Communication isn't just talking. What's new in AAC.
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Cultural Diversity↗
Biomedical subjects
A Ratcliffe
Publications and source records attributed to A Ratcliffe.
Determination of collagen-proteoglycan interactions in vitro.
The objective of this study was to characterize the physical interactions of the molecular networks formed by mixtures of collagen and proteoglycan in vitro. Pure proteoglycan aggrecan solutions, collagen (type II) suspensions and mixtures of these molecules in varying proportions and concentrations were subjected to viscometric flow measurements using a cone-on-plate viscometer. Linear viscoelastic and non-Newtonian flow properties of these solutions and suspensions were described using a second-order statistical network theory for polymeric fluids (Zhu et al., 1991, J. Biomechanics 24, 1007-1018). This theory provides a set of material coefficients which relate the macroscopic flow behavior of the fluid to an idealized molecular network structure. The results indicated distinct differences between the flow properties of pure collagen suspensions and those of pure proteoglycan solutions. The collagen network showed much greater shear stiffness and more effective energy storage capability than the proteoglycan network. The relative proportion of collagen to proteoglycan is the dominant factor in determining the flow behavior of the mixtures. Analysis of the statistical network theory indicated that the collagen in a collagen-proteoglycan mixture enhances molecular interactions by increasing the amount of entanglement interactions and/or the strength of interaction, while aggrecan acts to reduce the number and/or strength of molecular interactions. These results characterize the physical interactions between type II collagen and aggrecan and provide some insight into their potential roles in giving articular cartilage its mechanical behavior.
Aggrecans↗
Biochemical markers in synovial fluid identify early osteoarthritis of the glenohumeral joint.
The objective of this study on the glenohumeral joint was to assess the (1) accuracy of clinical diagnosis of osteoarthritis compared with arthroscopic diagnosis, and (2) the ability of biochemical markers in synovial fluid to detect osteoarthritis. Patients (96) were examined clinically and the preoperative diagnosis of osteoarthritis was recorded. At surgery (arthroscopy or arthroplasty), the glenohumeral joint was inspected for signs of osteoarthritis, and the joint osteoarthritis grade (I-IV) was recorded. At surgery, synovial fluid lavage was obtained from the joint, and later analyzed to determine levels of aggrecan components: total sulfated glycosaminoglycan and keratan sulfate epitope, link protein and the chondroitin sulfate epitope recognized by antibody 3B3 (3B3(-)). Compared with arthroscopic diagnosis of osteoarthritis, the results showed that the clinical diagnosis did not wrongly identify joints without osteoarthritis, and was always able to identify joints with advanced (Grade IV) osteoarthritis. Grade II osteoarthritis was rarely identified (10% of the time), and Grade III osteoarthritis was identified 50% of the time. Biochemical assessment of the synovial fluid showed that the catabolic markers (sulfated glycosaminoglycan, keratan sulfate and link protein) were elevated in fluids from joints with moderate (Grade III) and advanced osteoarthritis (Grade IV), and the 3B3(-) epitope was elevated in Grades II, III, and IV. These results show that arthroscopic diagnosis for osteoarthritis, of the glenohumeral joint is particularly useful for early and moderate osteoarthritis, where clinical (nonarthroscopic) diagnosis is poor, and that biochemical analysis of the synovial fluids corresponds well to arthroscopic diagnosis of shoulder osteoarthritis.
Arthroscopy↗
Structure of the human aggrecan gene: exon-intron organization and association with the protein domains.
The complete exon-intron organization of the human aggrecan gene has been defined, and the exon organization has been compared with the individual domains of the protein core. A yeast artificial chromosome containing the aggrecan gene was selected from the Centre d'Etude du Polymorphisme Humaine yeast artificial chromosome library. A cosmid sulibrary was created from this, and direct sequencing of individual cosmids was used to provide the exon-intron organization. The human aggrecan gene was found to be composed of 19 exons ranging in size from 77 to 4224 bp. Exon 1 is non-coding, whereas exons 2-19 code for a protein core of 2454 amino acids with a calculated mass of 254379 Da. Intron 1 of the gene is at least 13 kb. Overall, the sizes of the 18 introns range from 0.5 to greater than 13 kb. Each intron begins with a GT and ends with an AG, thus obeying the GT/AG rule of splice-junction sequences. The entire coding region is contained in 39.4 kb of the gene. The organization of exons is strongly related to the specific domains of the protein core. The A loop of G1 and the interglobular domain are encoded by exons 3 and 7 respectively. The B and B' loops of G1 are encoded by exons 4-6, and those of G2 are encoded by exons 8-10. These sets of exons, coding for the B and B' loops, are identical in size and organization. This is supported by the intron classes associated with these exons. Exon 11 codes for the 5' half of the keratan sulphate-rich region, and exon 12 codes for the 3' half of the keratan sulphate-rich region as well as the entire chondroitin sulphate-rich region. G3 is encoded by exons 13-18, including the alternatively spliced epidermal growth factor-like and complement regulatory protein-like domains. The correspondence between the exon organization and the protein domains argues strongly for modular assembly of the aggrecan gene.
