Runx2 Deletion in Dental-Epithelium Impair Ameloblast Differentiation and Enamel Formation

Objectives: Runx2 is an essential transcription factor for osteoblast differentiation and bone formation. However, molecular regulators of ameloblast differentiation and enamel secretion are less understood. The goal of this study is to uncover the role of Runx2 in ameloblast differentiation and enamel synthesis.
Methods: Runx2 gene was deleted specifically in ameloblasts using Keratin-14-Cre (K14-Cre) transgene. Molecular and histological techniques were used to evaluate ameloblast differentiation and enamel synthesis in newborn, 10-day, and 1-month-old littermates.
Results: K14-Cre activity specifically in the developing ameloblasts was confirmed by Td-reporter mice. The homozygous (Runx2^{F/F K14}) mice survived to adulthood and showed comparable size and positions of incisors and molars with distinct pre-dentin but no enamel matrix at birth. Comparable morphology of ameloblast at the pre-secretory and secretory-stages and the expression of marker-genes in Runx2^{F/F K14} littermates indicates that Runx2 is not essential for the initial stages of ameloblast differentiation. Analysis of 10-day-old WT-littermates revealed the presence of secretory and cuboidal mature ameloblast and the enamel matrix. Surprisingly, Runx2^{F/F K14} mice showed only elongated secretory ameloblast and a large amount of enamel-matrix in both molars and incisors. Comparable expression of Amelogenin, Enamelin, and Amelotin suggest that enlarged enamel-matrix in Runx2^{F/F K14} teeth is not due to increased synthesis. Interestingly, expression of mature ameloblast marker, Kallikrein-related-peptidase-4 (KLK4) was nearly absent in Runx2^{F/F K14} mice, indicating a failure in the degradation of enamel-matrix. The erupted molar and incisor of 1-month-old Runx2^{F/F K14} mice showed significant attrition of enamel at the cusp and incisal edge. The µCT of mandibular teeth revealed a 75% reduction in the enamel volume and density. A patchy enamel was noted only on the labial surface and molar groves of Runx2^{F/F K14} mice. Interestingly, KLK4 expression showed a significant reduction in the 1-month-old Runx2^{F/F K14} incisor.
Conclusions: Runx2 is essential for amelogenesis and the development of enamel.