Biomedical subjects

L A Setton

Publications and source records attributed to L A Setton.

At least 19 recordsLinked to original sources

PubMed2006

Zonal variations in the three-dimensional morphology of the chondron measured in situ using confocal microscopy.

OBJECTIVE: Chondrocytes in articular cartilage are surrounded by a narrow pericellular matrix (PCM), which together with the enclosed cell(s) are termed the "chondron". Although the precise function of this tissue region is unknown, previous studies provide indirect evidence that the PCM plays an important role in governing the local mechanical environment of chondrocytes. In particular, theoretical models of the chondron under mechanical loading suggest that the shape, size, and biomechanical properties of the PCM significantly influence the stress-strain and fluid flow environment of the cell. The goal of this study was to quantify the three-dimensional morphology of chondron in situ using en bloc immunolabeling of type VI collagen coupled with fluorescence confocal microscopy. METHODS: Three-dimensional reconstructions of intact, fluorescently labeled chondrons were made from stacks of confocal images recorded in situ from the superficial, middle, and deep zones of porcine articular cartilage of the medial femoral condyle. RESULTS: Significant variations in the shape, size, and orientation of chondrocytes and chondrons were observed with depth from the tissue surface, revealing flattened discoidal chondrons in the superficial zone, rounded chondrons in the middle zone, and elongated, multicellular chondrons in the deep zone. CONCLUSIONS: The shape and orientation of the chondron appear to reflect the local collagen architecture of the interterritorial matrix, which varies significantly with depth. Quantitative measurements of morphology of the chondron and its variation with site, disease, or aging may provide new insights into the influence of this structure on physiology and the pathology of articular cartilage.

Animals↗

PubMed2006

Pathogenesis of osteoarthritis-like changes in the joints of mice deficient in type IX collagen.

OBJECTIVE: To examine the pathogenetic mechanisms of osteoarthritis (OA)-like changes in Col9a1-/- mice, which are deficient in type IX collagen. METHODS: Knee joints and temporomandibular joints (TMJs) from Col9a1-/- mice and their wild-type (Col9a1+/+) littermates were examined by light microscopy. Immunohistochemical staining was performed to examine the expression of matrix metalloproteinase 3 (MMP-3) and MMP-13, degraded type II collagen, and the discoidin domain receptor 2 (DDR-2) in knee joints. Cartilage mechanics were also evaluated for compressive properties by microindentation testing of the tibial plateau and for tensile properties by osmotic loading of the femoral condyle. RESULTS: Histologic analysis showed age-dependent OA-like changes in the knee and TMJs of Col9a1-/- mice starting at the age of 3 months. At the age of 6 months, enhanced proteoglycan degradation was observed in the articular cartilage of the knee and TMJs of the mutant mice. The expression of MMP-13 and DDR-2 protein and the amount of degraded type II collagen were higher in the knee joints of Col9a1-/- mice than in their wild-type littermates at the age of 6 months. Changes in cartilage mechanics were observed in the femoral and tibial plateaus of Col9a1-/- mice at 6 months, including a decrease in the compressive modulus and uniaxial modulus. At 3 and 6 months of age, tibial cartilage in Col9a1-/- mice was found to be more permeable to fluid flow, with an associated compromise in the fluid pressurization mechanism of load support. All of these changes occurred only at medial sites. CONCLUSION: Lack of type IX collagen in Col9a1-/- mice results in age-dependent OA-like changes in the knee joints and TMJs.

Animals↗

PubMed2003

Osteoarthritis-like changes and decreased mechanical function of articular cartilage in the joints of mice with the chondrodysplasia gene (cho).