Aggrecans↗
Quantitative polymerase chain reaction assay for aggrecan and link protein gene expression in cartilage.
A procedure has been developed to quantify the levels of aggrecan and link protein mRNAs in small amounts of various tissues, including cartilage, using the power of PCR to amplify extremely low levels of specific templates. The PCR protocol which was selected allows for a simple assay procedure, with standards in different tubes from the samples. This straightforward procedure is quantitative, inexpensive, and allows for many samples to be analyzed at one time.
Aggrecans↗
Chondrocyte deformation and local tissue strain in articular cartilage: a confocal microscopy study.
It is well accepted that mechanical forces can modulate the metabolic activity of chondrocytes, although the specific mechanisms of mechanical signal transduction in articular cartilage are still unknown. One proposed pathway through which chondrocytes may perceive changes in their mechanical environment is directly through cellular deformation. An important step toward understanding the role of chondrocyte deformation in signal transduction is to determine the changes in the shape and volume of chondrocytes during applied compression of the tissue. Recently, a technique was developed for quantitative morphometry of viable chondrocytes within the extracellular matrix using three-dimensional confocal scanning laser microscopy. In the present study, this method was used to quantify changes in chondrocyte morphology and local tissue deformation in the surface, middle, and deep zones in explants of canine articular cartilage subjected to physiological levels of matrix deformation. The results indicated that at 15% surface-to-surface equilibrium strain in the tissue, a similar magnitude of local tissue strain occurs in the middle and deep zones. In the surface zone, local strains of 19% were observed, indicating that the compressive stiffness of the surface zone is significantly less than that of the middle and deep zones. With this degree of tissue deformation, significant decreases in cellular height of 26, 19, and 20% and in cell volume of 22, 16, and 17% were observed in the surface, middle, and deep zones, respectively. The deformation of chondrocytes in the surface zone was anisotropic, with significant lateral expansion occurring in the direction perpendicular to the local split-line pattern. When compression was removed, there was complete recovery of cellular morphology in all cases. These observations support the hypothesis that deformation of chondrocytes or a change in their volume may occur during in vivo joint loading and may have a role in the mechanical signal transduction pathway of articular cartilage.
Animals↗
Dose-dependent response of gamma irradiation on mechanical properties and related biochemical composition of goat bone-patellar tendon-bone allografts.
We studied the effects of gamma irradiation on the dimensions, mechanical and material properties, and mature hydroxypyridinium crosslink density of collagen in goat patellar tendon-bone specimens. Left and right patellar tendon-bone units were removed from 10 adult female goats and were bisected longitudinally. Each tendon half was frozen, and then the left halves were exposed to 4, 6, or 8 Mrad (40,000, 60,000, or 80,000 Gy) of gamma irradiation. The contralateral tendon halves served as controls (no irradiation). Each specimen then was loaded to failure in tension, and its soft-tissue midsubstance was processed to measure collagen content and hydroxypyridinium crosslink density. Dose-dependent reductions in the mechanical properties were found, including 46% (p < 0.01) and 18% (p < 0.05) reductions in maximum force and stiffness, respectively, at 4 Mrad. Similar reductions were noted in material properties, including 37% (p < 0.005) and 8% (p > 0.05) reductions in maximum stress and modulus, respectively, at 4 Mrad. These results are consistent with our previous report involving 2 and 3 Mrad (20,000 and 30,000 Gy) of exposure. We also found significant decreases in hydroxypyridinium crosslink density with 6 Mrad of irradiation (p < 0.05). However, since only one biomechanical parameter (modulus) correlated significantly with only one biochemical measure (hydroxypyridinium crosslink density) (p < 0.05), other possible mechanisms also are being explored to more fully explain these dose-dependent changes.
Analysis of Variance↗
Changes in proteoglycan synthesis of chondrocytes in articular cartilage are associated with the time-dependent changes in their mechanical environment.
