(LD) and aperiodic fibrils (APF) separates the two tissues. The POs extend cell processes toward the APFs. (SR) Stellate reticulum; (SI) stratum intermedium.
Epithelial-Ectomesenchymal Morphogenetic Regulation of Odontogenesis
During the 1930s, the science of experimental embryology developed hand-in-hand with advances in organ culture technology. It soon became possible to grow whole and disassociated tooth buds in vitro. Enamel organs, when separated from the dental papillae by trypsin digestion of the basement membrane, were cultured alone or in various recombination with non-oral mesenchymal tissues (Figs 1-9 and 1-10). Isolated cap stage enamel organ, grown either in vivo as a transplant or in vitro in an organ culture system, failed to produce ameloblasts. Dental papilla cells failed to differentiate into odontoblasts unless grown in contact with the enamel organ. These studies established the need for contact between the epithelium (enamel organ) and the ectomesenchyme (dental papilla) as a preliminary condition for the differentiation of ameloblasts and odontoblasts. It was also observed that the dental papilla, once established, controlled the shape of the tooth and gained the ability to direct the differentiation of overlying epithelium (see Figs 1-9 and 1-10).34–36
When it was discovered that the odontogenic inductive interaction could take place across a thin, porous filter, the search for diffusible soluble factors responsible for inducing the differentiation of ameloblasts and odontoblasts became the mission of several dental researchers. In the late 1960s and early 1970s, as the science of molecular biology was being developed, it was speculated that the transfer of informational messenger ribonucleic acid (mRNA) across the basement membrane might control the differentiation of odontogenic cells. In the 1970s, electron microscopic studies showed that cell-to-cell contacts were formed between preodontoblasts and preameloblasts during the cytodifferentiation stage of tooth development. It was proposed that such contacts might provide informational clues responsible for initiating differentiation. Because additional evidence in support of these hypotheses was not forthcoming, attention was directed to the extracellular matrix as a potential communication link between the enamel organ and the dental papilla. This premise was supported by the apparent importance of the basal lamina during odontoblast differentiation.
Fig 1-9 Control of tooth shape by the dental papilla (DP). Dissociation of the enamel organ from the dental papilla by low calcium and trypsin digestion of the basement membrane makes it possible to study the development of various recombinations. Organ cultures of recombined tissues demonstrate the controlling influence of ectomesenchyme (dental papilla) on final tooth form. (EO) Enamel organ. (Based on the findings of Kollar and Baird.34,35)
Fig 1-10 Inductive action of mesenchyme on epithelial differentiation. Organ cultures of dental epithelium recombined with skin mesenchyme develop skin epidermis, complete with skin appendages. When skin epithelium is cultured in contact with dental mesenchyme, a tooth is formed, complete with enamel organ. These results demonstrate the inductive influence of mesenchyme on epithelium. (Based on the findings of Kollar.36)
Role of matrix-mediated signaling
The discovery that enamel organs expressed amelogenin transcripts when cultured on a basement membrane gel, but not when grown on a laminin-coated filter, reinforced the concept that cell-matrix interactions had a permissive effect on gene transcription during tooth development. Research was soon focused on the interactions of cell membrane receptors with specific extracellular matrix ligands as important signaling events that might regulate odontogenic cell differentiation. These findings led Ruch et al to state:
Experimental data demonstrate that dental histomorphogenesis and cytodifferentiation are controlled by an alternative flux of information circulating between ectomesodermal and epithelial cells. They are matrix-mediated signals. The basement membrane is a dynamic, asymmetric interface demonstrating compositional and conformational modulations. The spatial pattern and timing of these changes result from specific activities of adjacent cells.4
Based on numerous in vitro experiments, Ruch et al4 proposed that basement membrane modifications are causally related to successive steps of odontogenesis. The following are the essential points of this hypothesis:
1. Time- and space-specific information is encoded in the basement membrane constituents.
2. This information is read by cell membrane receptor molecules of adjacent cells.
3. Receptor-ligand interactions act on the cytoskeleton and/or cytoplasmic enzymes, which subsequently influence transcriptional and posttranscriptional events.
To date, fibronectin, fibronectin receptors, tenascin, and syndecan have been implicated as participants in matrix-mediated signaling during Odontogenesis.
The distribution of cell adhesion molecules and substrate adhesion molecules as potential control factors in tooth development has been a subject of increasing interest. Syndecan, a proteoglycan cell adhesion molecule located in the cell membrane, is expressed prior to tooth formation in the ectomesenchymal cells that underlie the dental epithelium.37 Tenascin, a large substrate adhesion molecule, is expressed in the ectomesenchyme during the downgrowth of the dental lamina and during the subsequent condensation of the dental papilla.38 It has been proposed that the binding of membrane-bound syndecan molecules to extracellular tenascin molecules is responsible for the condensation of the ectomesenchymal cells.37,39
An alternative explanation is that tenascin interferes with cell-to-fibronectin attachment, leading to decreased migration of the ectomesenchymal cells, causing them to aggregate in the form of the dental papilla. Adhesion of fibroblasts is weaker to fibronectin than to tenascin.40 It has also been shown that when cells express syndecan they have a reduced ability to invade a collagen gel. Thus, the appearance of syndecan on the cell surface of ectomesenchymal cells may have a direct, negative effect on their ability to migrate, thereby causing them to form aggregates, such as the dental papilla.
Tissue separation and recombination studies have demonstrated that the expression of syndecan and tenascin in tooth ectomesenchyme is induced during specific epithelial-mesenchymal interactions.5 In situ hybridization studies indicate that mRNA for tenascin is expressed in high amounts in cells of the inner enamel epithelium and the preodontoblasts. Redundant pathways regulating cell condensation are undoubtedly present, because tooth development has been shown to proceed normally in mice lacking tenascin expression.41
Role of growth factors
Advances in organ culture technique have made it possible to grow developing teeth in chemically defined culture media. Yamada and coinvestigators42 demonstrated that explants of developing teeth could undergo complete cell differentiation and matrix mineralization in a chemically defined medium. They concluded that autocrine and paracrine factors coordinate the sequence of cellular differentiation events during tooth development. This stimulated the search for diffusible growth and regulatory factors that might be involved in odontogenesis.
Using chemically defined culture media, Chai et al43 showed that tooth size and rate of development are regulated in part by transforming growth factor β2 (TGF-β2). When antisense oligonucleotides against TGF-β2 are added to tooth organ cultures, development is accelerated and the tooth buds grow larger than controls.43 Addition of exogenous TGF-β2 reverses the effect of antisense nucleotides, leading to normal growth.
The advent of powerful molecular biologic approaches marked the beginning