OBJECTIVE: To investigate whether heterozygosity for a loss-of-function mutation in the gene encoding the alpha1 chain of type XI collagen (Col11a1) in mice (chondrodysplasia, cho) causes osteoarthritis (OA), and to understand the biochemical and biomechanical effects of this mutation on articular cartilage in knee and temporomandibular (TM) joints. METHODS: Articular cartilage from the knee and TM joints of mice heterozygous for cho (cho/+) and their wild-type littermates (+/+) was examined. The morphologic properties of cartilage were evaluated, and collagen fibrils were examined by transmission electron microscopy. Immunohistochemical staining was performed to examine the protein expression levels of matrix metalloproteinase 3 (MMP-3) and MMP-13 in knee joints. In 6-month-old animals, fixed-charge density was determined using a semiquantitative histochemical method, and tensile stiffness was determined using an osmotic loading technique. RESULTS: The diameter of collagen fibrils in articular cartilage of knee joints from heterozygous cho/+ mice was increased relative to that in control cartilage, and histologic analysis showed OA-like degenerative changes in knee and TM joints, starting at age 3 months. The changes became more severe with aging. At 3 months, protein expression for MMP-3 was increased in knee joints from cho/+ mice. At 6 months, protein expression for MMP-13 was higher in knee joints from cho/+ mice than in joints from their wild-type littermates, and negative fixed-charge density was significantly decreased. Moreover, tensile stiffness in articular cartilage of knee joints from cho/+ mice was moderately reduced and was inversely correlated with the increase in articular cartilage degeneration. CONCLUSION: Heterozygosity for a loss-of-function mutation in Col11a1 results in the development of OA in the knee and TM joints of cho/+ mice. Morphologic and biochemical evidence of OA appears to precede significant mechanical changes, suggesting that the cho mutation leads to OA through a mechanism that does not initially involve mechanical factors.

Animals↗

PubMed2001

Intervertebral disc cells exhibit differences in gene expression in alginate and monolayer culture.

STUDY DESIGN: The mRNA levels of aggrecan and collagen were quantified in intervertebral disc cells cultured under three conditions: primary alginate culture, monolayer culture, and re-encapsulation in alginate after monolayer culture. OBJECTIVES: To compare the phenotype of intervertebral disc cells under different culture conditions and to investigate the reversibility of cell phenotype after re-encapsulation in alginate after monolayer culture. SUMMARY OF BACKGROUND DATA: The intervertebral disc contains heterogeneous populations of cells that vary with anatomic region. These cells possess significant differences in phenotype that can be preserved in vitro, although the effect of culture conditions on the phenotype of these cells is poorly understood. METHODS: The intervertebral disc cells of 4-5-month-old pigs were isolated enzymatically from three anatomic zones: anulus fibrosus (AF), transition zone (TZ), and nucleus pulposus (NP). Gene expression levels of aggrecan and collagen Types I and II were measured using a quantitative reverse transcriptase--polymerase chain reaction. RESULTS: Gene expression levels of anulus fibrosus and transition zone cells were shifted in monolayer compared with alginate, although the shift was partially reversed when re-encapsulated in alginate. However, NP cells appeared to be insensitive to culture conditions. Furthermore, characteristic patterns of gene expression among AF, TZ, and NP cells in primary alginate culture did not exist in monolayer culture, but they were also observed after re-encapsulation in alginate. CONCLUSION: The findings of this study suggest that anulus fibrosus and transition zone cells undergo a reversible shift in phenotype when cultured in monolayer compared with alginate. These differences suggest that the culture system exerts a strong influence on cell phenotype and may play a role in the response of these cells to biophysical and biochemical stimuli in vitro.

Aggrecans↗

PubMed2001

A noncontacting method for material property determination for articular cartilage from osmotic loading.

Articular cartilage is one of several biological tissues in which swelling effects are important in tissue mechanics and function, and may serve as an indicator of degenerative joint disease. This work presents a new approach to quantify swelling effects in articular cartilage, as well as to determine the material properties of cartilage from a simple free-swelling test. Samples of nondegenerate and degenerate human patellar cartilage were subjected to osmotic loading by equilibrating the tissue in solutions of varying osmolarity. The resulting swelling-induced strains were measured using a noncontacting optical method. A theoretical formulation of articular cartilage in a free-swelling configuration was developed based on an inhomogeneous, triphasic mechano-chemical model. Optimization of the model predictions to the experimental data was performed to determine two parameters descriptive of material stiffness at the surface and deeper cartilage layers, and a third parameter descriptive of thickness of the cartilage surface layer. These parameters were used to determine the thickness-averaged uniaxial modulus of cartilage, H(A). The obtained values for H(A) were similar to those for the tensile modulus of human cartilage reported in the literature. Degeneration resulted in an increase in thickness of the region of "apparent cartilage softening," and a decrease in the value for uniaxial modulus at this layer. These findings provide important evidence that collagen matrix disruption starts at the articular surface and progresses into the deeper layers with continued degeneration. These results suggest that the method provides a means to quantify the severity and depth of degenerative changes in articular cartilage. This method may also be used to determine material properties of cartilage in small joints in which conventional testing methods are difficult to apply.

