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Biomedical subjects

A R Poole

Publications and source records attributed to A R Poole.

At least 127 records · Page 7Linked to original sources
PubMed1988

Degenerative changes in dog articular cartilage induced by a unilateral tibial valgus osteotomy.

We present preliminary details of the changes that occur in a new model for the study of degenerative arthritis which does not involve intraarticular surgery. A unilateral valgus tibial osteotomy was performed in a group of 8 skeletally mature dogs of mixed breed and sex. Proximal tibial cartilages, femoral condyles and patellas were examined in operated and unoperated knees of experimental and unoperated dogs. Lesions were examined histologically and analyses were made of 35SO4 uptake and uronic acid content as measures of proteoglycan synthesis and content, respectively. Water contents were also determined. Dogs were examined from 22 to 62 weeks after osteotomy. Angulation induced by the osteoarthritis was varied from animal to animal in this preliminary study. Histologically, degeneration of the patella was observed in 3 of 5 experimental dogs and in both operated and unoperated limbs in two of the dogs. Lesions were seen in both limbs and were most common in the tibial plateau where they were present both in the lateral (covered by meniscus) and medical (uncovered) compartments of the operated and unoperated limb. In the tibial plateau initial increases in proteoglycan content and synthesis were recorded in both limbs: later after 30/36 weeks these declined to normal. In the femoral condyles, increases in water content with angulation were seen in the lateral and medial condyles of operated limbs and in the medial condyles of unoperated limbs. Other changes in proteoglycan content were observed in both condyles with a loss of proteoglycan with increased angulation being recorded in the lateral condyle of the operated limb and an increase in proteoglycan with angulation in the medial condyle of the unoperated limb. These early changes are presumably in response to alterations in load bearing in both limbs. CLINICAL RELEVANCE. This study permits an analysis of degenerative changes that can occur when articulation is changed by an osteotomy. Since the changes occur in both limbs, are potentially reversible and are induced by extraarticular surgery, we have an attractive new model for studying degenerative arthritis.

Animals↗
PubMed1988

Kniest dysplasia is characterized by an apparent abnormal processing of the C-propeptide of type II cartilage collagen resulting in imperfect fibril assembly.

Epiphyseal and growth plate cartilages from four cases of Kniest dysplasia have been studied. In each case collagen fibril organization appeared abnormal by electron microscopy compared with age-matched normal cartilages: fibrils were much thinner, of irregular shape and did not exhibit the characteristic banding pattern. This was associated with the absence (compared with normal cartilage) of the C-propeptide of type II collagen (chondrocalcin) from the extracellular matrix of epiphyseal cartilages, although it was detected (as in normal cartilages) in the lower hypertrophic zone of the growth plate in association with calcifying cartilage. The C-propeptide was abnormally concentrated in intracellular vacuolar sites in Kniest cartilages and its total content was reduced in all cases but not in all cartilages. Moreover, it was not a part of the procollagen molecule. In contrast, type II collagen alpha-chain size was normal, indicating the formation of a triple helix. Also type II collagen content was normal and it was present in extracellular sites and only occasionally detected intracellularly. These observations suggest that the defect in Kniest dysplasia may result from the secretion of type II procollagen lacking the C-propeptide and abnormal fibril formation, and that the C-propeptide is normally required for fibril formation.

Calcium-Binding Proteins↗
PubMed1988

Cell-free translation of mRNA encoding an arterial smooth muscle cell proteoglycan core protein.

The size and immunological reactivity of the primary gene products of a small non-aggregating dermatan sulfate proteoglycan from bovine and monkey arterial smooth muscle cells were examined after cell-free translation of mRNA. Antisera against the dermatan sulfate proteoglycans from bovine articular cartilage, DSPG II [Rosenberg et al. J. Biol. Chem. 260, 6304 (1985)] and human skin fibroblasts [Glossl et al. J. Biol. Chem. 259, 14144 (1984)] were used to show that the unmodified smooth muscle precursor core protein was immunologically related to both the cartilage and fibroblast core proteins. The size of the precursor core proteins within each species was identical regardless of the tissue source. Comparison of the precursor core proteins synthesized by primate and bovine cells revealed that the bovine core proteins were approximately 1500 Da larger than the primate core proteins as determined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. A similar size difference was observed when the mature core proteins of monkey smooth muscle cells and bovine articular chondrocytes were compared after removal of the glycosaminoglycan chains. These results indicate that arterial smooth muscle cells synthesize a dermatan sulfate proteoglycan whose core protein is similar to, if not the same as, the cartilage and fibroblast dermatan sulfate proteoglycan core proteins. These core proteins may be encoded by the same gene that has diverged in size during speciation.