Explant loading experiments were conducted to investigate the effect of load duration on proteoglycan synthesis. A compressive load of 0.1 MPa applied for 10 min was found to stimulate proteoglycan synthesis, while the same load applied for 20 h suppressed synthesis. This bimodal response suggests that the cells are responding to different mechanical stimuli as time progresses. A theoretical model has therefore been developed to describe the mechanical environment perceived by cells within soft hydrated tissues (e.g. articular cartilage) while the tissue is being loaded. The cells are modeled, using the biphasic theory, as fluid-solid inclusions embedded in and attached to a biphasic extracellular matrix of distinct material properties. A method of solution is developed which is valid for any axisymmetric loading configuration, provided that the cell radius, a, is small relative to the tissue height, h (i.e. h/a >> 1). A closed-form analytical solution for this inclusion problem is then presented for the confined compression configuration. Results from this model show that the mechanical environment in and around the cells is time dependent and inhomogeneous, and can be significantly influenced by differences in properties between the cell and the extracellular matrix.
Algorithms↗
Cartilage metalloproteases in disuse atrophy.
A canine knee model of disuse atrophy produced by nonrigid fixation (sling) was characterized in respect to variables of proteoglycan size distribution, as well as biomechanical properties versus controls. Using this model, we found, in addition to the accepted dogma attributing changes to reduced protein synthesis by chondrocytes, that there is elevation of proteases and depression of tissue inhibitor of metalloproteases (TIMP) in atrophic knee cartilage. The findings are suggestive of cartilage remodelling reminiscent of bone remodelling in disuse atrophy reported by others. Whether the abnormal changes of protease-TIMP balance in knee cartilage can be retarded prophylactically by concurrent treatment with pentosan polysulfate and insulin like growth factor 1 remains uncertain.
Animals↗
High levels of aggrecan aggregate components are present in synovial fluids from human knee joints with chronic injury or osteoarthrosis.
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Adult↗
Regional variation in tensile properties and biochemical composition of the human lumbar anulus fibrosus.
STUDY DESIGN: The structure-function relationship of anulus fibrosus of nondegenerate lumbar intervertebral discs was investigated. OBJECTIVES: The tensile properties and biochemical composition of single lamella specimens from human anulus fibrosus and their variations with anatomic region were determined. SUMMARY OF BACKGROUND DATA: Regional differences in composition and ultrastructure suggest differences in tensile properties. METHODS: Single lamella specimens were isolated from the anulus, equilibrated in 0.15 mol/L NaCl and tested in uniaxial tension using a slow strain-rate protocol. Adjacent specimens were used to determine biochemical composition (including hydration, collagen, proteoglycan, and hydroxypyridinium crosslink density). Tensile properties, biochemical composition, and anatomic location were compared. RESULTS: Significant radial and circumferential variations in tensile properties of anulus were detected, with the anterior being stiffer than the posterolateral regions, and the outer being stiffer than the inner regions. CONCLUSIONS: The regional differences in tensile properties may result predominantly from structural rather than compositional variations and may contribute to the clinical frequency of anulus failure in the posterolateral region.
Collagen↗
Compressive mechanical properties of the human anulus fibrosus and their relationship to biochemical composition.
To enhance understanding of the biomechanical role of the intervertebral disc, the compressive properties and biochemical composition of nondegenerate samples of anulus fibrosus were determined as a function of radial position, region, and level. Because of the large swelling propensity of this tissue, a method was developed to test excised specimens while maintaining their in situ geometry and hydration. Using an analysis based on linear biphasic theory, the compressive modulus, hydraulic permeability, and isometric swelling pressure of the anulus fibrosus were determined and correlated with the tissue composition. The findings indicate that the anulus fibrosus is inhomogeneous, with regional and radial variations in both material properties and biochemical composition. The results of this study suggest that both structural and compositional factors may determine the mechanical behavior.
Adult↗
Differential levels of synovial fluid aggrecan aggregate components in experimental osteoarthritis and joint disuse.
