralized cartilage, with the exception of the growth cartilage sample. The most striking result, however, was the abrupt change of mineral particle orientation at the interface between the two tissues. While the particles were aligned perpendicular to the interface in cartilage, they were oriented parallel to it in bone, reflecting the morphology of the underlying organic matrices. The tight bonding of mineralized cartilage to bone suggests a mechanical role for the interface of the two elastically different tissues, bone and cartilage.     

New insight into deformation-dependent hydraulic permeability of gels and cartilage,and dynamic behavior of agarose gels in confined compression

Equilibrium, creep, and dynamic behaviors of agarose gels (2.0-14.8%) in confined compression were investigated in this study. The hydraulic permeabilities of gels were determined by curve-fitting creep data to the biphasic model (J. Biomech. Eng. 102 (1980) 73) and found to be similar in value to those published in the literature (AIChE J. 42 (1996) 1220). A new relationship between intrinsic permeability and volume fraction of water was found for agarose gel, capable of predicting deformation-dependent permeabilities of bovine articular cartilage and 2% agarose gel published in literature. This relationship is accurate for gels and cartilage over a wide range of permeabilities (four orders of magnitude variation). The dynamic stiffness of the gels increases with gel concentration and loading frequency (0.01-1.0Hz). The increase in dynamic stiffness with loading frequency is less pronounced for gels with higher concentrations. The results of this study provide a new insight into deformation-dependent permeability behavior of agarose gel and cartilage, and are important for understanding biological responses of cells to interstitial fluid flow in gel or in cartilage under dynamic mechanical loading.