Aggrecans↗
PubMed1987

The identification and characterization of two populations of aggregating proteoglycans of high buoyant density isolated from post-natal human articular cartilages of different ages.

After chromatography on Sepharose CL-2B under associative conditions, high-buoyant-density human articular-cartilage proteoglycans were analysed biochemically and by radioimmunoassay with monoclonal antibodies to a core-protein-related epitope and to keratan sulphate. An examination of proteoglycans from individuals of different ages revealed the presence at 1 year of mainly a single polydisperse population containing chondroitin sulphate (uronic acid) and keratan sulphate. From 4 years onwards a smaller keratan sulphate-rich and chondroitin sulphate-deficient population appears in increasing amounts until 15 years. At the same time the larger population shows a progressive decrease in size from 1 year onward. By 23 years and after the proportion of keratan sulphate in the larger chondroitin sulphate-rich proteoglycan increases. Both adult proteoglycan populations are shown immunologically to aggregate with hyaluronic acid, with the smaller showing a greater degree of interaction. The larger population is richer in serine and glycine, and the smaller population contains more glutamic acid/glutamine, alanine, phenylalanine, lysine and arginine; its protein content is also higher. Whether the larger post-natal population represents a different gene product from the single polydisperse population found in the human fetus, which has a different amino acid composition, remains to be established. The smaller population, which represents approximately one-third the mass of the larger population in the adult, may represent a degradation product of the larger population, in which the hyaluronic acid-binding region and keratan sulphate-rich region are conserved.

Aged↗
PubMed1987

The dermatan sulfate proteoglycans of bovine sclera and their relationship to those of articular cartilage. An immunological and biochemical study.

Dermatan sulfate proteoglycans were isolated from adult bovine sclera and adult bovine articular cartilage. Their immunological relationships were studied by enzyme-linked immunosorbent assays using polyclonal antibodies raised against the large and small dermatan sulfate proteoglycans from sclera and a polyclonal and monoclonal antibody directed against the small dermatan sulfate proteoglycans from cartilage. The small dermatan sulfate proteoglycans from sclera and cartilage displayed immunological cross-reactivity while there was no convincing evidence of shared epitope(s) with the larger dermatan sulfate proteoglycans, nor did these larger proteoglycans share any common epitopes with each other. A hyaluronic acid binding region was detected immunologically on the larger scleral dermatan sulfate proteoglycan but was absent from the larger dermatan sulfate proteoglycan of cartilage and both the small dermatan sulfate proteoglycans. These antibodies were used in immunofluorescence microscopy to localize the scleral proteoglycans and molecules containing these epitopes in the eye. The large scleral dermatan sulfate proteoglycan was restricted to sclera while molecules related to the small scleral and cartilage proteoglycans were found in the sclera, anterior uveal tract, iris, and cornea. Amino acid sequencing of the amino-terminal regions of the core proteins of the small dermatan sulfate proteoglycans from sclera and articular cartilage showed that all the first 14 amino acids analyzed were identical and the same as reported earlier for the small bovine skin and tendon dermatan sulfate proteoglycans. These studies demonstrate that the larger dermatan sulfate proteoglycans of sclera and cartilage are chemically unrelated to each other and to the smaller dermatan sulfate proteoglycans isolated from these tissues. The latter have closely related core proteins and probably represent a molecule with a widespread distribution in which the degree of epimerization of glucuronic acid and iduronic acid varies between tissues.

Amino Acid Sequence↗
PubMed1987

Proteoglycan-induced arthritis in BALB/c mice. Clinical features and histopathology.

