Biomedical subjects

M D Buschmann

Publications and source records attributed to M D Buschmann.

At least 19 recordsLinked to original sources

PubMed2006

Chitosan-glycerol phosphate/blood implants increase cell recruitment, transient vascularization and subchondral bone remodeling in drilled cartilage defects.

OBJECTIVE: Marrow-stimulation techniques are used by surgeons to repair cartilage lesions although consistent regeneration of hyaline cartilage is rare. We have shown previously that autologous blood can be mixed with a polymer solution containing chitosan in a glycerol phosphate (GP) buffer (chitosan-GP), and that implantation of this polymer/blood composite onto marrow-stimulated chondral defects in rabbit and sheep leads to the synthesis of more chondral repair tissue with greater hyaline character compared to marrow-stimulation alone. In the current study, we examined the modulation of cell recruitment and repair tissue characteristics at early post-surgical time points (from day 1 to 56) in a rabbit model to elucidate potential mechanisms behind this improved repair outcome. DESIGN: Thirty-three skeletally mature New Zealand White rabbits underwent bilateral arthrotomies, with each trochlea receiving a cartilage defect (3.5 mm x 4.5mm) bearing four microdrill holes (0.9 mm diameter, approximately 4 mm deep) into the subchondral bone. One defect per rabbit was treated with a chitosan-GP/blood implant, while the other defect was left as a microdrilled control. Repair tissues were stained by histochemistry, for collagen types I, II, and X by immunohistochemistry and analyzed using quantitative stereological tools. RESULTS: Histological analyses demonstrated that control defects followed a typical healing sequence observed previously in marrow-stimulation animal models while chitosan-GP/blood implants led to three significant modifications in the healing sequence at early stages: (1) increased inflammatory and marrow-derived stromal cell recruitment to the microdrill holes, (2) increased vascularization of the provisional repair tissue in the microdrill holes, and (3) increased intramembranous bone formation and subchondral bone remodeling (BR). CONCLUSIONS: These results suggest that the greater levels of provisional tissue vascularization and BR activity are main factors supporting improved cartilage repair when chitosan-GP/blood implants are applied to marrow-stimulated cartilage lesions.

Animals↗

PubMed2006

Chitosan-glycerol phosphate/blood implants elicit hyaline cartilage repair integrated with porous subchondral bone in microdrilled rabbit defects.

OBJECTIVE: We have previously shown that microfractured ovine defects are repaired with more hyaline cartilage when the defect is treated with in situ-solidified implants of chitosan-glycerol phosphate (chitosan-GP) mixed with autologous whole blood. The objectives of this study were (1) to characterize chitosan-GP/blood clots in vitro, and (2) to develop a rabbit marrow stimulation model in order to determine the effects of the chitosan-GP/blood implant and of debridement on the formation of incipient cartilage repair tissue. METHODS: Blood clots were characterized by histology and in vitro clot retraction tests. Bilateral 3.5 x 4 mm trochlear defects debrided into the calcified layer were pierced with four microdrill holes and filled with a chitosan-GP/blood implant or allowed to bleed freely as a control. At 1 day post-surgery, initial defects were characterized by histomorphometry (n=3). After 8 weeks of repair, osteochondral repair tissues between or through the drill holes were evaluated by histology, histomorphometry, collagen type II expression, and stereology (n=16). RESULTS: Chitosan-GP solutions structurally stabilized the blood clots by inhibiting clot retraction. Treatment of drilled defects with chitosan-GP/blood clots led to the formation of a more integrated and hyaline repair tissue above a more porous and vascularized subchondral bone plate compared to drilling alone. Correlation analysis of repair tissue between the drill holes revealed that the absence of calcified cartilage and the presence of a porous subchondral bone plate were predictors of greater repair tissue integration with subchondral bone (P<0.005), and of a higher total O'Driscoll score (P<0.005 and P<0.01, respectively). CONCLUSIONS: Chitosan-GP/blood implants applied in conjunction with drilling, compared to drilling alone, elicited a more hyaline and integrated repair tissue associated with a porous subchondral bone replete with blood vessels. Concomitant regeneration of a vascularized bone plate during cartilage repair could provide progenitors, anabolic factors and nutrients that aid in the formation of hyaline cartilage.

