Analysis of eosinophilic round bodies formed after injection of enamel matrix derivative into the backs of rats.

ABSTRACT

BACKGROUND: Enamel matrix derivative (EMD) is used in dental clinics for the regeneration of alveolar bone. Its effects have not yet been clarified, although it induces eosinophilic round bodies (ERBs) and cartilage formation at the injection site. The objective of this experiment was to examine the histopathologic and biochemical properties of ERBs formed after EMD injection. METHODS: The backs of Sprague-Dawley rats injected with various concentrations of EMD were examined histopathologically. For biochemical examinations, ERBs were microdissected out from the sections. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), matrix-assisted laser-desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS), and database analysis of ERBs were carried out. RESULTS: The histopathological findings were consistent with a foreign body reaction. Numerous ERBs were observed 7 days after injection of 30.0 mg/ml EMD. Histopathologically, ERBs did not contain polysaccharide, amyloid, or hemosiderin. The cells surrounding ERBs were not macrophages or vascular endothelial cells. SDS-PAGE of the microdissected ERBs revealed an intense band at around the 40-kDa region. MALDI-TOF MS showed that the spectrum for ERBs has only a single strong ion intensity. Analysis of the amino acid sequence revealed that the ERBs were composed of various molecular fragments, which all contained an identical seven amino acid sequence. In addition, these peptides are a component of amelogenin. CONCLUSIONS: A high concentration of EMD induces ERBs that consist of a 40-kDa protein which includes a constituent part of amelogenin. The ERBs (or remaining EMD) might promote mesenchymal cell differentiation into hard tissue-forming cells around the EMD injection site.