Immunization with chondroitinase ABC-digested fetal human cartilage proteoglycan and Freund's complete adjuvant induced polyarthritis and ankylosing spondylitis in female BALB/c mice. The initial external symptoms of the joint inflammation were swelling and redness. This was associated with edema of the synovium and periarticular tissues and gross proliferation of cells, which reached a peak during weeks 7-9 of the experiment. Mononuclear cell infiltration, with perivascular concentration and occlusion of small vessels, was common. Synovitis increased in severity, villous pannus developed, and erosions of bone, articular cartilage, and occasionally, growth plate were observed. The lumbar spine and the proximal intervertebral discs of the tail also exhibited inflammatory and degenerative changes. As the arthritis progressed, sometimes with acute inflammatory exacerbations, more joints became involved and, by the sixteenth to the twentieth weeks of the experiment, a progressive polyarthritis, with gross joint deformities and restricted function, developed in the majority of the limb joints. Clinical and morphologic features of the disease correlated well with radiologic analysis and with an increased deposition of 99mTc-methylene diphosphonate (determined by radionuclide imaging). The development of this arthritis was accompanied by the expression of cell-mediated and humoral immunity to the immunizing antigen. However, this immunity was also observed, although it was generally less well developed, in mice that received the intact or digested proteoglycan without adjuvant. These mice did not usually develop arthritis. Control mice that received only adjuvant did not develop arthritis.

Animals↗
PubMed1987

Immunity to cartilage proteoglycans in BALB/c mice with progressive polyarthritis and ankylosing spondylitis induced by injection of human cartilage proteoglycan.

Intraperitoneal injection of human fetal cartilage proteoglycan (depleted of chondroitin sulfate) in Freund's complete or incomplete adjuvant induces a chronic erosive polyarthritis and spondylitis in all female BALB/c mice. This occurrence is strain-specific but not haplotype-specific, and it is sex-related. The development of the arthritis is associated with the natural presence of cellular immunity to the immunizing antigen and to chondroitinase ABC-treated mouse cartilage proteoglycan. In addition, relatively more antibody to the immunizing proteoglycan is elicited in arthritic mice, and antibodies are produced that cross-react with native mouse proteoglycan. This combination of immune responses is not observed in mice that do not develop arthritis. Associated with the arthritis is the development of cytotoxicity to mouse chondrocytes and, in some animals, of rheumatoid factor, immune deposits in joint tissues and kidneys, and the production of autoantibodies to mouse type II collagen. These observations might be related to our earlier demonstration that immunity to human cartilage proteoglycan is observed in some patients with ankylosing spondylitis.

Adult↗
PubMed1987

The immunologic detection and characterization of cartilage proteoglycan degradation products in synovial fluids of patients with arthritis.

Antibodies were used in radioimmunoassays with gel chromatography to detect the hyaluronic acid-binding region, core protein, and keratan sulfate of human cartilage proteoglycan in the synovial fluids of patients with rheumatoid arthritis, juvenile rheumatoid arthritis, and osteoarthritis. All fluids contained proteoglycan that was mainly included on Sepharose CL-4B; this result indicates cleavage of proteoglycan (which is normally excluded). The hyaluronic acid-binding region was the smallest and most commonly detected fragment. It was relatively free of keratan sulfate and core protein, and it could sometimes bind to hyaluronic acid. Other larger fragments containing core protein and/or keratan sulfate were detected in every fluid.

Aggrecans↗
PubMed1987

The calcification of cartilage matrix in chondrocyte culture: studies of the C-propeptide of type II collagen (chondrocalcin).

We have shown that when chondrocytes are isolated by collagenase digestion of hyaline cartilage from growth plate, nasal, and epiphyseal cartilages of bovine fetuses they rapidly elaborate an extracellular matrix in culture. Only growth plate chondrocytes can calcify this matrix as ascertained by incorporation of 45Ca2+, detection of mineral with von Kossa's stain and electron microscopy. There is an extremely close direct correlation between 45Ca2+ incorporation in the first 24 h of culture and the content of the C-propeptide of type II collagen, measured by radioimmunoassay, at the time of isolation and during culture. Moreover, growth plate cells have an increased intracellular content of the C-propeptide per deoxyribonucleic acid and, during culture, per hydroxyproline (as a measure of helical collagen) compared with nasal and epiphyseal chondrocytes. In growth plate chondrocytes 24,25-dihydroxycholecalciferol (24,25-[OH]2D3), but not 1,25-dihydroxycholecalciferol alone, stimulates the net synthesis of the C-propeptide and calcification; proteoglycan net synthesis is unaffected. Together, these metabolites of vitamin D further stimulate C-propeptide net synthesis but do not further increase calcification stimulated by 24,25-(OH)2D3. These observations further demonstrate the close correlation between the C-propeptide of type II collagen and the calcification of cartilage matrix.