Adult↗

PubMed2001

Collagen gene expression and mechanical properties of intervertebral disc cell-alginate cultures.

Cells of the intervertebral disc have a limited capacity for matrix repair that may contribute to the onset and progression of degenerative disc changes. In this study, the biosynthetic capacity of cells isolated from specific regions of the porcine intervertebral disc was evaluated in vitro. Using a competitive reverse transcription-polymerase chain reaction technique, gene expression levels for types I and II collagen were quantified in cells cultured for up to 21 d in a three-dimensional alginate culture system and compared to levels obtained for cells in vivo. The mechanical properties of cell-alginate constructs were measured in compression and shear after periods of culture up to 16 weeks. Cells from the anulus fibrosus expressed the most type I collagen mRNA in vivo and in vitro, while cells from the transition zone expressed the most type II collagen mRNA in vivo and in vitro. Mechanical testing results indicate that a mechanically functional matrix did not form at any time during the culture period; rather, decreases of up to 50% were observed in the compressive and shear moduli of the cell-alginate constructs compared to alginate with no cells. Together with results of prior studies, these results suggest that intervertebral disc cells maintain characteristics of their phenotype when cultured in alginate, but the molecules they synthesize are not able to form a mechanically functional matrix in vitro.

Alginates↗

PubMed2001

Altered mechanics and histomorphometry of canine tibial cartilage following joint immobilization.

Joint immobilization is associated with altered cartilage biosynthesis and catabolism that may affect cartilage mechanics and joint function. In this study, the mechanical behavior of articular cartilage was studied in an experimental model of joint immobilization, in which the canine knee was cast-immobilized at 90 degrees of flexion for 4 weeks. Articular cartilage from the medial tibial plateau was tested in compression and in shear. Biochemical assays for water and glycosaminoglycan content and histomorphometric grading were performed on site-matched samples. Significant decreases in the equilibrium and dynamic shear moduli, but not compressive moduli, were observed in cartilage after 4 weeks of joint immobilization as compared to cartilage from a separate control population. Importantly, there was also evidence of a decrease in the compressive and shear moduli of tibial cartilage from the contralateral knee joints compared to control joints that were not immobilized. No significant effect of immobilization on the biochemical parameters or histomorphometric scores was detected, expect for a significant loss of proteoglycan staining following immobilization. These findings for changes in the tibial cartilage following cast immobilization are consistent with a mild form of cartilage degeneration.

Analysis of Variance↗

PubMed2001

Anisotropic and inhomogeneous tensile behavior of the human anulus fibrosus: experimental measurement and material model predictions.

The anulus fibrosus (AF) of the intervertebral disc exhibits spatial variations in structure and composition that give rise to both anisotropy and inhomogeneity in its material behaviors in tension. In this study, the tensile moduli and Poisson's ratios were measured in samples of human AF along circumferential, axial, and radial directions at inner and outer sites. There was evidence of significant inhomogeneity in the linear-region circumferential tensile modulus (17.4+/-14.3 MPa versus 5.6+/-4.7 MPa, outer versus inner sites) and the Poisson's ratio v21 (0.67+/-0.22 versus 1.6+/-0.7, outer versus inner), but not in the axial modulus (0.8+/-0.9 MPa) or the Poisson's ratios V12 (1.8+/-1.4) or v13 (0.6+/-0.7). These properties were implemented in a linear an isotropic material model of the AF to determine a complete set of model properties and to predict material behaviors for the AF under idealized kinematic states. These predictions demonstrate that interactions between fiber populations in the multilamellae AF significantly contribute to the material behavior, suggesting that a model for th

Adult↗

PubMed2000

Longitudinal characterization of synovial fluid biomarkers in the canine meniscectomy model of osteoarthritis.

