Ole W. Wiebkin

Glycosaminoglycans of human gingival epithelium and connective tissue.

The glycosaminoglycan (GAG) components of the proteoglycans (PG) in the epithelial and connective tissue extracellular compartments of human gingivae have been determined. Following proteolytic digestion of the separated tissues. GAG were identified electrophoretically. These were hyaluronic acid (HA), heparan sulfate (HS), dermatan sulfate (DS) and chondroitin-4 sulfate (ChS-4). Neither ChS-6 nor keratan sulfate (KS) was observed. Confirmation of the nature of the molecular species was obtained by the use of Streptomyces hyaluronidase, chondroitinase AC II and degradation with nitrous acid. Quantitatively, the major GAG component of the epithelial specimens was HS (59.6%), whilst DS (60.6%) constituted the major GAG of human gingival connective tissue.

Surgicel®: Macrophage processing of the fibrous component

Previous reports have demonstrated that Surgicel, a local haemostatic agent, is absorbed from implantation sites. In an earlier study, it was shown that the material consists of a uronic acid component and a fibrous residue. A chemically quantified loss of the uronic acid component within 18 h of implantation was demonstrated. The aim of the present study was to examine the fate of the fibrous residue in rat tissues, using both light and electron microscopy. Results indicated that this fibrous component is phagocytosed by macrophages at the site of implantation. A model for the clearance of Surgicel from tissue implantation sites is presented.

Molecular Weight Estimation of Sulfated Glycosaminoglycans in Human Gingivae

The number-average molecular weights of human gingival epithelium and connective tissue glycosaminoglycans (GAG) have been determined. Radioactive labelled GAG were extracted from separated gingival epithelium and connective tissue following alkaline degradation of the tissue in the presence of tritiated sodium borohydride. They were identified by electrophoresis as heparan sulfate (HS), hyaluronic acid (HA), dermatan sulfate (DS) and chondroitin 4-sulfate (ChS-4). Following densitometric quantitation of the sulfated GAG (HS, DS and ChS-4), the amount of radioactivity associated with each species was determined by liquid scintillation of each band staining positively for these GAG. The number-average molecular weights for each GAG were determined by end group analysis. The values obtained for each sulfated GAG indicated a degree of similarity in molecular weight distribution between the two tissue types ranging from 15,000 to 27,000.

Insulin-like growth factor binding proteins (IGF-BPs) in bovine articular and ovine growth-plate chondrocyte cultures: their regulation by IGFs and modulation of proteoglycan synthesis

Cultured chondrocytes respond to insulin-like growth factors (IGFs) by increasing the production of proteoglycans and insulin-like growth factor binding proteins (IGF-BPs). To investigate the biological effects of IGFs and IGF-BPs, isolated bovine articular and ovine growth-plate chondrocytes were cultured at high density in the presence of IGF-1, and its truncated form, des (1-3) IGF-I. Both growth factors stimulated the production of IGF-BPs in articular and growth-plate chondrocyte monolayers. Western ligand blots showed that bovine articular chondrocytes released two forms of IGF-BPs into conditioned medium with molecular weights of 29 and 31 kDa. Ovine growth-plate chondrocytes released four different forms of IGF-BPs of approx. 22, 24; 29-30 and 34 kDa. IGF-I and des (1-3) IGF-I stimulated total proteoglycan synthesis by articular chondrocytes up to 1.5-fold. The truncated analogue was more potent at lower concentrations, particularly in stimulating incorporation of newly synthesized proteoglycans into the cell-layer. The maximal stimulation of proteoglycan synthesis in ovine growth-plate chondrocyte culture was 3-fold with des (1-3) IGF-I, while IGF-I enhanced proteoglycan production by only 2-fold over the concentrations used. Our results suggest that endogenous IGF-BPs in chondrocyte cultures act as a part of a feed-back mechanism which diminishes the bioactivity of IGF-I.

