Figure legends
Figure 1. The histological sections of human deciduous molar germ from
the cap-to-early bell stage. At embryonic stages of 12 weeks (A, late
cap stage), 15 weeks (B, early bell stage), and 18 weeks (C, bell
stage), the tooth germ gradually enlarges and differentiates. Scale bar,
100 μm. DP, dental papilla; IDE, inner dental epithelium; ODE, outer
dental epithelium; DF, dental follicle; SR, stellate reticulum; SI,
stellate intermedium; PEK, primary enamel knot; PA, preameloblast.
Figure 2. The overall identified proteins in different stages in human
deciduous molar germ. (A) Venn diagram illustrating the overlapping
proteins among the number of proteins identified in different stages of
the human deciduous molar. (B) Inter-individual variation analysis of
different stages. (C) The summary of high-coefficient of variation (CV)
proteins and low-CV proteins in human tooth germ. (D) Results of PANTHER
protein class illustrating overlapping proteins during the cap-to-bell
transition in human embryonic tooth germ. (E) Venn diagram displaying
the tissue-specific proteins of molar proteome compared with proteins in
Human Protein Atlas (HPA) and Genotype-Tissue Expression project (GTEx)
databases. Functional enrichment results and the protein are listed. (F)
The temporal expression of proteins involved in the important
developmental pathways retrieved from the common knowledge databases
(Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, Reactome, and
GeneCards).
Figure 3. The expression analyses of all identified proteins. (A)
Clustering dendrogram of all proteins based on dissimilarity algorithm
and assignment modules. (B) Module-trait correlation heatmap between
clinical traits and modules. (C) Correlation scatter plot between
protein significance and module membership in MEpaleturquoise and
MEgreenyellow modules. (D) Functional enrichment analysis of proteins in
MEpaleturquoise and MEgreenyellow modules. (E) Hub proteins expression
heatmap. (F) UMAP plots showed distributions of dental cell
subpopulations of public database (top); The bar charts (below)
representing the cell ratios of different cell populations. (G) Average
gene expression heatmap plot of different dental cell populations (top);
Temporal gene expression in four types of dental cells (below).
Figure 4. The protein expression analyses of differentially expressed
proteins (DEPs). (A) Fuzzy c-means clustering identifying five temporal
expression patterns and the corresponding biological processes. TFs,
transcription factors. (B) Top 20 proteins with the highest degree
scores by Cytohubba algorithm. (C) Average expression heatmap plot of
the top 20 proteins with five temporal expression patterns (top);
Temporal gene expression of the top 20 proteins in different cell
populations (below).
Figure 5. The validation of four DEPs (AFM, GJA1, PAX9, and KRT15) by
IHC analysis. (A) The normalized mass-spectrometry intensity of DEPs
from cap to early bell stage. (B) The representative images and relative
quantitative value of IHC staining. Arrow indicated dental epithelial
cells. Magnification, 200×. DEPs, differentially expressed proteins;
AFM, Afamin; GJA1, Gap junction alpha-1; PAX9, Paired box 9; KRT15,
keratin 15.