Our aim was to investigate, in combination with histomorphology and immunohistochemistry, the development and disintegration of Hertwig's epithelial root sheath (HERS) and associated phenotypic alterations throughout molar root formation in mice. ICR mice,3 through 28 days-old, were used and serial sagittal sections were prepared from the lower first molars. The development of HERS began with the formation of a bilayered extension of the inner and outer dental epithelium from the cervical loop of the enamel organ. The epithelial double layer continued to grow in an apical direction outlining the shape of the future root of the tooth. Through FGF signals HERS cells mediated the differentiation of dental papilla cells into dentin-secreting odontoblasts. HERS proliferation rates did not match the rate of odontoblast proliferation or root elongation, resulting in the separation at the cervical region from the coronal enamel epithelium and the fenestration or disintegration of the epithelial sheath Structure attached to the root dentin. The disintegration of HERS allowed for mesenchymal cells of the dental follicle to come in contact with the outer dentin surface. HERS-derived epithelial cells proved to have diverse rates: migration away from the root surface and incorporation into the cementum. Some of the migrated epithelial cells remained intact as the rests of Malasses in the periodontium, whereas the rest of the migrated cells appeared to undergo epithelial-mesenchymal transformation. The present study using ssDNA antibodies further proved that only a few or some HERS cells underwent apoptosis or cell death during root formation. The results obtained support the theory that HERS acts as a barrier between dental papillae and dental follicle and a guide for root elongation accompanying harmonious root dentinogenesis and cementogenesis. In this sense, the disruption of HERS appears to be a key event during early root formation and eruption.