Animals↗

PubMed2006

Defects in articular cartilage metabolism and early arthritis in fibroblast growth factor receptor 3 deficient mice.

Fibroblast growth factor (FGF) receptor 3 has been identified as a key regulator of endochondral bone development and of post-natal bone metabolism through its action on growth plate chondrocytes and osteoblasts, respectively. It has also been shown to promote chondrogenesis and cartilage production by cultured pre-chondrogenic cells in response to FGF18. In the current studies, we show that the absence of signaling through Fgfr3 in the joints of Fgfr3(-/-) mice leads to premature cartilage degeneration and early arthritis. Degenerative changes in cartilage matrix included excessive proteolysis of aggrecan core protein and type II collagen, as measured by neo-epitope immunoreactivity. These changes were accompanied by increased expression of metalloproteinase MMP13, type X collagen, cellular hypertrophy and loss of proteoglycan at the articular surface. Using a novel micro-mechanical indentation protocol, it was shown that articular cartilage in the humeral head of 4-month-old Fgfr3(-/-) mice was less resistant to compressive force and less stiff than that of littermate controls. These results identify Fgfr3 signaling as a potential target for intervention in degenerative disorders of cartilage metabolism.

Aggrecans↗

PubMed2005

Low calcium levels in serum-free media maintain chondrocyte phenotype in monolayer culture and reduce chondrocyte aggregation in suspension culture.

OBJECTIVE: Extracellular calcium influences chondrocyte differentiation and synthesis of extracellular matrix. Previously, calcium concentrations ranging from 0.1 mM to 2 mM have been used in vitro and these studies indicated that low calcium concentrations were generally favorable for chondrocyte culture. Our objective was to extend these findings to yet lower calcium concentrations and to comprehensively examine effects on morphology and phenotype in two culture systems. METHODS: Serum-free media containing 1 mM, 50 microM or 15 microM of calcium and a serum-containing medium were used to culture chondrocytes in suspension and in monolayer, at high and low inoculation density. RESULTS: In monolayer, at low and high density, removing serum and decreasing calcium concentration decreased cell spreading and lowered collagen type I expression whereas collagen type II expression remained stable. In suspension, cells aggregated for all media tested; however, aggregates were smaller and looser in the absence of serum. CONCLUSION: The serum-free 50 microM and 1 mM calcium media provide good alternatives to classical media for monolayer culture since both growth and chondrocyte phenotype were maintained. In suspension culture, the serum-free 1mM calcium medium also possesses the beneficial properties of limiting aggregate size while maintaining growth and phenotype.

Animals↗

PubMed2005

Tissue engineering of cartilage using an injectable and adhesive chitosan-based cell-delivery vehicle.

OBJECTIVE: Adult articular cartilage shows a limited intrinsic repair response to traumatic injury. To regenerate damaged cartilage, cell-assisted repair is thus viewed as a promising therapy, despite being limited by the lack of a suitable technique to deliver and retain chondrogenic cells at the defect site. DESIGN: We have developed a cytocompatible chitosan solution that is space-filling, gels within minutes, and adheres to cartilage and bone in situ. This unique combination of properties suggested significant potential for its use as an arthroscopically injectable vehicle for cell-assisted cartilage repair. The primary goal of this study was to assess the ability of this polymer system, when loaded with primary articular chondrocytes, to support cartilage formation in vitro and in vivo. The chitosan gel was cultured in vitro, with and without chondrocytes, as well as injected subcutaneously in nude mice to form subcutaneous dorsal implants. In vitro and in vivo constructs were collectively analyzed histologically, for chondrocyte mRNA and protein expression, for biochemical levels of glycosaminoglycan, collagen, and DNA, and for mechanical properties. RESULTS: Resulting tissue constructs revealed histochemical, biochemical and mechanical properties comparable to those observed in vitro for primary chondrocytes cultured in 2% agarose. Moreover, the gel was retained after injection into a surgically prepared, rabbit full-thickness chondral defect after 1 day in vivo, and in rabbit osteochondral defects, up to 1 week. CONCLUSIONS: The in situ-gelling chitosan solution described here can support in vitro and in vivo accumulation of cartilage matrix by primary chondrocytes, while persisting in osteochondral defects at least 1 week in vivo.

