Abstract
Odontogenesis relies on the reciprocal signaling interactions between dental epithelium and neural crest-derived mesenchyme, which is regulated by several signaling pathways. Subtle changes in the activity of these major signaling pathways can have dramatic effects on tooth development. An important regulator of such subtle changes is the fine tuning function of microRNAs (miRNAs). However, the underlying mechanism by which miRNAs regulate tooth development remains elusive. This study determined the expression of miRNAs during cytodifferentiation in the human tooth germ and studied miR-34a as a regulator of dental papilla cell differentiation. Using microarrays, miRNA expression profiles were established at selected times during development (early bell stage or late bell stage) of the human fetal tooth germ. We identified 29 differentially expressed miRNAs from early bell stage/late bell stage comparisons. Out of 6 miRNAs selected for validation by qPCR, all transcripts were confirmed to be differentially expressed. miR-34a was selected for further investigation because it has been previously reported to regulate organogenesis. miR-34a mimics and inhibitors were transfected into human fetal dental papilla cells, mRNA levels of predicted target genes were detected by quantitative real-time PCR, and levels of putative target proteins were examined by western blotting. ALP and DSPP expression were also tested by qPCR, western blotting, and immunofluorescence. Findings from these studies suggested that miR-34a may play important roles in dental papilla cell differentiation during human tooth development by targeting NOTCH and TGF-beta signaling.
Highlights
Odontogenesis involves three major processes: morphogenesis, histogenesis, and cytodifferentiation [1]
Identification of differentially expressed miRNAs The developing human tooth germ can be divided into 4 main stages: dental lamina, bud stage, cap stage and bell stage
From a total of 1887 human miRNAs assayed, we identified transcripts that were differentially expressed with adjusted p-values of less than 0.05 between early bell stage and late bell stage human tooth germ
Summary
Odontogenesis involves three major processes: morphogenesis, histogenesis, and cytodifferentiation [1]. Terminal differentiation, which is controlled by cell-matrix interactions involving several signaling pathways, starts from the bell stage. The signaling interactions between ectoderm-derived dental epithelium and neural crest-derived mesenchyme are regulated by several pathways, including TGF-beta, SHH, WNT, FGF, and NOTCH [2,3,4]. These growth factors interact in an intricate network regulated by spatial and temporal expression during odontogenesis [5,6,7,8]. The regulation of tooth development by major signaling pathways has been studied [15,16,17,18,19,20], but the fine tuning of this network via microRNAs (miRNAs) has not yet been fully elucidated
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