The levels of proteoglycan aggregate components (link protein, keratan sulfate epitope, and total sulfated glycosaminoglycan) were determined in the synovial fluid lavages of dogs with experimental osteoarthritis or disuse atrophy. A model of experimental osteoarthritis was created by transection of the anterior cruciate ligament of the right knee; studies were carried out 6 and 12 weeks after surgery. Joint disuse was studied at 4 and 8 weeks after initiation of the disuse. Recovery after disuse also was studied in joints that had 3 weeks of remobilization after 4 or 8 weeks of disuse. Synovial fluid lavages from the right knee joints of untreated animals were used as controls. The concentrations of keratan sulfate epitope, sulfated glycosaminoglycan, and link protein in the synovial fluid lavages at 6 and 12 weeks after transection of the anterior cruciate were elevated compared with the control values. Similar analysis of the fluid after disuse showed that the levels of keratan sulfate epitope and sulfated glycosaminoglycan were increased compared with the control levels and the levels after transection. However, the concentration of link protein in the fluid after disuse was not significantly different from the control level. The levels of keratan sulfate epitope and sulfated glycosaminoglycan in the synovial fluid lavages after disuse with recovery were high, but the levels of link protein remained low. The results indicate that the catabolism of proteoglycan aggregates in articular cartilage during early osteoarthritis and disuse is different. The determination of keratan sulfate epitope in synovial fluid lavages appears to provide a relatively general indication of proteoglycan catabolism, whereas increased levels of link protein may be more indicative of cartilage degeneration.
Aggrecans↗
Mechanical and biochemical changes in the superficial zone of articular cartilage in canine experimental osteoarthritis.
The changes in the tensile mechanical properties and biochemical composition of the superficial zone of articular cartilage were examined in a canine model of early osteoarthritis generated by transection of the anterior cruciate ligament. Sixteen weeks following ligament transection, the tensile stiffness of the articular cartilage was decreased by 44% and the ion-induced stress relaxation of the tissue was increased by 57% compared with the contralateral control. Biochemical analyses indicated that the water content of the experimental tissue was increased by 13%, which was reflected as an apparent 37% decrease in the proteoglycan content and a 36% decrease in the collagen content (expressed per wet weight). The hydroxypyridinium crosslink density was decreased in the experimental tissue by 11%. A significant negative correlation was found between the ion-induced stress relaxation and the hydroxypyridinium crosslink density in both control tissue (R = -0.56) and experimental tissue (R = -0.70). No correlation was noted between the tensile stiffness and the biochemical composition of the tissue. These results suggest that, in the superficial zone of articular cartilage, the structure of the tissue may play a more important role than the composition in the determination of its mechanical properties. A major event observed in the model of early osteoarthritis appears to be the disruption and remodeling of the collagen network in the superficial zone of the articular cartilage.
Animals↗
Differential rates of aggrecan synthesis and breakdown in different zones of the bovine growth plate.
This study examines the basic metabolic events of aggrecan synthesis and breakdown in the growth plate at different depths and at different stages of development. Growth plate was harvested from the distal tibia of fetal and calf tissue and maintained as explants in serum-free-conditions. The tissue was sectioned into three equal depths (resting/proliferative zone, upper hypertrophic zone, and lower hypertrophic zone) and (a) cultured for three days with daily media change for studies of proteoglycan breakdown rates, or (b) incubated with [35S]-sulfate to determine relative rates of proteoglycan synthesis. Rates of both aggrecan synthesis and turnover were highest in the resting/proliferative zone compared to the upper or lower hypertrophic zones, and was greater in the calf compared to the fetal tissue. In situ hybridization studies showed that aggrecan gene expression in the cells of the resting/proliferative zone and the upper hypertrophic zones were similar, and was reduced in the deepest cells of the lower hypertrophic zone, adjacent to the zone of calcification. Proteoglycan structure was characterized by associative and dissociative Sepharose CL2B chromatography. These results showed that approximately 90% of the newly synthesized proteoglycan, and the total proteoglycan population, was able to aggregate and that the monomers were relatively large. The proteoglycan released into the media had a reduced ability to aggregate and the monomers were of a more variable size. These data support the hypothesis that the matrix proteoglycan content is controlled both by the rate of synthesis and breakdown, but in the lower regions the rate of synthesis may play a more dominant role. The higher metabolic activity of aggrecan in the calf than fetal growth plate may be a result of environmental stimuli (i.e., soluble mediators, loading) during different stages of development.
Aggrecans↗
The effects of matrix compression on proteoglycan metabolism in articular cartilage explants.