Animals↗

PubMed2004

Tetrapolar measurement of electrical conductivity and thickness of articular cartilage.

A tetrapolar method to measure electrical conductivity of cartilage and bone, and to estimate the thickness of articular cartilage attached to bone, was developed. We determined the electrical conductivity of humeral head bovine articular cartilage and subchondral bone from a 1- to 2-year-old steer to be 1.14+/-0.11 S/m (mean+/-sd, n =11) and 0.306+/-0.034 S/m, (mean+/-sd, n =3), respectively. For a 4-year-old cow, articular cartilage and subchondral bone electrical conductivity were 0.88+/-0.08 S/m (mean+/-sd, n =9) and 0.179+/-0.046 S/m (mean+/-sd, n =3), respectively. Measurements on slices of cartilage taken from different distances from the articular surface of the steer did not reveal significant depth-dependence of electrical conductivity. We were able to estimate the thickness of articular cartilage with reasonable precision (<20% error) by injecting current from multiple electrode pairs with different inter-electrode distances. Requirements for the precision of this method to measure cartilage thickness include the presence of a distinct layer of calcified cartilage or bone with a much lower electrical conductivity than that of uncalcified articular cartilage, and the use of inter-electrode distances of the current injecting electrodes that are on the order of the cartilage thickness. These or similar methods present an attractive approach to the non-destructive determination of cartilage thickness, a parameter that is required in order to estimate functional properties of cartilage attached to bone, and evaluate the need for therapeutic interventions in arthritis.

Animals↗

PubMed2003

A validated 1H NMR method for the determination of the degree of deacetylation of chitosan.

A method for the determination of the degree of deacetylation (DDA) of chitosan by 1H NMR spectroscopy has been formally validated. Chitosans with DDA ranging from 48 to 100% have been used for the validation. The method is found to be simple, rapid and more precise than other known techniques like IR or titration for %DDA measurements. The precision, ruggedness, robustness, specificity, stability and accuracy of the technique are discussed in this paper.

Acetylation↗

PubMed2003

Strain-rate dependent stiffness of articular cartilage in unconfined compression.

The stiffness of articular cartilage is a nonlinear function of the strain amplitude and strain rate as well as the loading history, as a consequence of the flow of interstitial water and the stiffening of the collagen fibril network. This paper presents a full investigation of the interplay between the fluid kinetics and fibril stiffening of unconfined cartilage disks by analyzing over 200 cases with diverse material properties. The lower and upper elastic limits of the stress (under a given strain) are uniquely established by the instantaneous and equilibrium stiffness (obtained numerically for finite deformations and analytically for small deformations). These limits could be used to determine safe loading protocols in order that the stress in each solid constituent remains within its own elastic limit. For a given compressive strain applied at a low rate, the loading is close to the lower limit and is mostly borne directly by the solid constituents (with little contribution from the fluid). In contrast, however in case of faster compression, the extra loading is predominantly transported to the fibrillar matrix via rising fluid pressure with little increase of stress in the nonfibrillar matrix. The fibrillar matrix absorbs the loading increment by self-stiffening: the quicker the loading the faster the fibril stiffening until the upper elastic loading limit is reached. This self-protective mechanism prevents cartilage from damage since the fibrils are strong in tension. The present work demonstrates the ability of the fibril reinfored poroelastic models to describe the strain rate dependent behavior of articular cartilage in unconfined compression using a mechanism of fibril stiffening mainly induced by the fluid flow.

