Moon-Il Cho

Bacteriolysis of Streptococcus mutans BHT by lysozyme and inorganic anions normally present in human saliva

Abstract Liberation of DNA from Streptococcus mutans BHT was elicited by hen egg-white lysozyme in the presence of low concentrations of additive inorganic anions. Significant lysis was observed at salt molarities within normal salivary concentrations. Lysis increased with anion concentration until apparent saturation plateau levels were attained. Thiocyanate and bicarbonate anions were more effective than chloride and fluoride anions at all salt molarities tested. The order of anion potency in the liberation of DNA was SCN− >HCO−3 >Cl− >F−, although differences between SCN− and HCO−3 as well as between Cl− and F− could be minimized by varying the relative enzyme, anion and bacterial-cell concentration. Although DNA was not released at low anion concentrations when the apparent amount of lysozyme bound per cell was high, liberation did occur at low lysozyme concentrations. After release of cell-bound lysozyme in low-molarity bicarbonate solutions, strain BHT retained approx. 10 times as much enzyme from reaction mixtures with high enzyme concentrations as from those with low. The results suggest that physiologic concentrations of various anions present in saliva may be sufficient to trigger lysis of oral microorganisms by lysozyme.

A freeze-fracture study of ruffle-ended post-secretory ameloblasts.

The post-secretory portion of the rat incisor enamel organ was prepared for routine transmission electron microscopy and freeze-fracture replication in order to define further the structural surface features of the ruffle-ended ameloblasts. Surface views of the distal plasma membrane of the ruffle-ended ameloblasts revealed a well-developed zonula occludens junction with from six to ten rows of tight junctional strands. Gap junctions were also observed just proximal to the tight junctional strands. The membranes of the ruffled border contained a rich supply of intramembrane particles (IMP). The IMPs were approximately 7 to 8 nm in diameter and preferentially located on the P-face profiles of the membrane. The density of IMPs on the membranes of the ruffled border was higher than that on the lateral borders of the cell. It is suggested that the IMPs of the ruffled border may represent enzymatic proteins in the basal cell membrane of absorptive ameloblasts. In addition, the large, highly-developed zonula occludens appeared structurally capable of sealing the intercellular spaces between the ruffle-ended ameloblasts.

Ultrastructural and cytochemical studies of the fate of unsecreted collagen precursors after administration of colchicine to mice.

The administration of colchicine disrupts the normal organization of the Golgi complex and blocks the secretion of collagen precursors in periodontal ligament fibroblasts of the mouse. The fate of the unsecreted collagen precursors contained in Golgi-derived saccules and newly formed dense bodies was followed by electron microscopy. A progressive condensation of saccule content along with phase separation of electron-dense and electron-lucent material was observed. Fusion of saccules with dense secretory bodies gave rise to larger inclusions (zebra bodies; ZB) filled with a combination of electron-dense and electron-lucent material. In some ZB, these materials appeared to polymerize into fibrillar units. The fibrillar units stained with silver methenamine like normal collagenous fibrils. These results suggest that unsecreted collagen precursors accumulate in vesicular compartments within which partial polymerization can occur. This finding may explain some reports of intracellular collagenous fibrils in fibroblasts of pathologically altered connective tissues.

Maturation of human gingival keratinocytes cultured with fibroblasts from keratinizing and non-keratinizing epithelia

When keratinocytes from epidermis, gingiva and buccal mucosa are cultured in vitro they form a stratified squamous epithelium that lacks evidence of orthokeratinization or parakeratinization. We attempted to induce orthokeratinization or parakeratinization in cultured gingival keratinocytes by co-cultivation with fibroblasts from human skin, gingiva and buccal mucosa. Keratinization was defined by the morphological appearance of the cultured cells and by the presence of large molecular weight keratin proteins (63,000 and 67,000 mol. wt). Using these criteria, the chosen fibroblasts failed to induce any alteration in the pattern of keratinization. We conclude that under our present culture conditions, fibroblasts alone cannot induce keratinization in cultured keratinocytes.

Radioautographic analysis of [3H]-fucose utilization by mouse odontoblasts with emphasis on intracytoplasmic and plasma membrane glycoproteins

[3H]-fucose utilization by odontoblasts was studied by light and electron microscopic radioautography. At 10 min after injection, fucose label was concentrated in the Golgi area. By 20-30 min, there was a progressive decline in Golgi labelling with label present at the plasma membrane, terminal web, odontoblast process and predentine matrix. At 4 h, the predentine and the predentine-dentine junction were heavily labelled. At the ultrastructural level, Golgi labelling at 10 min was mostly localized to cisternal elements and at 20 and 30 min secretory granules and dense bodies were also labelled. Most of the silver grains observed in the terminal web were associated with microfilaments near the plasma membrane. In the predentine, the matrix itself accounted for 23.0 per cent of the label at 4 h and the plasma membrane of the odontoblast process accounted for 19 per cent. The results indicate that odontoblasts, in addition to secreting glycoproteins into the dentinal matrix, also continuously manufacture glycoproteins for incorporation into the cell surface, the lysosomal system and the terminal web.

Immuno-electron microscopic study of antigenic surface components of Actinomyces naeslundii in human dental plaque

Abstract Rabbit antiserum to formalin-killed Actinomyces naeslundii I was used to investigate the ultrastructural location and distribution of the antigenic sites of A. naeslundii . Antigenic sites were identified by an indirect technique using goat anti-rabbit IgG coupled to horseradish peroxidase, also used to identify individual cells of A. naeslundii in a mixed bacterial population and in freshly isolated dental plaque. The bound antibody and associated reaction product were visible in ultrathin sections as an electron-dense amorphous material (100–150 nm thick) in juxtaposition to the bacterial cell wall. The location of the immuno-reactants suggested that the antigens are distributed superficially and evenly over the entire bacterial cell surface. Bridge-like extensions of the immuno-reactants connected adjacent cells, suggesting that a limited amount of antigenic material might extend out from the cell wall to provide structural continuity with similar material on adjacent cells.