R.C. Herold

Phylogenetic distribution of enamel proteins: Immunohistochemical localization with monoclonal antibodies indicates the evolutionary appearance of enamelins prior to amelogenins

SummaryThe hard covering tissues, enamel or enameloid, of representative vertebrate teeth were immunohistochemically stained using specific monoclonal antibodies against bovine amelogenins and bovine enamelins in order to determine the phylogenetic distribution of enamelin and amelogenin proteins. Immunohistochemically, only enamelin proteins were present in lower vertebrate (shark, bony fish, and larval amphibian) teeth and dermal denticles. Both enamelin and amelogenin proteins were present in higher vertebrate (mammal, reptile, and adult amphibian) teeth. Large hydroxyapatite crystal size and high levels of mineralization, characteristics common to both enamel and enameloid, are probably due to the presence of the common protein enamelin. The evolution of enamel from enameloid in the tetrapods seems to have involved the development of the gene for amelogenin.

Inhibition of Programmed Cell Death in Mouse Embryonic Palate in Vitro by Cortisol and Phenytoin: Receptor Involvement and Requirement of Protein Synthesis

Abstract In an in vitro model Cortisol and phenytoin inhibit the precisely timed process of palatal development, the lysosomally mediated cell death of the medial edge palatal epithelium. This inhibition of programmed cell death of the palatal midline epithelium by each drug is virtually completely blocked by the antiglucocorticoid, cortexolone, whose blocking action results from competitive binding of the glucocorticoid receptor site. The inhibition produced by each of these drugs is prevented by the protein synthesis blocker, cycloheximide. Thus, blockade of programmed cell death by each of these drugs involves the glucocorticoid receptor site and requires protein synthesis.

Development and ultrastructural changes of sarcosomes during honey bee flight muscle development

Abstract The indirect flight muscle of the honey bee forms by the accumulation of myoblasts into a syncytium. Incorporation of myoblasts into the muscle fiber continues during much of the pupal period. The mitochondria of the honey bee flight muscles have the following structure during developmental stages: Myoblast mitochondria are ovoid and are incorporated in the muscle fiber along with the cytoplasm. The early pupal stage muscle fiber contains ovoid and filamentous mitochondria. The middle pupal muscle fiber contains filamentous, ovoid and vesicular mitochondria with few internal tubules. The late pupal stage muscle contains only ovoid mitochondria containing granules and some internal tubules. The newly emerged honey bee muscle contains ovoid mitochondria containing many internal tubules and a few granules. The mature honey bee muscle contains large ovoid mitochondria (sarcosomes) packed with internal tubules. Flight muscles sarcosomes increase in size and show a twelvefold increase in volume from late pupal period to adult maturation. Correlation of sarcosome ultrastructure and cytochrome changes is carried out.

Ultrastructure of odontogenesis in the pike (Esox lucius). Role of dental epithelium and formation of enameloid layer

The role of the dental epithelial cells in the formation of teeth in the pike (Esox lucius) was investigated by electron microscopy. Initial dentine deposition, consisting primarily of bundles of coarse collagen bundles, formed under the influence of relatively undifferentiated dental epithelial cells similar to those related to initial dentine formation in developing mammalian and amphibian teeth. Following initial tooth dentine deposition, a calcifying granular enameloid layer, 0.5 μm thick, formed on the tooth surface between the inner dental epithelial cells (IDE) and the dentine. The inner dental epithelial cells which produced the enameloid matrix showed ultrastructural features similar to secreting mammalian ameloblasts. The thin calcified surface enameloid layer in pike teeth is regarded to be similar to true ectodermal enamel. It lacked collagen, was produced by IDE cells (ameloblasts), and showed crystal arrangement perpendicular to the tooth surface. The differentiation of the outer dental epithelium, which occurred at the same time as enameloid deposition and tooth calcification, showed fine structure specialization for efficient transport of ions and water. An uncalcified cuticle, produced by the IDE, is deposited on the calcified tooth enamel surface.

