Translocation of enamel proteins from inner enamel epithelia to odontoblasts during mouse tooth development

The developmental problem of how dental epithelia and/or dental papilla ectomesenchyme induce and/or up- or down-regulate tooth formation are as yet unresolved issues. We have designed studies to map the synthesis and fate pathways of secreted amelogenin proteins from Kallenbach differentiation zones II-IV during in vivo and in vitro mouse mandibular first molar tooth development (M1). Tooth organs from cap, bell, and crown stages were processed for reverse transcriptase/polymerase chain reaction (RT-PCR) and high resolution Protein A immunocytochemistry using anti-amelogenin and anti-peptide antibodies. Cap stage M1 were cultured for periods ranging from 10-21 days in vitro using either serum-less, or 15% fetal calf sera-supplemented, chemically-defined medium. Amelogenin transcripts are expressed in the mouse embryonic molar from E15 through early postnatal development. Amelogenin antigens were first detected in Kallenbach's differentiation zone II. Amelogenin proteins secreted from preameloblasts were identified along cell processes and cell surfaces of odontoblasts adjacent to forming mantle dentine extracellular matrix (ECM) prior to biomineralization. Amelogenin proteins were restricted to forming endocytotic vesicles, clathrin-coated vesicles, and lysosomes within odontoblasts. At later stages (e.g. 2 days postnatal development), enamel proteins were not identified in odontoblasts or predentine matrix following mineralization. Comparable observations for stages of development were noted for in vitro cultured tooth explants. Preameloblasts synthesize and secrete amelogenin proteins which bind to odontoblast cell surfaces possibly through the process of receptor-mediated endocytosis. We conclude that amelogenin proteins secreted from preameloblasts, prior to the initiation of biomineralization, were translocated to odontoblasts to serve as yet unknown biological functions.