Damage to the meniscus can lead to posttraumatic osteoarthritis. Early markers of joint injury and tissue disease may be useful in developing and administering clinical treatment. We investigated the effects of total medial meniscectomy on biomarkers measured serially in synovial lavage fluid each month for 3 months. Following meniscectomy in dogs, four biomarkers were evaluated: cartilage oligomeric matrix protein, keratan sulfate epitope (5D4), the 3B3(-) neoepitope of chondroitin-6-sulfate, and the 3B3(+) chondroitinase-generated epitope of chondroitin-6-sulfate. Meniscectomy led to statistically significant elevations of all four biomarkers, with levels peaking at 4 weeks. By 12 weeks, the level of the 5D4 epitope returned to the preoperative baseline level whereas that of cartilage oligomeric matrix protein, 3B3(-), and 3B3(+) remained above the baseline. Concentrations of these biomarkers in the knees not operated on did not change significantly from the baseline. The levels of cartilage oligomeric matrix protein and 3B3(-) relative to 3B3(+) remained constant in all knees. In contrast, the level of 5D4 relative to 3B3(+) declined over time in the knee operated on but remained constant in the knee not operated on. These results demonstrate a quantitative change in the molecular components of synovial fluid after meniscectomy, as well as a qualitative change evinced by an alteration in the relative proportions of these epitopes. Extensive analyses showed a strong correlation between serum levels of 3B3(-) from the femoral and cephalic veins; however, serum 3B3(-) was not correlated with synovial fluid 3B3(-). These findings support the hypothesis that the concentrations of select cartilage biomarkers in synovial fluid are altered following meniscectomy and are promising tools for objectively monitoring the induction of osteoarthritis in this model system.

Animals↗

PubMed2000

Simultaneous changes in the mechanical properties, quantitative collagen organization, and proteoglycan concentration of articular cartilage following canine meniscectomy.

The mechanical properties and microstructure of articular cartilage from the canine tibial plateau were studied 12 weeks after total medial meniscectomy. The organization of the birefringent collagen network was measured with quantitative polarized light microscopy to determine the thickness and the degree of organization of the superficial and deep zones. The zonal concentration of sulfated glycosaminoglycan was quantified with digital densitometry of safranin-O staining. Equilibrium compressive and shear properties, as well as dynamic shear properties, were measured at sites adjacent to those of microstructural analysis. The results evinced significant loss of cartilage function following meniscectomy, with decreases of 20-50% in the compressive and shear moduli. There was no evidence of alterations in the degree of collagen fibrillar organization, although a complete loss of the surface zone was seen in 60% of the samples that underwent meniscectomy. Meniscectomy resulted in a decreased concentration of sulfated glycosaminoglycan, and significant positive correlations were found between the equilibrium compressive modulus and the glycosaminoglycan content. Furthermore, the shear properties of cartilage correlated directly with collagen fibrillar organization measured at the superficial zone of corresponding sites. These findings demonstrate that meniscectomy leads to impaired mechanical function of articular cartilage, with significant evidence of quantitative correlations between cartilage microstructure and mechanics.

Animals↗

PubMed2000

A linear material model for fiber-induced anisotropy of the anulus fibrosus.

The anulus fibrosus (AF) is a lamellar, fibrocartilaginous component of the intervertebral disc, which exhibits highly anisotropic behaviors in tension. These behaviors arise from the material's unique collagen structure. We have investigated the use of a linear, fiber-induced anisotropic model for the AF using a quadratic strain energy density formulation with an explicit representation of the collagen fiber populations. We have proposed a representative set of intrinsic material properties using independent datasets of the AF from the literature and appropriate thermodynamic constraints. The model was validated by comparing predictions with previous experimental data for AF behavior and its dependence on fiber angle. The model predicts that compressible effects may exist for the AF, and suggests that physical effects of the equivalent "matrix," "fiber," "fiber-matrix," and "fiber-fiber," interactions may be important contributors to the mechanical behavior of the AF.

Anisotropy↗

PubMed2000

The micromechanical environment of intervertebral disc cells: effect of matrix anisotropy and cell geometry predicted by a linear model.

Cells of the intervertebral disc exhibit spatial variations in phenotype and morphology that may be related to differences in their local mechanical environments. In this study, the stresses, strains, and dilatations in and around cells of the intervertebral disc were studied with an analytical model of the cell as a mechanical inclusion embedded in a transversely isotropic matrix. In response to tensile loading of the matrix, the local mechanical environment of the cell differed among the anatomic regions of the disc and was strongly influenced by changes in both matrix anisotropy and parameters of cell geometry. The results of this study suggest that the local cellular mechanical environment may play a role in determining both cell morphology in situ and the inhomogeneous response to mechanical loading observed in cells of the disc.