Animals↗

PubMed2003

Mechanical anisotropy of the human knee articular cartilage in compression.

Articular cartilage exhibits anisotropic mechanical properties when subjected to tension. However, mechanical anisotropy of mature cartilage in compression is poorly known. In this study, both confined and unconfined compression tests of cylindrical cartilage discs, taken from the adult human patello-femoral groove and cut either perpendicular (normal disc) or parallel (tangential disc) to the articular surface, were utilized to determine possible anisotropy in Young's modulus, E, aggregate modulus, Ha, Poisson's ratio, v and hydraulic permeability, k, of articular cartilage. The results indicated that Ha was significantly higher in the direction parallel to the articular surface as compared with the direction perpendicular to the surface (Ha = 1.237 +/- 0.486 MPa versus Ha = 0.845 +/- 0.383 MPa, p = 0.017, n = 10). The values of Poisson's ratio were similar, 0.158 +/- 0.148 for normal discs compared with 0.180 +/- 0.046 for tangential discs. Analysis using the linear biphasic model revealed that the decrease of permeability during the offset compression of 0-20 per cent was higher (p = 0.015, n = 10) in normal (from 25.5 x 10(-15) to 1.8 x 10(-15) m4/N s) than in tangential (from 12.3 x 10(-15) to 1.3 x 10(-15) m4/N s) discs. Based on the results, it is concluded that the mechanical characteristics of adult femoral groove articular cartilage are anisotropic also during compression. Anisotropy during compression may be essential for normal cartilage function. This property has to be considered when developing advanced theoretical models for cartilage biomechanics.

Adult↗

PubMed2003

Investigation of mechanical behavior of articular cartilage by fibril reinforced poroelastic models.

The fibril reinforced poroelastic models have been found successful in describing some mechanical behaviors of articular cartilage in unconfined compression that were not understood previously, including the strong and nonlinear transient response, the strain-magnitude and strain-rate dependent cartilage stiffness and the depth-varying stresses and strains. It has been demonstrated that a better description for the mechanical behavior of cartilage is obtained by introducing a fibrillar matrix into a poroelastic model, in addition to the nonfibrillar matrix and water. This paper reports the development of the nonlinear fibril reinforced homogeneous and nonhomogeneous models and further explores the potentials of the models for investigation of cartilage mechanical response. Some comments are made in regard to further applications of the models and improved accuracy of the material representation.

Biomechanical Phenomena↗

PubMed2002

Streaming potentials maps are spatially resolved indicators of amplitude, frequency and ionic strength dependant responses of articular cartilage to load.

Streaming potential distributions were measured on the surface of articular cartilage in uniaxial unconfined compression using a linear array of microelectrodes. Potential profiles were obtained for sinusoidal and ramp/stress-relaxation displacements and exhibited dependencies on radial position, sinusoidal amplitude and frequency, time during stress relaxation, and on ionic strength. The measurements agreed with trends predicted by biphasic and related models. In particular, the absolute potential amplitude was maximal at the disk center, as was the predicted fluid pressure and the potential gradient (the electric field) was seen to be maximal at the disk periphery, as was the predicted fluid velocity. We also observed a similarity between non-linear behavior of streaming potential amplitude and load amplitude with respect to sinusoidal displacement amplitude. Taken together, these results support many of the phenomena concerning relative fluid-solid movement and fluid pressurization predicted by biphasic and related models, and they indicate the general utility of spatially resolved measurements of streaming potentials for the investigation of electromechanical phenomena in tissues. For example, these streaming potential maps could be used to non-destructively diagnose cartilage extracellular matrix composition and function, as well as to quantify spatially and temporally varying physical signals in cartilage that can induce cellular and extracellular biological responses to load.

