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Biomedical subjects

K G Vogel

Publications and source records attributed to K G Vogel.

At least 19 recordsLinked to original sources
PubMed1999

In situ expression of collagen and proteoglycan genes during development of fibrocartilage in bovine deep flexor tendon.

A region of fibrocartilage develops in bovine deep flexor tendon where the tissue wraps around bone and is subjected to compressive and shear forces in addition to tension. There is no fibrocartilage at this location in fetal tendon or in adjacent adult tendon that is subjected to tensional load only. We investigated the development of fibrocartilage in tendon using in situ hybridization to localize cells that express collagen and proteoglycan genes typical of either tendon or cartilage. The signal for type I collagen and decorin was high in cells throughout fetal and newborn tendon, as is expected in a growing tissue composed predominantly of type I collagen. No signal for aggrecan was seen in either fetal or newborn tendon. No hybridization with any of the probes for collagen or proteoglycan was detected in cells in the tensional region of adult tendon, indicating that the cells in this tissue are normally quiescent. However, the cells in the fibrocartilage of adult tendon displayed a high level of expression for types I and II collagen, decorin, biglycan, and aggrecan. This suggests that the fibrocartilage in adult tendon is a dynamic tissue. Expression of type IIA collagen is considered a marker of prechondrocytes. Type IIA collagen gene expression was present throughout both the tensional and compressed regions of fetal and newborn tendon but was absent in cartilage and adult tendon. This suggests that cells located throughout fetal tendon may have the capacity to develop as chondrocytes. Fibrocartilage signal was detected for type I collagen in 75% of the cells and for type II collagen in 50% of the cells at one location in adult tendon, suggesting that some cells in this tissue could have expressed mRNA for both type I and type II collagen.

Animals↗
PubMed1998

Centromeric protein B null mice are viable with no apparent abnormalities.

The centromere protein B (CENP-B) is a centromeric DNA/binding protein. It recognizes a 17-bp sequence motif called the CENP-B box, which is found in the centromeric region of most chromosomes. It binds DNA through its amino terminus and dimerizes through its carboxy terminus. CENP-B protein has been proposed to perform a vital role in organizing chromatin structures at centromeres. However, other evidence does not agree with this view. For example, CENP-B is found at inactive centromeres on stable dicentric chromosomes, and also mitotically stable chromosomes lacking alpha-satellite DNA have been reported. To address the biological function of CENP-B, we generated mouse null mutants of CENP-B by homologous recombination. Mice lacking CENP-B were viable and fertile, indicating that mice without CENP-B undergo normal somatic and germline development. Thus, both mitosis and meiosis are able to proceed normally in the absence of CENP-B.

Animals↗
PubMed1998

Proteoglycan synthesis by fibroblasts from different regions of bovine tendon cultured in alginate beads.

The ability of cell shape to modulate proteoglycan synthesis in tendon fibroblasts was investigated by placing freshly isolated tendon fibroblasts and chondrocytes into primary culture either as adherent cells on a polystyrene substratum or as rounded cells in alginate beads. Chondrocytes and cells from the compressed region of adult tendon synthesized predominantly large proteoglycan when maintained either as dense monolayers, where actin stress fibers in the cytoskeleton were prominent, or in alginate beads, where actin fibers could not be detected. After three rounds of proliferation as elongated adherent cells the synthesis of large proteoglycan was greatly reduced, i.e. the chondrocytic cells underwent 'dedifferentiation'. Cells from the tensional region of adult tendon synthesized predominantly small proteoglycan when in primary culture as a monolayer, after proliferation on a flat substratum, or as round cells in alginate beads. Fibroblasts from the tensional region of newborn tendon showed no tendency toward increased synthesis of large proteoglycan when maintained as round cells in alginate beads for 7 weeks. In tendon there appears to be a mechanically induced developmental transition from fibroblastic to chondrocytic cells. However, neither the change to a rounded cell shape nor the lack of organized cytoskeletal actin fibers was sufficient to induce chondrocyte-like proteoglycan synthesis in differentiated tendon fibroblasts in culture.