Animals↗

PubMed1999

Viscoelastic properties of intervertebral disc cells. Identification of two biomechanically distinct cell populations.

STUDY DESIGN: A combined experimental and theoretical biomechanical study to quantify the mechanical properties of living cells of the porcine intervertebral disc. OBJECTIVES: To quantify zonal variations in the mechanical properties and morphology of cells isolated from the intervertebral disc. SUMMARY OF BACKGROUND DATA: Cellular response to mechanical stimuli is influenced by the mechanical properties of cells and of the extracellular matrix. Significant zonal variations in intervertebral disc matrix properties have been reported. No information is currently available on the corresponding regional variations in the mechanical properties of intervertebral disc cells, despite evidence of significant differences in cellular phenotype and biologic response to loading. METHODS: The micropipette aspiration test was used in combination with a three-parameter viscoelastic solid model to measure the mechanical properties of cells isolated from the anulus fibrosus, transition zone, and nucleus pulposus. RESULTS: Intervertebral disc cells exhibited viscoelastic solid behaviors. Highly significant differences were observed in the morphology, cytoskeletal arrangement, and biomechanical properties of the nucleus pulposus cells as compared with anulus fibrosus or transition zone cells. Cells of the nucleus pulposus were approximately three times stiffer and significantly more viscous than cells of the anulus fibrosus or transition zone. CONCLUSIONS: The findings of this study provide new evidence for the existence of two biomechanically distinct cell populations in the intervertebral disc. These differences in mechanical behavior may be related to observed differences in the cytoskeletal architecture between these cells, and may further play an important role in the development, maintenance, and degeneration of the intervertebral disc.

Animals↗

PubMed1999

Compressive and shear properties of alginate gel: effects of sodium ions and alginate concentration.

The equilibrium and viscoelastic properties of alginate gel crosslinked with Ca2+ were determined as a function of alginate concentration and duration of exposure to physiologic concentrations of NaCl. Compressive and shear stress relaxation tests and oscillatory shear tests were performed to measure the material properties at two time periods after storage in NaCl compared to no NaCl exposure. The effect of concentration was determined by testing 1-3% alginate gel in a bath of physiological NaCl and CaCl2. After 15 h of exposure to NaCl, the compressive, equilibrium shear, and dynamic shear moduli decreased by 63, 84, and 90% of control values, respectively. The material properties exhibited no further changes after 7 days of exposure to NaCl. The loss angle and amplitude of the relaxation function in the shear also decreased, indicating less viscous behaviors in both dynamic and transient configurations. All moduli, but not the loss angle, significantly increased with increasing alginate concentration. The observed decrease in compressive and shear stiffness for alginate gel after exposure to Na+ was significant and indicated that physiological conditions will soften the gel over a time period of up to 7 days after gelation. The alginate gel retains significant solid-like behaviors, however, as measured by a loss angle of approximately 3 degrees. This study provides the first available data for material properties of alginate gel tested in physiological saline.

Alginates↗

PubMed1999

Diffusion tensor microscopy of the intervertebral disc anulus fibrosus.

Morphologically accurate biomechanical models of the intervertebral disc anulus fibrosus (AF) require precise knowledge of its lamellar architecture; however, available methods of assessment are limited by poor spatial resolution or the destructive nature of the technique. In a novel approach, diffusion tensor microscopy was used in this study to characterize the microstructure of excised porcine AF samples. Results show diffusion in the AF to be anisotropic. The orientations of anisotropy exhibit a layered morphology that agrees with light micrographs of the corresponding samples, and the behavior of the orientation angles is consistent with the known AF collagen fiber architecture. A static magnetic field-dependent relaxation anisotropy was observed in the AF, which has methodological implications for magnetic resonance (MR) imaging of ordered collagenous tissues. These findings present MR diffusion tensor microscopy as a potentially valuable tool to assess quantitatively and nondestructively water diffusion anisotropy and lamellar structure of the intervertebral disc AF.

Algorithms↗

PubMed1999

Tensile properties of articular cartilage are altered by meniscectomy in a canine model of osteoarthritis.