24,25-Dihydroxyvitamin D 3↗
PubMed1987

Age-related changes in the role of matrix vesicles in the mandibular condylar cartilage.

A combined approach of light microscopy, immunofluorescence, transmission electron microscopy and electron energy loss spectroscopy (EELS) was used to study age-related changes in the condylar cartilage in mice. Chondrocalcin, a cartilage matrix calcium-binding protein, was demonstrated by indirect immunofluorescence microscopy using monospecific antibodies. In one week old animals the most intense staining was observed in the matrix around the hypertrophic cells in the mineralising zone, to a lesser degree around the cells in the zone of chondroblasts, while no staining was noted in the zone of chondroprogenitor cells and in the matrix around the early hypertrophic cells. In the mineralisation zone the distribution of chondrocalcin correlated with that of mineral deposits as revealed by the von Kossa stain. The matrix between the early hypertrophic cells as shown by transmission electron microscopy revealed the presence of matrix vesicles and demonstrated a gradual accumulation of hydroxyapatite in the mineralising zone. In one month old animals chondrocalcin localisation was mainly confined to the lower hypertrophic zone which also demonstrated positive von Kossa staining was seen along the articular surface. In older animals multiple electron-dense structure that resembled matrix vesicles were observed in the non-mineralising portions of the condylar cartilage. Use of the EELS method confirmed the almost complete lack of calcium ions in these structures. In contrast, with the use of the same method, detectable amounts of calcium were recorded in vesicles in the mineralising zones of all age groups. Hence what appear ultrastructurally as structures similar to matrix vesicles represent atypical vesicles that might characterise an ageing and degenerative articular cartilage and are not necessarily associated with the mineralisation process.

Aging↗
PubMed1987

Immunodetection and characterization of the degradation of cartilage proteoglycans in vitro and in vivo.

The large aggregating cartilage proteoglycans can be detected immunologically using antibodies directed against the hyaluronic acid-binding region, core protein, keratan sulfate, chondroitin sulfate, and hyaluronic acid. With radioimmunoassays we have detected proteoglycans released from adult human articular cartilage in vitro and in vivo. In vitro cleavage appears to occur mainly adjacent to the hyaluronic acid binding region with the release of the rest of the molecule. Two distinct populations have been observed, the larger of which is chondroitin sulfate-rich and the smaller keratan sulfate-rich. Although they are a little smaller and probably aggregate less with hyaluronic acid, these populations correspond in size and composition to two similar populations detected in situ in healthy adult human articular cartilage. In vivo studies of synovial fluid have revealed that the proteoglycan fragments are smaller and the most commonly detected fragment is the hyaluronic acid binding region. This suggests that in vivo further degradation occurs, probably due to degradation mediated by the inflamed synovium and polymorphonuclear leucocytes present in the synovial fluid of inflamed joints.

Binding Sites↗
PubMed1986

Chondrocalcin is identical with the C-propeptide of type II procollagen.

The primary structure of the cartilage matrix molecule chondrocalcin has been found to be identical with that of the C-propeptide of type II procollagen by comparing sequence analyses of the N-terminal regions and of tryptic peptides derived from chondrocalcin. This implies that in type II collagen the C-propeptide of type II collagen is employed not only in the assembly of the triple helix of type II collagen, as demonstrated previously, but in calcifying cartilage it may also be involved in those events leading to cartilage calcification, as earlier indicated.

Amino Acid Sequence↗
PubMed1986

Age-related changes in protein-related epitopes of human articular-cartilage proteoglycans.

Monoclonal antibodies were prepared that recognize different age-related epitopes on proteoglycan subunits of high buoyant density isolated from human epiphysial and articular cartilages. Antibody EFG-4 (IgG1) recognizes a proteinase-sensitive segment associated with the core protein. Antibody BCD-4 (IgG1) reacts with keratan sulphate bound to core protein. Both epitopes are minimally expressed in foetal cartilage and increase with age after birth to become maximally expressed in adult cartilage by about 30 years of age. In contrast, monoclonal antibody alpha HFPG-846 (IgM) recognizes a core-protein-related epitope that is maximally expressed in young foetal cartilage, declines up to birth and thereafter and is almost absent after about 30 years of age. Antibody alpha HFPG-846 was used to isolate by immuno-affinity chromatography two subpopulations of proteoglycan subunits from a 16-year-old-human cartilage proteoglycan subunit preparation. Only the antibody-unbound population showed a significant reaction with antibodies EGF-4 and BCD-4. The amino acid and carbohydrate compositions of these proteoglycan fractions were different, and one (antibody-bound) resembled those of foetal and the other (antibody-unbound) resembled those of adult proteoglycans isolated from 24-27-week-old-foetal and 52-56-year-old-adult cartilage respectively. These observations demonstrate that human cartilages contain at least two chemically and immunochemically distinct populations of proteoglycans, the proportions and content of which are age-dependent. It is likely that these populations represent the products of different genes, though their heterogeneity may be compounded by the result of different post-translation modifications.