Actins↗
PubMed1997

Mechanical loading and TGF-beta regulate proteoglycan synthesis in tendon.

Fibrocartilage is found in tendon at sites where the tissue is subjected to transverse compressive loading in vivo. A significant characteristic of the tissue transition from tendon to fibrocartilage in bovine deep flexor tendon is increased gene expression, synthesis, and accumulation of both a large proteoglycan, aggrecan, and a small proteoglyoan, biglycan. In order to investigate the cellular events involved in this response, segments of fetal bovine deep flexor tendon were subjected in vitro to cyclic compressive load for 72 h. Following loading, the level of aggrecan mRNA in cells from loaded tissue was increased 200-450% compared to matched nonloaded tissue segments, as determined by slot-blot analysis. The level of biglycan mRNA increased 100%, and the level of versican mRNA increased 130% in the loaded tissue. The level of decorin mRNA remained virtually unchanged, while expression of alpha 1(I) collagen increased only 40%. When tissue segments were cultured in the presence of transforming growth factor (TGF)-beta 1 (1 ng/ml), the synthesis and expression of mRNA for both aggrecan and biglycan increased, whereas decorin expression was not affected. Similarity in both the direction and the pattern of the cellular response to mechanical load and TGF-beta suggested a causal relationship. Both loading of tendon segments and TGF-beta treatment increased expression of mRNA for TGF-beta by approximately 40% compared to control tissue. In addition, the amount of newly synthesized TGF-beta immunoprecipitated from extracts of loaded tissue was several-fold greater than that from nonloaded tissue. The experiments of this study support a hypothesis suggesting that one aspect of the response of cells in fetal tendon to compressive load is increased TGF-beta synthesis which, in turn, stimulates synthesis of extracellular matrix proteoglycans and leads toward fibrocartilage formation.

Aggrecans↗
PubMed1997

Biology of the rotator cuff tendon.

Tendons are complex composite material composed primarily of water, collagen, proteolycans, and cells, designed to transmit tensile loads from muscle to bone. Although rotator cuff tendons differ in many ways from other tendons in the body, a knowledge of basic tendon structure and function is helpful in understanding rotator cuff tendon biology, injury, and repair. In addition to type I collagen, rotator cuff tendons contain small amounts of type III collagen, which play a role in healing and repair. In comparison with other tendons, the increased glycosaminoglycan and proteoglycan content seen in rotator cuff tendons may be adaptive, pathologic, or both. The etiology of rotator cuff pathology is probably related to trauma, aging, and degeneration. As our understanding of these processes increases, we will be able to develop and implement improved preventative and therapeutic interventions for rotator cuff pathology.

Animals↗
PubMed1996

Proteoglycans of human rotator cuff tendons.

Rotator cuff and biceps tendons that appeared grossly normal were procured from adult cadavers without a history of shoulder problems. These tendons were analyzed for the amount and type of glycosaminoglycan, type of proteoglycan, and histology. When compared with the distal/tensional region of biceps tendon, the glycosaminoglycan content of supraspinatus, infraspinatus, and subscapularis tendons was 2.5-fold higher and the glycosaminoglycan content of the proximal/compressed region of biceps tendon was 3-fold higher. The ratio of hyaluronic acid to chondroitin sulfate/dermatan sulfate in all three cuff tendons was approximately 1. Rotator cuff tendons contained large proteoglycan similar to aggrecan, as demonstrated by sodium dodecyl sulfate-polyacrylamide gel migration elution from Sepharose CL-4B, and content of both chondroitin sulfate and keratan sulfate chains. Both decorin and biglycan were also present, as demonstrated by migration in sodium dodecyl sulfate-polyacrylamide gels and core protein immunoreactivity. In contrast decorin was the only proteoglycan prominent in distal/tensional regions of biceps tendon. Histological analysis showed layers of loosely organized alcian blue-stained material running between the longitudinal collagen fiber bundles. The proteoglycan content of rotator cuff tendons was similar to fibrocartilage in tendons that have been subjected to compressive loads in situ. This suggests that cells of normal adult rotator cuff tendons have adapted to loads distinct from pure tension. However, the histological organization did not resemble mature fibrocartilage. The increased amount of proteoglycan in rotator cuff tendons may serve to separate and lubricate collagen bundles as they move relative to each other during normal shoulder motion.