Loss of or damage to the meniscus alters the pattern of loading in the knee joint and frequently leads to cartilage degeneration and osteoarthritis. The mechanical properties of articular cartilage have been shown to reflect the extent of cartilage degeneration in human osteoarthritis and in experimental models of joint disease, but there is little experimental data documenting changes in cartilage mechanics following meniscectomy. We hypothesized that the tensile properties of the surface zone of articular cartilage are altered following total medial meniscectomy. Twelve mongrel dogs underwent complete resection of the medial meniscus in the right knee, and the femoral cartilage was studied 12 weeks after the operation. We performed uniaxial, tensile stress-relaxation tests to determine the equilibrium tensile modulus of surface-zone cartilage. Water and glycosaminoglycan content were also measured at site-matched locations. The tensile moduli of the cartilage decreased significantly following meniscectomy. The linear region modulus decreased by 40%, from 25.5 +/- 7.7 to 15.3 +/- 7.2 MPa. There was a weak (r = -0.45), but significant, correlation between the linear region modulus and the gross morphological grade for cartilage damage. Water and glycosaminoglycan content did not change following meniscectomy. Composition was not correlated with mechanical properties or morphological grade, suggesting that cartilage structure may play a more important role than composition in determining the mechanical properties. The observed decrease in cartilage material properties provides a quantitative measure of the loss of cartilage function following meniscectomy and reflects a pattern of change that is consistent with damage to the collagen-proteoglycan solid network.

Animals↗

PubMed1999

Nonuniform swelling-induced residual strains in articular cartilage.

Swelling effects in cartilage originate from an interstitial osmotic pressure generated by the presence of negatively charged proteoglycans in the tissue. This swelling pressure gives rise to a non-zero residual strain in the cartilage solid matrix in the absence of externally applied loads. Previous studies have quantified swelling effects in cartilage as volumetric or dimensional change of excised samples in varying osmotically active solutions. This study presents a new optical technique for measuring two-dimensional swelling-induced residual strain fields in planar samples of articular cartilage attached to the bone (i.e., in situ). Osmotic loading was applied to canine cartilage bone samples by equilibration in external baths of varying NaCl concentration. Non-zero swelling-induced strains were measured in physiological saline, giving evidence of the existence of residual strains in articular cartilage. Only one component of planar strain (i.e., in thickness direction) was found to be non-zero. This strain was found to be highly non-uniform in the thickness direction, with evidence of compressive strain in the deep zone of cartilage and tensile strain in the middle and surface zones. The obtained results can be used to characterize the material properties of the articular cartilage solid matrix, with estimated values of 26 M Pa for the tensile modulus for middle zone cartilage. The method provides the basis to obtain material properties of the cartilage solid matrix from a simple, free-swelling test and may be useful for quantifying changes in cartilage properties with injury, degeneration and repair.

Analysis of Variance↗

PubMed1999

Altered mechanics of cartilage with osteoarthritis: human osteoarthritis and an experimental model of joint degeneration.

OBJECTIVE: Studies of cartilage mechanics seek to determine the fundamental relationships between mechanical behavior and the composition and structure of healthy cartilage and to determine mechanisms for changes associated with degeneration. METHOD: The mechanics of normal and osteoarthritic (OA) human articular cartilage are reviewed. Studies of the initiation and pathogenesis of cartilage degeneration in the anterior cruciate ligament transection (ACLT) model of joint instability are also presented. RESULTS: In human cartilage with OA, tensile, compressive and shear behaviors are dramatically altered. These changes present as decreases in the modulus or stiffness of OA cartilage in tension, compression and shear loading, and increases in the propensity to swell as compared to healthy cartilage. In the ACL transection model of OA, similar changes in the mechanics of cartilage have been observed. In addition, changes in structure, composition, and as metabolism consistent with human OA have been found. Deterioration of the collagen-proteoglycan solid network, which appears to be focused at the articular surface, has been the earliest cartilage changes in the model. It remains to be determined if the initial disruption of the cartilage surface is a direct result of mechanical forces or a product of altered chondrocyte activity. CONCLUSIONS: These data and continued research using experimental models of OA provide a basis for our understanding of the pathogenesis and the time course of events in OA and will lead to the development of better procedures for disease intervention and treatment.

Animals↗