Behavior of Hyaluronic ACID from Gingival Epithelium and Connective Tissue on the Analytical Ultracentrifuge

The molecular weights of hyaluronic acid (HA) isolated from separated specimens of human gingival epithelium and connective tissue as well as standard hyaluronic acid preparations have been estimated. The values were determined following substitution of sedimentation values into a previously determined empirical relationship between the reciprocal of the sedimentation coefficient at zero concentration (S-1)0 and molecular weights estimated by sedimentation-diffusion (MsD). The values of (S-1)0 for connective tissue and standard low-molecular weight HA preparations obtained by linear regression of all points indicated molecular weights (MsD) of 340,000 and 205,000 respectively. However, epithelial and standard high-molecular weight HA behaved differently during ultracentrifugation generating a curvilinear relationship between s-1 and concentration. Nevertheless linear extrapolation of a line of best fit of the very lowest concentrations (those which approached zero concentration) provided molecular weight estimates of 860,000 and 2,500,000 respectively. Moreover, similar treatment of s-1 values derived from the previously published data of Laurent, Ryan and Pietruszkiewicz has validated the use of linear regression of s-1 at the lower concentrations alone, to calculate high-molecular weight HA. The curvilinear relationship for s-1 throughout the whole concentration range (0.15-2.3 mg/ml) has been regarded only as a qualitative indication that the HA samples are of relatively high-molecular weight, while a straight line through such data points implies a qualitatively lower molecular weight for HA.

Experimental forward mandibular displacement in sheep.

In order to investigate growth modifications of the temporomandibular joint (TMJ) during dentofacial orthopaedic treatment, specific functional appliances have been used experimentally to prompt the mandible into a protrusive position in various animal models. The purpose of this study was to test the effectiveness of a functional appliance specially designed for sheep and to evaluate the sheep as a model for dentofacial orthopaedic research. Eight, 4-month-old, castrated male Merino sheep were randomly assigned to experimental or control groups, with four in each group. Cast functional appliances were fabricated for the animals in the experimental group. The treatment period was 15 weeks. Dental casts, endosseous implant markers and cephalograms were used to analyse the displacement of the mandible. Undemineralised sagittal sections of TMJ were used to evaluate the tissue responses induced by the appliances. The weight of the animals was measured monthly to monitor their growth. The growth of the metacarpus was also measured. During the experimental period, the animals maintained their weight within the normal range and grew normally. The appliance displaced the mandible to a downward and forward position. The adaptive responses in the TMJ induced by the appliances included changed anteroposterior shape of the condylar process, anteriorly thickened condylar cartilage, and a thickened compact bone layer along the anterior surface of the posterior wall of the glenoid fossa. The sheep coped well with the experimental procedures and the appliance used was demonstrably effective in inducing adaptive responses in the TMJ. Consequently, it is believed that the sheep is an appropriate animal model to study growth modifications in the TMJ region.

Identification of basal lamina acidic glycoconjugates, particularly heparan sulphate proteoglycans, using a poly-l-lysine-gold probe in induced oral carcinomas

Acidic glycoconjugates represent the major non-fibrous macromolecular components that form the extracellular and cell-associated matrices of all animal tissues. The constituent molecules are principally structural glycoproteins and proteoglycans. While their protein component is determined by gene pools, it is the polyanionic (acidic) nature of the polysaccharides, determined by their degrees of carboxylation and sulphation, which confers both functional and diagnostic status on these molecules. Sulphated glycoconjugates in the basal laminae have been reported to play a role in tumour invasion and metastasis. In this study, we used cationic colloidal gold together with transmission electron microscopic methods to compare the expression of acidic glyconconjugates in the basal lamina of both normal rat tongue mucosa and experimentally induced oral carcinomas. Results indicated that heparan sulphate rich glycoconjugates were predominant and were mostly confined to the lamina lucida of the basal lamina in normal oral mucosa. Conversely, observation of basal laminae associated with induced carcinomas showed less intense and more widely dispersed gold labelling for heparan sulphate. The observed differences in gold labelling may reflect modified metabolism of sulphated glycoconjugates or result from the action of degradative enzymes in the induced tumours.