Adult↗
PubMed1996

The effect of compressive loading on proteoglycan turnover in cultured fetal tendon.

A fibrocartilaginous tissue develops in tendon at the point where the tendon wraps under bone and is subjected to transverse compressive loading in addition to tension. This tissue is characterized by a high level of large proteoglycan (aggrecan), which could accumulate because of increased synthesis, diminished turnover, or both. To examine the effect of loading on proteoglycan turnover segments of fetal tendon in sterile culture were subjected to cyclic, uniaxial compression loading to 30% of initial thickness once every 6 sec. for 72 h, and then allowed to incorporate 35S-sulfate for 12 h. The rate of loss of newly-synthesized 35S-proteoglycans from tissue was determined during subsequent culture for up to 12 days, with or without continued loading. Proteoglycan was lost from fetal tendon segments rapidly during the first 3 days of culture and slowly thereafter. Loss of newly-synthesized proteoglycan from adult tendon fibrocartilage was linear, with a half life of 12 d. Segments of fetal tendon subjected to cyclic compression before labeling synthesized more proteoglycan. These segments lost a greater percent of labeled proteoglycan to medium during a subsequent 12-day culture period than matched segments that had not experienced loading. Analysis of medium and tissue proteoglycans by SDS polyacrylamide gel electrophoresis and sieve chromatography indicated that small proteoglycans (decorin and biglycan) were retained in both loaded and non-loaded tissue whereas large proteoglycans (migrating in the Vo of a Sepharose CL-4B column) were readily lost. It is concluded that the 3-day loading regimen did not diminish turnover of large proteoglycan. To the contrary, although synthesis of large proteoglycan was enhanced by the loading regimen, these proteoglycans were still rapidly lost from the fetal tissue.

Animals↗
PubMed1994

The in vitro interaction of proteoglycans with type I collagen is modulated by phosphate.

Binding of proteoglycans to type I collagen in vitro was assessed using radiolabeled decorin, biglycan, and large proteoglycans and acid-extracted bovine tendon collagen. Decorin, biglycan, and large proteoglycans were all bound to collagen fibrils in phosphate-buffered saline (PBS) containing 3 mM sodium phosphate. Only decorin was bound when the phosphate concentration in PBS was increased to 30 mM. These distinct binding characteristics were not altered by the presence of 10% serum, by purification of the proteoglycans in 7 M urea and 4 M guanidine HCl, or by digestion of the collagen with pepsin. In addition to being affected by phosphate, both glycosaminoglycan and proteoglycan binding to collagen was inhibited by sulfate, an anion with similar structure, and by molecules that contain sulfate or sulfonate groups, such as chondroitin sulfate and N-tris[hydroxymethyl]methyl-2-aminoethanesulfonic acid (Tes). The rate of in vitro collagen fibrillogenesis was retarded by increasing concentrations of phosphate. Decorin decreased the rate of collagen fibrillogenesis in all buffers and virtually abolished fibril formation when added in buffer containing both 30 mM phosphate and 30 mM Tes. It is concluded that decorin binds to collagen through interaction between collagen and the decorin core protein, whereas biglycan and large proteoglycans bind to collagen fibrils through their glycosaminoglycan chains. This glycosaminoglycan-collagen interaction is inhibited by phosphate, sulfate, and sulfonate. These observations may clarify contradictory results among previous in vitro studies of proteoglycan-collagen interaction. Since the phosphate concentration of blood and interstitial fluid is estimated to be approximately 1 mM, collagen-glycosaminoglycan interactions could occur in tissue.

Aggrecans↗
PubMed1994

Aggrecan in bovine tendon.