The effects of compressive stress on the rate of proteoglycan synthesis and release were determined in bovine articular cartilage from 4-5-month-old animals. Full depth cartilage explants were compressed in an unconfined configuration at various stresses ranging up to 1.0 MPa. At mechanical equilibrium (after 24 h), no significant changes were detected in the rate of [35S]-sulfate (35SO4) incorporation at the low level of compressive stresses used (less than 0.057 MPa). At an intermediate level of compressive stress (0.057, 0.1, 0.5 MPa), 35SO4 incorporation rates were reduced to approximately 60% of control values. At the highest level compressive stress (1.0 MPa) studied, 35SO4 incorporation rates were further reduced to approximately 20% that of controls. Recovery experiments at intermediate stress levels showed increased rates of 35SO4 incorporation at 24 h after compression. In explants loaded for 24 h at stresses of 0.1 MPa or higher, there was a stress-dose dependent inhibition of proteoglycan release into the media (up to 61% at 1.0 MPa), and proteoglycan release rates did not return to control values following a 24 h recovery period. While cartilage composition and biosynthetic activity were found to vary significantly with depth in control cartilage, the observed suppression (% change) in biosynthetic activity was relatively uniform with depth in both loading and recovery experiments. The study indicates that compression of the tissue to physiological strain magnitudes serves as a signal to modulate chondrocyte biosynthetic and catabolic responses through the depth of cartilage, while prolonged compression at higher strains may be responsible for tissue and cell damage.
Animals↗
Restoration of Injured or Degenerated Articular Cartilage.
Intra-articular fractures, ligamentous and meniscal injuries, and articular cartilage breakdown are major causes of degenerative joint disease. Lesions on the articular surface seem to have a limited capacity for repair and often progress inexorably toward osteoarthritis. Recent studies on joint immobilization and cartilage atrophy, however, have shown that repair and remodeling of articular cartilage may be possible. Currently used clinical methods of stimulating cartilage repair and remodeling include alteration of the loading on degenerated joints (primarily by using osteotomies), introduction of new cartilage-forming cells by perforation of subchondral bone, and soft-tissue arthroplasty. These procedures provide temporary relief in selected patients, but they often do not predictably restore long-term joint function. Experimentally, cartilage repair has been stimulated successfully with the use of allografts of periosteum and perichondrium, which serve as sources of cells with chondrogenic potential; introduction of cells grown in culture (stem cells or chondrocytes); stimulation by fibrin clot formation; artificial collagen matrices combined with cell transplants; and chondrogenic growth factors. The long-term success of all these methods has not been explored thoroughly, even in animal studies. Nevertheless, some research results are sufficiently encouraging to suggest that repair of the degenerating articular cartilage may be possible in the future.
Journal Article↗
Estrogen protection against bone resorbing effects of parathyroid hormone infusion. Assessment by use of biochemical markers.
OBJECTIVE: Because parathyroid hormone (PTH) stimulates bone resorption, resistance to its actions might help maintain bone mass. We tested the hypothesis that the effects of estrogen on bone are accomplished in part by decreasing the sensitivity of the skeleton to the resorbing effects of PTH. STUDY DESIGN: Comparison of response to PTH infusion in untreated and estrogen-treated postmenopausal women with osteoporosis. INTERVENTION: (1-34) human PTH, 0.55 U/(kg.h), was infused intravenously over 20 hours. SETTING: The inpatient clinical research unit of a referral hospital. PATIENTS: Women with primary postmenopausal osteoporosis who were untreated (n = 15) or treated with estrogen (n = 17). MAIN OUTCOME MEASURES: Skeletal turnover indices including hydroxyproline, deoxypyridinoline, pyridinoline, tartrate-resistant acid phosphatase, alkaline phosphatase, bone Gla protein, and insulin-like growth factor-1. RESULTS: All basal indices were higher in untreated than in estrogen-treated women, but statistical differences were seen only for deoxypyridinoline and pyridinoline. During the 20-hour infusion, hydroxyproline/creatinine increased 0.023 mumol/mumol in untreated women but only 0.010 mumol/mumol in estrogen-treated women (P < 0.05). Corresponding changes for deoxypyridinoline/creatinine were 14.6 mumol/mumol and 3.5 mumol/mumol (P = 0.06). Tartrate-resistant acid phosphatase and pyridinoline increased only in untreated group. A circadian rhythm in circulating bone Gla protein was seen in both groups without clear PTH-induced effects or differences between groups. Alkaline phosphatase levels decreased and insulin-like growth factor-1 levels increased in both groups with no distinction between untreated and estrogen-treated women [corrected]. CONCLUSION: The estrogenized postmenopausal osteoporotic skeleton is less sensitive to the bone resorbing effects of acutely administered PTH. There are no differential effects on bone formation.
Aged↗