Immunohistochemical localization of amelogenins in enameloid of lower vertebrate teeth.

The indirect method of immunofluorescence was used to demonstrate the presence of amelogenins in the enameloid of teeth and dermal denticles of Chondrichthyes; in the enameloid of Teleostei and Amphibia; and in the enamel of Reptilia. Nonmammalian amelogenins are formed in the ectodermal cells of tooth organs and chemically are so similar to mammalian amelogenins that they interact with antiserum prepared from bovine enamel matrix.

Immunocytochemical localization of enamelin proteins in developing bovine teeth.

Enamelins were localized at both the light and electron microscopic level using an antienamelin monoclonal antibody and indirect immunogold methods. Bovine fetal incisors (crown-rump length 17-30 cm) were preserved in Karnovsky's fixative and embedded in Epon. For light microscopy, 2 microM thick sections were immunostained by the indirect method using the monoclonal antibody and goat anti-mouse IgG linked to 5 nM gold particles, followed by silver enhancement to increase the sensitivity of the method. For electron microscopy, thin sections were immunostained (indirect) with the antienamelin monoclonal antibody and goat anti-mouse IgG linked to 5 or 15 nM gold. Control samples were treated with an unrelated monoclonal antibody. The localization of enamelins was confined in the light microscopic sections to the extracellular enamel matrix. No gold staining was observed in the ameloblasts or other enamel organ cells even though the gold-silver technique is extremely sensitive. Ultrastructurally, enamelin was localized in the enamel extracellular matrix and associated ameloblasts. Both the crystal-containing and granular matrix were positively stained, with most gold particles being closely associated with the crystals. Counting of gold particles indicated more than 4 times as many amelogeninas enamelin-reactive antigenic sites in similar regions. Decalcification did not increase immunostaining with the anti-enamelin antibody in the extracellular matrix. Within ameloblasts, the gold particles were associated with secretory granules and Golgi complexes. Thus it appears that enamelins are synthesized in ameloblasts and secreted into the extracellular matrix in a similar manner to amelogenins and are preferentially associated with matrix hydroxyapatite crystals. Transient levels of enamelins within the ameloblasts are apparently too low to be detected by light microscopy.

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.

An immunohistochemical study of the effects of fluoride on enamel development in the rat incisor.

A monoclonal antiamelogenin antibody was used to investigate the effects of fluoride on enamel development in the rat incisor. The results suggested that during secretion the enamel matrix molecules are arranged in such a way as to mask the epitope recognized by the monoclonal antibody. However, during the transition stage of development as the matrix begins to be degraded the epitope becomes exposed and labelling intensity increases to reach a maximum at the end of transition/start of maturation. The effect of fluoride is to delay the appearance of labelling within the enamel matrix until the end of transition. This suggests that the fluoride may inhibit enzymatic degradation or disaggregation of the matrix, the resulting residual matrix then inhibiting crystal growth.

Monoclonal antibodies to different epitopes in amelogenins from fetal bovine teeth recognize high-molecular-weight components.

Enamel proteins were extracted and partitioned into amelogenin and enamelin fractions. Although several attempts were made to raise monoclonal antibodies to each protein fraction, monoclonal antibodies were only obtained against the amelogenin fraction. Six monoclonal antibodies were generated, and these could be classified into three groups recognizing different epitopes by a competitive enzyme-linked immuno-absorbent assay. A model for the arrangement of the epitopes is proposed. In Western-blotting experiments, all six monoclonal antibodies recognized amelogenin components of approx. 45,000 and 60,000 daltons as well as lower molecular-weight components of 10,000 to 30,000. It is proposed that the 45,000 and 60,000 dalton components are precursors of the lower molecular-weight components.

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.

Prosthetic joint infections secondary to rapidly growing Mycobacterium fortuitum.

Infection is an uncommon but catastrophic complication of joint arthroplasty, usually requiring removal of the implant. In a 30-year-old woman a knee arthroplasty was infected with the rapidly growing mycobacterium Mycobacterium fortuitum. Review of other reports of arthroplasties infected with this organism illustrates the problem, diagnosis, and treatment. M. fortuitum is widely distributed in nature, and although usually of low pathogenicity, it can cause infection in conditions of reduced local tissue resistance, i.e. hypodermic abscesses, implant inflammations, and trauma. Only six cases of M. fortuitum prosthetic joint infection have been previously described. Persistent drainage characterized cases in which the prosthesis was left in place. Although antibiotic treatment temporarily suppressed the signs and symptoms of infection, cure required removal of the prosthesis, as in the present case. Diagnosis of M. fortuitum infection is difficult because acid-fast stains of the organisms are often negative. Routinely bacterial cultures are continued for less than about five days, a period not long enough for growth of M. fortuitum. M. fortuitum infections should be considered in draining prosthetic joints with negative bacterial cultures and in those that have had repeated glucocorticoid intraarticular injections.

Abnormal tooth tissue in human odontodysplasia.

The affected teeth in this case of odontodysplasia exhibited abnormal hypoplastic enamel, abnormal dentin containing extensive interglobular regions and completely lacking a peritubular matrix, and an abnormal irregular tissue consisting of highly calcified and partially fused granules located within the dentin of the tooth tips. Electron micrographs of the irregular tissue showed that the granules consisted of crystallites densely and radically packed in a noncollagenous amorphous matrix. Granules were surrounded by a slightly calcified, irregularly arranged, collagenous matrix. The irregular tissue formed in the pulp of the tooth tip befofe and independently of the dentin. It was at least partially formed by pulpal calcification. Abnormal dentin matrix was formed by odontoblasts which were less differentiated than normal. Odontodysplastic teeth showed abnormal differentiation of odontoblasts and ameloblasts resulting in defective enamel and dentin and, in extreme cases, in extensive pulpal calcification.

Odontogenic tumors in mice carrying albumin-myc and albumin-rats transgenes.

Odontogenic tumors that produce abnormal tooth-like structures are repeatedly observed in mandibles of mice that carry both albumin-myc and albumin-ras transgenes. The earliest lesions appear among the periodontal ligament mesenchymal cells, but later lesions include an epithelial component. Subsequent tumor development recapitulates the process of normal tooth formation, which requires multiple sequential cell signals, and results in cell differentiation, matrix secretion, and mineralization. Tumor cells with epithelial morphology produce ras oncoprotein, consistent with an epithelial origin of these tumors. As albumin regulatory sequences direct oncogene expression in these mice, our findings also suggest that some of the albumin present in normal teeth may be locally produced and have a role in tooth mineral formation. The reproducibility of this phenotype makes these mice an excellent model for studies of both normal and neoplastic odontogenesis.