Large proteoglycans were purified by ion-exchange chromatography, gel filtration and CsCl gradient centrifugation from the compressed and tensional regions of adult bovine deep flexor tendon. Tryptic peptide maps of proteoglycan from the compressed region were very similar to maps of aggrecan from bovine articular cartilage, with evidence for the presence of all fifteen previously identified markers from the G1, G2 and G3 domains. The presence of aggrecan in these samples was confirmed by sequencing the G1 peptide YPIHTPR. The equivalent maps for large proteoglycan from tensional tendon were also consistent with the presence of aggrecan, and this was confirmed by sequencing three marker peptides from each of the G2 and G3 domains. However, G1 marker peptides were conspicuously absent from tensional samples. Northern blots for aggrecan mRNA showed high levels in cells from compressed tendon and articular cartilage. Extended exposure revealed a lower level of hybridization to RNA from tensional tendon as well. The results confirm that aggrecan, which is similar in core protein structure to articular cartilage aggrecan, is the predominant chondroitin sulfate-bearing large proteoglycan of compressed tendon. The results also indicate that aggrecan fragments lacking the G1 domain can account for the small amounts of chondroitin sulfate-bearing large proteoglycan in tensional regions of adult tendon.

Aggrecans↗
PubMed1994

Regional expression of mRNA for proteoglycans and collagen in tendon.

Regions of tissue that ressemble cartilage develop at the point where tendon wraps under a bone and receives compressive forces in addition to tension. Northern blot analysis was used to assess expression of mRNA for several extracellular matrix constituents in cells immediately after their isolation from tensional and compressive regions of fetal, adult and old adult bovine deep flexor tendon. Messenger RNA for aggrecan and type II collagen, as well as for biglycan and decorin, was highly expressed in cells from the compressed region of adult tendon, indicating that this tissue contains cells with a chondrocytic phenotype. In contrast, only mRNA for decorin was highly expressed in cells from the tensional region of adult tendon. The major developmental changes in mRNA expression in the compressed region of tendon included a approximately 25-fold increase in aggrecan expression between fetal and adult tissue and an increase in type II collagen mRNA from undetectable in fetal tendon to expression in adult tendon that was nearly as high as the level expressed in cells from adult articular cartilage. Developmental changes in the tensional region consisted of a approximately 10-fold decrease in type I collagen expression and a 4-fold increase in decorin expression in cells from adult tissue, as compared to fetal tissue. These observations indicate that levels of gene expression for proteoglycans and collagen in tendon correlate with the mechanical environment in the tissue. The regional distinctions in mRNA expression were lost when cells were grown in monolayer culture for one week.(ABSTRACT TRUNCATED AT 250 WORDS)

Aggrecans↗
PubMed1993

Proteoglycan synthesis in fetal tendon is differentially regulated by cyclic compression in vitro.

The predominant proteoglycan in tensional regions of tendon is the small proteoglycan decorin. However, a fibrocartilaginous tissue containing large amounts of aggrecan and biglycan develops at points where tendon wraps under bone and is subjected to compressive loading in addition to tension. The hypothesis that local compression regulates the development of fibrocartilage in tendon was tested by assessing the effect of in vitro compressive loading on proteoglycan synthesis. Fetal bovine deep flexor tendon explants from the region which would have become fibrocartilage were subjected to 3 days of continuous cyclic uniaxial compression (unconfined) to 30% strain, at a frequency of 1 cycle/6 s (0.17 Hz). Compression was perpendicular to the long axis of the tendon. Large proteoglycan, biglycan, and decorin were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and [35S]sulfate incorporated into each proteoglycan was quantitated by liquid scintillation counting of gel slices. The primary effect of compression was to stimulate selectively synthesis of large proteoglycan and biglycan. Incorporation of [35S]sulfate into large proteoglycan was increased 100-300% and incorporation into biglycan was increased 50-150% in compressed tissue compared to matched uncompressed tissue segments. Incorporation into decorin was unchanged. A similar effect on radio-sulfate incorporation was seen following loading of tissue from the tensional region of tendon, which does not normally develop into fibrocartilage. Proteoglycans from compressed tissue were larger, due to slightly longer glycosaminoglycan chains. Disaccharide analysis showed that the C6S/C4S ratio was higher in both the large and the small proteoglycan populations from compressed tissue. Aggrecan mRNA levels were increased approximately fivefold in loaded tissue, and SDS-PAGE analysis of [3H]leucine-labeled core proteins indicated that large proteoglycan core protein synthesis was increased by compression. The selective changes in large proteoglycan and biglycan synthesis, and in the sulfate composition and size of the glycosminoglycan chains, are consistent with what might be expected during development of fibrocartilage in vivo. These observations support the hypothesis that compressive force can regulate the development of fibrocartilaginous tissue in tendon.

