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Biomedical subjects

J Sodek

Publications and source records attributed to J Sodek.

At least 163 records · Page 9Linked to original sources
PubMed1979

Procollagen synthesis and processing in periodontal ligament in vivo and in vitro. A comparative study using slab-gel fluorography.

A combination of dodecylsulphate/polyacrylamide gel electrophoresis and fluorography has been used to quantify the synthesis of type I and type III collagens by periodontal ligament in situ and periodontal-ligament fibroblasts in vitro. The separation of 14C-labelled collagen alpha chains was achieved by introducing an interrupted reduction step, and the total radioactivity in the alpha-chain bands related to the fluorographic response by a series of standard curves. From these curves an accurate assessment of the relative amounts of type I and III collagen synthesized could be made. The same system also allowed the synthesis and processing of the respective procollagens to be analyzed. For the study in vivo, 200-g male rats were injected with 2 mCi [14C]glycine and killed 0.5-6 h later. Periodontal ligament was dissected from the mandibular molars and the newly-synthesized collagens extracted with 0.45 M sodium chloride. In the study in vitro, confluent monkey periodontal-ligament fibroblasts were cultured in the presence of [14C]proline and [14C]glycine. Analysis of labelled collagens showed a rapid conversion of type I procollagen to collagen but type III collagen was recovered as a procollagen intermediate both in vitro and in vivo. Analysis of duplicate samples after pepsin digestion showed type III collagen synthesis to comprise 15% of the total collagen synthesized in vivo and 20% in early subcultures in vitro. However, the proportion of type III synthesized by the fibroblasts decreased on subculturing. The data demonstrate that fibroblasts in vitro retain the basic characteristics of collagen synthesis and procollagen processing found in vivo, but the overall phenotypic expression of the cells is not stable in culture.

Animals↗
PubMed1979

The effect of oxygen partial pressure on protein synthesis and collagen hydroxylation by mature periodontal tissues maintained in organ cultures.

Mature periodontal tissues from adult-mouse first mandibular molars were cultured in a continuous-flow organ-culture system which allowed the regulation of both ascorbic acid concentration and pO(2) (oxygen partial pressure). Protein synthesis was measured by analysing the incorporation of [(3)H]proline into collagenous and non-collagenous proteins during the last 24h of a 2-day culture. At low pO(2) [16.0kPa (approx. 120mmHg)] approx. 60% of protein-incorporated [(3)H]proline was found in collagenous proteins. However, it was evident that this collagen was considerably underhydroxylated. At high pO(2) [56.0kPa (approx. 420mmHg)], both the amount of collagen deposited in the tissues and the degree of hydroxylation were increased considerably. In contrast, no significant effect on non-collagenous protein was observed. Tissues cultured at low pO(2) for the first 48h were unable to respond to a subsequent increase in pO(2) during the last 24h. Analysis of pepsin-solubilized collagen alpha-chains labelled with [(14)C]glycine demonstrated the synthesis of both type-I and type-III collagens by explants cultured for 48h at high pO(2). Type-III collagen comprised 20-30% of the radioactivity in alpha-chains in both the periodontal ligament and the tissues of the alveolar process. The pattern of protein synthesis in the alveolar tissues at high pO(2) was similar to that observed in these tissues in vivo. However, in the cultured periodontal ligament the proportions of non-collagenous proteins and type-III collagens were increased in comparison with the tissue in vivo.

Alveolar Process↗
PubMed1979

Specific immunohistochemical localization of type I collagen in porcine periodontal tissues using the peroxidase-labelled antibody technique.

Antibody against Type I collagen was raised in rabbits and purified by immunoadsorption on Sepharose-conjugated Types I and III collagen. The cross-reactivity of purified antibody to Type III collagen was found to be less than 0.5% by passive haemagglutination and less than 1.5% by radioimmunoassay. When paraffin sections of fixed and decalcified pig molars were incubated with purified antibody to Type I collagen, varying degrees of staining were observed in the ligament, gingiva, bone and cementum. The periodontal ligament adjacent to bone was more widely stained than that adjacent to cementum in some regions, whereas in others, no difference in staining could be discerned between the two halves of the ligament. The lamina propria of gingiva was stained, and this appeared to be most intense in the vicinity of the overlying epithelium. The fibrous component in the endosteal spaces, the dentine and the extracellular coronal elements in the pulp were generally stained. The impression obtained from the staining pattern is that Type I collagen is not restricted to particular regions of the periodontal ligament or the lamina propria of the gingiva.

Animals↗
PubMed1978

Cleavage of native type III collagen in the collagenase susceptible region by thermolysin.