Animals↗

PubMed2002

Detection and analysis of cartilage degeneration by spatially resolved streaming potentials.

Cartilage molecular changes in osteoarthritis are most commonly related to the degradation and loss of proteoglycan and collagen fibrils of the extracellular matrix, which directly influence tissue stiffness and compression-generated streaming potentials. In this study, we evaluated the potential of a new technique, spatially resolved mapping of streaming potentials, to non-destructively indicate cartilage health or degeneration. Matched pairs of bovine cartilage/bone explant disks were cultured for 11 days in a serum free medium with and without interleukin-lalpha (IL-1alpha). The electromechanical properties (static stiffness, dynamic stiffness and streaming potentials) of cartilage disks were measured during unconfined compression using a mechanical tester coupled with a linear array of eight 50 microm diameter platinum-iridium microelectrodes. After 11 days of culture, the proteoglycan content of IL-1alpha treated disks was significantly reduced and the denatured and cleaved collagen content was increased compared to control disks. These biochemical alterations were concomitant with the reductions in the amplitudes of the static stiffness, the dynamic stiffness and the streaming potential profile as well as changes in the shape of the streaming potential profile. We found that spatial mapping of streaming potentials presents several advantages for the development of a clinical instrument to evaluate the degeneration of articular cartilage.

Animals↗

PubMed2002

Alterations in mechanical behaviour of articular cartilage due to changes in depth varying material properties--a nonhomogeneous poroelastic model study.

The depth dependence of the material properties is present in normal adult cartilage and is believed to have significant implications in its normal mechanical function. Cartilage pathology may alter the depth dependence, e.g. a reduced depth dependence of the fibril stiffness has been observed in osteoarthritic cartilage. The objective of the present study is to investigate the alterations in the mechanical response of articular cartilage when the depth dependence of the material properties is varied to simulate healthy and pathological situations. This study is made possible by a recently developed nonhomogeneous poroelastic model. Depth variations of the strains and stresses for individual material phases (collagen, proteoglycan and fluid) are obtained for cartilage disks in unconfined compression using the finite element method. The mean nominal axial strain considered is up to 15%, while the axial strain at the articular surface can reach 33%. This paper demonstrates how the mechanical behaviours of cartilage are affected by individual depth dependent cartilage properties, while such observations are not fully available in experimental investigations. This study suggests the possibility of diagnosing cartilage health by analysing its mechanical behaviours.

Animals↗

PubMed2002

The role of fibril reinforcement in the mechanical behavior of cartilage.

Collagen fibril reinforcement was incorporated into a nonlinear poroelastic model for articular cartilage in unconfined compression. It was found that the radial fibrils play a predominant role in the transient mechanical behavior but a less important role in the equilibrium response of cartilage. The radial fibrils are in tension and can be highly stressed during compression, in contrast to low compressive stresses in all directions for the proteoglycan matrix after a small initial compression. The strain dependent fibril stiffening produces strong nonlinear transient response; the fibrils provide extra stiffness to balance a rising fluid pressure and to restrain stress increase in the proteoglycans. The fibril reinforcement, induced by the fluid pressure and flow, also accounts for a complex pattern of strain-magnitude and strain-rate dependence of cartilage stiffness.

Animals↗

PubMed2001

Soluble recombinant neprilysin induces aggrecanase-mediated cleavage of aggrecan in cartilage explant cultures.