Animals↗
PubMed1993

Proteoglycans in the compressed region of human tibialis posterior tendon and in ligaments.

Proteoglycan content and tissue morphology were examined in tendons and ligaments from 24 cadavers, ranging in age at the time of death from 1.5 months to 83 years. The region of the human tibialis posterior tendon that passes under the medial malleolus was characterized by cells having a rounded shape, positive staining with alcian blue, and higher glycosaminoglycanuronic acid content than in the more proximal region of the same tendon. Analysis of proteoglycans by sodium dodecyl sulfate/polyacrylamide gel electrophoresis indicated that the predominant small proteoglycan of the proximal/tensional region was decorin, whereas two types of small proteoglycans (decorin and biglycan) and large proteoglycans were present in the region passing under the medial malleolus and presumably subjected to compressive and shear forces in addition to tension. The pattern of proteoglycan accumulation in the compressed region of tendon was basically similar for all individuals and showed no distinctive trends related to age after puberty. In terms of type and amount of proteoglycan, the patellar tendon was like the tensional region of the tibialis posterior. Glycosaminoglycan content in the lateral collateral ligament and anterior cruciate ligament, however, was twofold higher than in the tendons. The ligaments contained large as well as small proteoglycans, just as in the compressed region of tendon.

Adolescent↗
PubMed1992

The interaction of decorin core protein fragments with type I collagen.

To further define the molecular interaction between decorin and type I collagen we generated a 20 kD fragment containing the N-terminal half of the core protein by Endoproteinase Arg C digestion and a 40 kD fragment including all leucine-rich repeats in the central part of decorin core by cleavage with 2-nitro-5-thiocyanobenzoate. The fragments did not influence collagen fibril formation, even at high concentration, and radioactive fragments showed little binding to collagen fibrils. Our observations suggest that neither the N-terminal half nor the central leucine-rich repeats of the decorin core protein can, by itself, interact fully with fibrillar collagen.

Animals↗
PubMed1992

Compression loading in vitro regulates proteoglycan synthesis by tendon fibrocartilage.

The regulation of proteoglycan synthesis in a fibrocartilaginous tissue by mechanical loading was assessed in vitro. Discs of bovine tendon fibrocartilage were loaded daily with unconfined, cyclic, uniaxial compression (5 s/min, 20 min/day) and the synthesis of large and small proteoglycans was measured by incorporation of [35S]sulfate. All discs synthesized predominantly large proteoglycan when first placed in culture. After 2 weeks in culture nonloaded discs synthesized predominantly small proteoglycans whereas loaded discs continued to produce predominantly large proteoglycan. The turnover of 35S-labeled proteoglycan was not significantly altered by the compression regime. Increased synthesis of large proteoglycans was induced by a 4-day compression regime following 21 days of culture without compression. Inclusion of cytochalasin B during compression mimicked this induction. Autoradiography demonstrated that cell proliferation was minimal and confined to the disc edges whereas 35S-labeled proteoglycan synthesis occurred throughout the discs. These experiments demonstrate that mechanical compression can regulate synthesis of distinct proteoglycan types in fibrocartilage.

Animals↗
PubMed1992

The effects of transforming growth factor-beta and serum on proteoglycan synthesis by tendon fibrocartilage.