Viscometric assays were used to demonstrate the activity of thermolysin (EC 3.4.24.4) on native type III collagen in solution. Analysis of the reaction products by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and electron microscopic visualisation of segment long spacing aggregates demonstrated localised cleavage of the collagen in the collagenase susceptible region.

Collagen↗
PubMed1978

Amino acid sequence of rabbit skeletal muscle alpha-tropomyosin. The COOH-terminal half (residues 142 to 284).

Amino acid sequence analysis of the large cyanogen bromide fragment (residues 142 to 281) derived from the COOH-terminal half of the mixed tropomyosin population of rabbit skeletal muscle has been carried out. The isolation and sequence analysis of peptides derived from chymotryptic digests and from tryptic digests of the maleylated fragment permitted the alignment of the complete sequence except for the assignment of acids or amides at residues 142, 144, and 145. Selected peptides from a Myxobacter 495 alpha-lytic protease digest have confirmed certain overlaps. Based on previously published data the sequence can be extended to residue 284, the COOH-terminal end of the protein. In fourteen positions, amino acid substitutions have been observed. In one of these (residue 199) the sequence evidence indicates a minimum of four different polypeptide chains in the mixed tropomyosin population. The assignment of particular amino acid residues to these positions for the major alpha-component of rabbit skeletal tropomyosin has been based on the relative recoveries of peptides containing different residues in these positions.

Amino Acid Sequence↗
PubMed1978

Nature of collagens synthesized by monkey periodontal-ligament fibroblasts in vitro.

1. First subcultures of fibroblast-like cells from adult monkey periodontal ligament were incubated in the presence of 14C-labelled amino acids and produced significant amounts of type-I and type-III collagens. 2. The proportion of type-III collagen produced was calculated on the basis of the recovery of procollagens from DEAE-cellulose chromatography to be approx. 20%, and at least 10% when analysed as collagens on CM-cellulose chromatography. 3. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of the procollagens, the collagens and their CNBr peptides was used to confirm the identity of the collagen types. 4. In serum-free media extensive conversion of type-I procollagen, but not of type-III procollagen, into collagen was observed, suggesting that a specific type-I procollagen peptidase was produced. 5. The pattern of collagen synthesis was not significantly different from that obtained with fibroblasts derived from skin corium of the same animals.

Animals↗
PubMed1978

Isolation and identification of epithelial-like cells in culture by a collagenase-separation technique.

An operational criterion for the identification and isolation of epithelial-like (E) cells, based on their ability to cover and protect a collagen gel from the action of collagenase, has been developed. The E cells isolated by this collagenase-separation technique (CST) exhibited the ultrastructural features, including desmosomes and abundant tonofilaments, that are considered characteristic of this cell type. Unlike confluent cultures of fibroblast-like (F) cells, E cells were not found to have large external transformation-sensitive (LETS) protein on their surface membranes. The CST provides a nondestructive and efficient means of identifying and isolating E cells from mixed populations.

Cell Separation↗
PubMed1976

A new approach to assessing collagen turnover by using a micro-assay. A highly efficient and rapid turnover of collagen in rat periodontal tissues.

Measurement of [3H]proline incorporation into newly synthesized and mature collagen in connective tissues was used to compare rates and efficiency of collagen turnover. The approach minimizes label-recycling problems. By using a micro-assay to determine hydroxyproline specific radioactivities, a highly efficient and rapid collagen turnover in rat periodontal tissues was demonstrated.

Animals↗
PubMed1972

Amino-acid sequence of rabbit skeletal tropomyosin and its coiled-coil structure.

A tentative amino-acid sequence for the COOH-terminal half of rabbit skeletal tropomyosin is reported. These studies confirm our previous conclusions that this tropomyosin consists of several different but similar polypeptide chains. In the sequence, nonpolar residues occur in two series at intervals of seven residues. Amino-acid residues in series I are three residues on the NH(2)-terminal side of, and four residues on the COOH-terminal side of, residues in series II. The presence of occasional charged or ambivalent residues in the positions of series I or II does not lead to a disruption of this long-range pattern. The majority of residues located between the nonpolar residues are charged or polar amino acids. Two highly similar or identical alpha-helices with the reported sequence can be packed together in parallel in a coiled-coil structure. These may be in register or staggered by seven residues or some multiple of it. The observation that groups of small hydrophobic side chains appear to alternate with groups of bulky side chains suggests that a staggered arrangement of the two alpha-helices would maximize the regularity and hydrophobic interactions of the coiled-coil. Model building considerations show that this would occur with a stagger of 14 residues. Such an arrangement could account for the end-to-end aggregation of tropomyosin in solution, and in crystal and tactoid filaments. However, a structure in which the two polypeptides are in register cannot be ruled out.

Amino Acid Sequence↗