Neprilysin (neutral endopeptidase, enkephalinase, CALLA, CD10, NEP) is a regulatory Zn metallopeptidase expressed in the brush border membranes of the kidney and has been found in porcine chondrocytes and rat articular cartilage as well as other cell types and tissues. Although its function in cartilage is not currently known, previous observations of high levels of NEP enzymatic activity in the synovial fluid of arthritic patients and on the chondrocyte membranes of human osteoarthritic cartilage have led to the hypothesis that NEP is involved in the inflammation or degradation pathways in articular cartilage. Our study localized endogenous NEP to the membranes of mature bovine articular chondrocytes in a tissue explant model and demonstrated that the addition of soluble recombinant NEP (sNEP) to the culture medium of bovine cartilage explants leads to the degradation of aggrecan through the action of aggrecanase. A 6-day exposure to sNEP was necessary to initiate the degradation, suggesting that the chondrocytes were responding in a delayed manner to an altered composition of regulatory peptides. This NEP-induced degradation was completely inhibited by the NEP inhibitors thiorphan and phosphoramidon. These results suggest that NEP is present as a transmembrane enzyme on articular chondrocytes where it can cleave regulatory peptides and lead to the induction of aggrecanase.

Aggrecans↗

PubMed2001

The asymmetry of transient response in compression versus release for cartilage in unconfined compression.

Observations in compression tests of articular cartilage have revealed unequal load increments for compression and release of the same amplitude applied to a disk with an identical previously imposed compression (in equilibrium). The mechanism of this asymmetric transient response is investigated here using a nonlinear fibril-reinforced model. It is found that the asymmetry is predominantly produced by the fibril stiffening with its tensile strain. In addition, allowing the hydraulic permeability to decrease significantly with compressive dilatation of cartilage increases the transient fibril strain, resulting in a stronger asymmetry. Large deformation also enhances the asymmetry as a consequence of stronger fibril stiffening.

Biomechanical Phenomena↗

PubMed2001

High frequency acoustic parameters of human and bovine articular cartilage following experimentally-induced matrix degradation.

Matrix degradation and proteoglycan loss in articular cartilag eare features of early osteoarthritis. To determine the effect of matrix degradation and proteoglycan loss on ultrasound propagation in cartilage, we used papain and interleukin-1alpha to degrade the matrix proteoglycans of human and bovine cartilage samples, respectively. There is also minor collagen alteration associated with these chemical degradation methods. We compared the speed of sound and frequency dependent attenuation (20-40 MHz) of control and experimental paired samples. We found that a loss of matrix proteoglycans and collagen disruption resulted in a 20-30% increase in the frequency dependent attenuation and a 2% decrease in the speed of sound in both human and bovine cartilage. We conclude that the frequency dependent attenuation and speed of sound in articular cartilage are sensitive to experimental modification of the matrix proteoglycans and collagen. These findings suggest that ultrasound can potentially be used to detect morphologic changes in articular cartilage associated with the progression of osteoarthritis.

Adult↗

PubMed2000

A fibril reinforced nonhomogeneous poroelastic model for articular cartilage: inhomogeneous response in unconfined compression.

The depth dependence of material properties of articular cartilage, known as the zonal differences, is incorporated into a nonlinear fibril-reinforced poroelastic model developed previously in order to explore the significance of material heterogeneity in the mechanical behavior of cartilage. The material variations proposed are based on extensive observations. The collagen fibrils are modeled as a distinct constituent which reinforces the other two constituents representing proteoglycans and water. The Young's modulus and Poisson's ratio of the drained nonfibrillar matrix are so determined that the aggregate compressive modulus for confined geometry fits the experimental data. Three nonlinear factors are considered, i.e. the effect of finite deformation, the dependence of permeability on dilatation and the fibril stiffening with its tensile strain. Solutions are extracted using a finite element procedure to simulate unconfined compression tests. The features of the model are then demonstrated with an emphasis on the results obtainable only with a nonhomogeneous model, showing reasonable agreement with experiments. The model suggests mechanical behaviors significantly different from those revealed by homogeneous models: not only the depth variations of the strains which are expected by qualitative analyses, but also, for instance, the relaxation-time dependence of the axial strain which is normally not expected in a relaxation test. Therefore, such a nonhomogeneous model is necessary for better understanding of the mechanical behavior of cartilage.

Biomechanical Phenomena↗