The effects of transforming growth factor-beta (TGF-beta) and serum on proteoglycan synthesis by tissue explants from the fibrocartilaginous region of adult bovine tendon and by cells in culture from this region were assessed. The most characteristic effect of added TGF-beta on both explant tissue and cells in culture was enhanced synthesis of one small proteoglycan-biglycan. Lowered serum concentration diminished incorporation of Na2 35SO4 into proteoglycans. Added TGF-beta (1 ng/ml) stimulated cell proliferation, increased overall proteoglycan synthesis, and increased the length of glycosaminoglycan chains on all secreted proteoglycans. The effect of TGF-beta on cells in culture was highly consistent whereas explants from different animals showed greater variability in the response. It was concluded that TGF-beta did not specifically promote or maintain the cartilaginous nature of this tissue because supplementing medium with TGF-beta did not significantly alter the ratio of large/small proteoglycans synthesized by tissue explants. However, the observation of enhanced biglycan synthesis by TGF-beta suggests that TGF-beta could be involved in differentiation of regions of tendon subjected to compression, because compressed tendon contains both decorin and biglycan small proteoglycans whereas tensional tendon contains primarily decorin. Excess decorin added to cell culture medium did not affect the ability of TGF-beta to enhance synthesis of biglycan.

Amino Acid Sequence↗
PubMed1990

Ultrastructure and proteoglycan composition in the developing fibrocartilaginous region of bovine tendon.

Clear distinctions in morphology and proteoglycan composition have been described in regions of adult tendon that pass under bone and are subjected to compressive as well as tensional forces. In this study, developing bovine deep flexor tendon from early fetal stages through 6 months of age was examined biochemically and by light and electron microscopy. Longitudinal collagen fibers were seen in the tensional region of tendon throughout development; whereas a well established network arrangement of collagen fibers with wide interfibrillar spaces was seen in the compressed region by 7 months of fetal age. Collagen fibril diameters of both regions increased with age with the mean diameter in tensional tissue always greater than in compressed tissue. Glycosaminoglycan hexosamine content of the tensional region remained low throughout development (approximately 0.2% of dry tissue weight), but increased in the compressed region from 0.4% of dry weight at the 7-month fetal stage, to 1.0% dry weight at 6 months. Keratan sulfate was not detectable in tensional tendon at any age as measured by inhibition ELISA, but was found in increasing quantities in the pressure bearing region of tendon from young calves. Small proteoglycans predominated in both tensional and compressed regions throughout fetal and early neonatal development, and were of both PG I (biglycan) and PG II (decorin) types. Large proteoglycans represented only a small proportion of total proteoglycans in both regions of fetal tendon. By SDS/PAGE analysis, immunoreactivity, and molecular sieve chromatography, large proteoglycans of fetal compressed tendon were similar to large proteoglycans of adult tensional tendon in that they contained only chondroitin-6-sulfate, with little if any KS, and appeared to be slightly smaller than cartilage large proteoglycans.(ABSTRACT TRUNCATED AT 250 WORDS)

Age Factors↗
PubMed1989

Characteristics of the in vitro interaction of a small proteoglycan (PG II) of bovine tendon with type I collagen.

Binding of the small dermatan sulfate proteoglycan of bovine tendon (PG II type/decorin-like) to type I collagen was characterized in an in vitro fibril-forming assay, using native collagen prepared from bovine tendon by acid extraction and radiolabeled proteoglycans synthesized by bovine tendon fibroblasts in culture. Substantial binding to collagen was noted for both intact small proteoglycan and core protein from which the glycosaminoglycan chain was removed. However, binding to collagen was minimal for free glycosaminoglycan chains or large proteoglycans. Binding of the small proteoglycan was optimal at approximately physiological conditions of salt concentration and pH. Scatchard analysis showed a binding affinity constant of 3.3 x 10(7) M-1 with 0.054 proteoglycan binding sites/collagen molecule, when about 0.25-6 micrograms proteoglycan was combined with 100 micrograms collagen. Binding to preformed fibrils of native tendon collagen and to pepsin-treated bovine skin collagen was similar to binding to native tendon collagen. Binding occurred in non-ionic detergents at concentrations up to 1% and once bound, the proteoglycan was not released by washing with up to 2 M NaCl. When both PG I and PG II small proteoglycans were added to collagen, only PG II was bound. This difference is not readily explained by differences in disulfide bond position. These studies indicate a strong, specific interaction between type I collagen fibrils and the core protein of the small (PG II) proteoglycan of tendon.

Animals↗