Adolescent↗
PubMed1986

Monoclonal antibodies to different protein-related epitopes of human articular cartilage proteoglycans.

Monoclonal antibodies produced against chondroitinase-treated human adult cartilage proteoglycans were selected for their ability to recognize epitopes on native proteoglycans. Binding analyses revealed that four of these monoclonal antibodies (BCD-4, BCD-7, EFG-4 and KPC-190) each recognized a different epitope on the same proteoglycan molecule which represents a subpopulation of a high buoyant density (D1) fraction of human articular cartilage proteoglycans (10, 30, 50 and 60% in fetal-newborn, 1.5 years old, 15 years old and 52-56 years old cartilages, respectively). Analysis of epitope specificities revealed that BCD-7 and EFG-4 monoclonal antibodies recognized epitopes on proteoglycan monomer which are associated with the protein structure in that they are sensitive to cleavage by Pronase, papain and alkali treatment and do not include keratan sulphate, chondroitin sulphate or oligosaccharides. The BCD-4 and KPC-190 epitopes also proved to be sensitive to Pronase or papain digestion or to alkali treatment, but keratanase or endo-beta-galactosidase also reduced the immunoreactivity of these epitopes. These observations indicate that the BCD-4 and KPC-190 epitopes represent peptides substituted with keratan sulphate or keratan sulphate-like structures. The BCD-4 epitope is, however, absent from a keratan sulphate-rich fragment of human adult proteoglycan, while the other three epitopes were detected in this fragment. None of these four epitopes were detected in the link proteins of human cartilage, in the hyaluronic acid-binding region of human newborn cartilage proteoglycan, in Swarm rat chondrosarcoma proteoglycan, in chicken limb bud proteoglycan monomer and in the small dermatan sulphate-proteoglycan of bovine costal cartilage. EFG-4 and KPC-190 epitopes were not detected in human fetal cartilage proteoglycans, although fetal molecules contained trace amounts of epitopes reactive with BCD-4 and BCD-7 antibodies.

Age Factors↗
PubMed1986

Biological roles of dermatan sulphate proteoglycans.

Dermatan sulphate-containing proteoglycans (DS-PGs) are widely distributed in the extracellular matrix of skin, sclera, tendon, cartilage and a variety of other connective tissues. Two species of dermatan sulphate proteoglycans, called DS-PGI and DS-PGII, have recently been isolated from mature bovine articular cartilages. In their monomeric forms, both DS-PGI and DS-PGII are polydisperse, have relative molecular masses (Mr) ranging from 80K to 140K, and possess protein cores with apparent Mr values of approximately 45K. DS-PGI readily self-associates whereas DS-PGII does not. Polyclonal and monoclonal antibodies against DS-PGII do not react with DS-PGI. DS-PGI and DS-PGII appear to possess different core proteins and represent two different species of dermatan sulphate proteoglycans. DS-PGs have dramatic effects on the biological functions of cells. For example, they inhibit the capacity of fibroblasts to adhere to a fibronectin substratum. BALB/c 3T3 cells were labelled with [3H]thymidine and plated onto dishes coated with plasma fibronectin, plasma fibronectin plus chondroitin sulphate proteoglycan (CS-PG, cartilage-specific proteoglycan monomer), or plasma fibronectin plus DS-PGs. In the absence of proteoglycan, approximately 55% of the cells were attached at 1 h. In the presence of CS-PG, cell attachment was slightly decreased. In the presence of DS-PGs, the adhesion of the fibroblasts to fibronectin was essentially abolished. Similar results were obtained if a plasma fibronectin substratum was preadsorbed with the DS-PGs and the DS-PGs were left in the attachment medium.

Animals↗