Immunofluorescent localization of amelogenins in developing bovine teeth

Abstract Antiserum was prepared to the proteins (amelogenins) isolated from fetal bovine enamel matrix. This antiserum was used to localize the amelogenins in the developing bovine molar by immunofluorescent microscopy. Amelogenins could be identified in the preameloblasts before enamel matrix deposition had begun as well as in the secretory ameloblasts. The closely adherent layer of stratum intermedium cells also contained some immunoreactive material, suggesting that they may contribute protein to the enamel matrix. The newly deposited enamel matrix consisted of brightly fluorescent particles. Mature enamel matrix did not contain the immunoreactive protein except in a thin layer along the dentino-enamel junction and adjacent to the ameloblasts. No other portion of the tooth bud or other tissues reacted with the specific antiserum.

Cytochrome changes during honey bee flight muscle development

Abstract The progressive changes in the composition of the cytochrome system of the wing muscles during development through pupal stages and early adulthood in honey bees ( Apis mellifera L.) was studied in homogenates with a low temperature difference-spectrum technique. Cytochromes b 5 and a + a 3 are the only ones found during most of the pupal period. The association of this system with myofibril synthesis is discussed. Cytochrome b 5 abruptly becomes undetectable 2 days before emergence. Late pupal and early adult life is characterized by the progressive growth of cytochromes a + a 3 , as well as the appearance and progressive growth of cytochromes b + c 1 and c. This growth parallels the growth of the specialized wing muscle mitochondria, the sarcosomes, and is completed 20 days after emergence. At this time the bee is capable of sustained flight, a faculty that is only gradually achieved during the sarcosome growth period. The association of the adult cytochrome system with the intense energy demand in the wing muscles during flight is discussed.

Production of a monoclonal antibody to enamelins which does not cross-react with amelogenins

SummaryDeveloping enamel matrix contains a complex mixture of proteins whose characterization is essential to an understanding of amelogenesis. It is not known whether each component is the product of an individual gene, or whether they are interrelated by physiologic or artifactual breakdown. To define these relationships, monoclonal antibodies were prepared to enamel proteins and we have previously reported the characterization of six antibodies to amelogenins which did not react with enamelins (Christner et al. (1985) Arch. Oral Biol. 30:849–854). We now report the isolation of antibody to enamelins which stains the enamel matrix but does not cross-react with amelogenins. These results suggest that amelogenins and enamelins are distinct classes of proteins.

Inhibition of Programmed Cell Death in the Fetal Palate by Cortisol

Abstract In an in vitro model, cortisol inhibits a precisely timed process of palatal development, the lysosomally mediated cell death of the medial edge palatal epithelium. These steroid effects occur only in palatal shelves from mouse strains with high incidences of corticosteroid-induced cleft palate. In studies in vivo using steroid-sensitive mice, corticosteroid delays shelf elevation but does not prevent shelf contact and alters lysosomal enzyme distribution in the medial edge palatal epithelia. Thus, susceptibility to corticosteroid-promoted palatal clefting is correlated with the inhibition of programmed cell death in the medial edge epithelium in vitro and in vivo.

Monoclonal antibody and immunogold cytochemical localization of amelogenins in bovine secretory amelogenesis

Amelogenin enamel-protein epitopes in developing incisors were ultrastructurally localized with high specificity resolution. They formed clumps scattered over the enamel organic matrix between the hydroxyapatite crystals, and were also present over islands of stippled or granular material at the forming surface of the enamel matrix demonstrating that this material consists in part of amelogenin enamel protein. The amounts of amelogenin, as judged by labelling density, were not greater in the stippled or the surface crystal-containing matrix as compared to the enamel matrix up to 50 micron deep. Amelogenins were also localized in the rough endoplasmic reticulum. Golgi apparatus and secretory granules of the ameloblasts, which suggests they are merocrine secretions.

Production of a monoclonal antibody to bovine tooth enamel proteins

Several monoclonal antibodies to bovine enamel proteins were produced using the mouse myeloma system. Each antibody recognized the same two protein bands on gel electrophoresis. The clones were tested in situ and clone 185 localized specifically in the enamel layer. Clones 185 and 121 were shown to recognize different antigenic determinants.