Gene expression analysis in cells of the dentine–pulp complex in healthy and carious teeth

Knowledge of the molecular events that occur in carious disease has so far been constrained due to difficulties in obtaining sufficient quantities of the dental tissues and cells involved. Our histological findings indicate that a pulp–odontoblast cellular complex can be obtained from carious and healthy human teeth when exposed to low-temperatures prior to pulpal extirpation and from rodent teeth processed at room-temperature. In contrast, pulpal tissue extracted from room-temperature processed human teeth and low-temperature processed rodent teeth resulted in the odontoblast layer remaining attached to the pulp chamber. Semi-quantitative RT-PCR (sq-RT-PCR) analysis confirmed that markers previously shown to be preferentially expressed in odontoblasts, namely dentin sialophosphoprotein (DSPP) and Nestin, amplified more readily from the extracted pulp–odontoblast complex, as compared to pulpal tissue alone, in both human and rodent samples. Subsequent gene expression analysis of collagen-1α and collagen-3α indicated levels were significantly higher in carious pulpal tissue. In addition, analysis characterising the expression of members of the transforming growth factor and bone morphogenic protein families and their receptors indicated in general, that these genes were expressed by healthy odontoblasts and up-regulated in both pulpal cells and odontoblasts in response to carious injury. Use of this temperature-sensitive dental tissue preparation procedure allows detection of differential gene expression in odontoblasts and other pulpal cells in healthy and carious tissue.