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Abstract
The main objective of this study was to map global gene expression in order to provide information about the populations of mRNA species participating in murine tooth development at 24 h intervals, starting at the 11th embryonic day (E11.5) up to the 7th post-natal day (P7). The levels of RNA species expressed during murine tooth development were mesured using a total of 58 deoxyoligonucleotide microarrays. Microarray data was validated using real-time RT-PCR. Differentially expressed genes (p < 0.05) were subjected to bioinformatic analysis to identify cellular activities significantly associated with these genes. Using ANOVA the microarray data yielded 4362 genes as being differentially expressed from the 11th embryonic day (E11.5) up to 7 days post-natal (P7), 1921 of these being genes without known functions. The remaining 2441 genes were subjected to further statistical analysis using a supervised procedure. Bioinformatic analysis results for each time-point studied suggests that the main molecular functions associated with genes expressed at the early pre-natal stages (E12.5–E18.5) were cell cycle progression, cell morphology, lipid metabolism, cellular growth, proliferation, senescence and apoptosis, whereas most genes expressed at post-natal and secretory stages (P0–P7) were significantly associated with regulation of cell migration, biosynthesis, differentiation, oxidative stress, polarization and cell death. Differentially expressed genes (DE) not described earlier during murine tooth development; Inositol 1, 4, 5-triphosphate receptor 3 (Itpr3), metallothionein 1(Mt1), cyclin-dependent kinase 4 (Cdk4), cathepsin D (Ctsd), keratin complex 2, basic, gene 6a (Krt2-6a), cofilin 1, non-muscle (Cfl1), cyclin 2 (Ccnd2), were verified by real-time RT-PCR.
Highlights
During murine tooth development substantial changes occur within a time-span of 24 h
GENES WITH EXPRESSION LEVEL CHANGING OVER TIME Microarray results showed that a total 4362 of non-constant genes are expressed during murine tooth development from E11.5 up to P7; 1921 genes with unknown function and very highly expressed at all time-points with net fluorescence intensities above 10000 and 2441 differentially expressed genes at all time- points with known Entrez Gene ID
The results suggest that time-courses assayed by real-time RT-PCR show a similar trend to expression data obtained using microarrays (Figure 2)
Summary
During murine tooth development substantial changes occur within a time-span of 24 h. From E11.5 up to P7, i.e., in the course of 16 days the tooth germ develops from one layered oral epithelium into a phenotypic molar tooth. The developing murine tooth germ is an excellent model for studying the time-course of gene expression in a rapidly, developing organ. Differentiation requires participation of a large number of genes, the expression of which is regulated in time and space. The numbers and types of genes involved, vary depending on the developmental stage. MiRNAs and lncRNAs are known to be involved during tooth development (Jevnaker and Osmundsen, 2008; Michon et al, 2010; Michon, 2011) providing an additional layer of complexity
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