Single-cell RNA sequencing breakthrough for tomorrow’s dentistry

New knowledge on the cellular makeup and growth of teeth can expedite developments in regenerative dentistry – a biological therapy for damaged teeth – as well as the treatment of tooth sensitivity. The study, which was conducted by researchers at Karolinska Institutet, is published in Nature Communications.

Teeth develop through a complex process in which soft tissue, with connective tissue, nerves and blood vessels, are bonded with three different types of hard tissue into a functional body part. As an explanatory model for this process, scientists often use the mouse incisor, which grows continuously and is renewed throughout the animal’s life.

Despite the fact that the mouse incisor has often been studied in a developmental context, many fundamental questions about the various tooth cells, stem cells and their differentiation and cellular dynamics remain to be answered.

Using a single-cell RNA sequencing method and genetic tracing, researchers at Karolinska Institutet, the Medical University of Vienna in Austria and Harvard University in the USA have now identified and characterised all cell populations in mouse teeth and in the young growing and adult human teeth.

 In-depth single-cell analysis of dental epithelium

a t-SNE dimensional reduction shows subpopulations of 268 single epithelial cells. 13 unbiased clusters (colors) reveal previously unrecognized stem, progenitor and mature epithelial subtypes. Inset: mitotic signature as defined by average expression of cell-cycle-related genes. b Identification of a previously unrecognized cellular subtypes of epithelial layer. RYR2+ cells in ameloblasts’ layer and THBD+ subpopulation of stratum intermedium organized into cuboidal layer found by immunohistochemistry. c Panel on the right shows localization of ACTA2-expressing cells inside the labial cervical loop (immunohistochemistry) and corresponding expression of Acta2 predicted from RNA-seq analysis (left panel). d Long-term (2 months) lineage tracing of a Acta2CreERT2/R26tdTomato dental epithelial stem cells shows the traced cells in both apical (near the cervical loop) and distal ameloblasts. Ameloblast character was proved both morphologically and by expression of CALB1 (immunohistochemistry). e Transcriptional program of ameloblasts differentiation. Four clusters corresponding to different stages of ameloblasts maturation (upper). Transcriptional states of ameloblasts progenitors were modeled as a single trajectory, which reveals sequence of cell state transitions and linked activity developmental gene modules (bottom). Heatmap: the cells (columns) are arranged according to estimated pseudotime, genes (rows) were clustered in nine modules. Smoothed gene expression profiles are shown. f Transient progenitor population found in labial cervical loop is demarcated by the expression of Egr1 and Fos. Panels in the bottom part shows the lineage tracing of FosCreERT2/R26ZsGreen1. Insets show the lineage traced cells in outer enamel epithelium. Of note, FosCreERT2/R26ZsGreen1 traced cells in epithelial and mesenchymal compartments are of distinct origins since compartments are spatially separated. (LaCL Labial Cervical Loop, SI Stratum Intermedium, Am. Ameloblasts). Scale bars: bde: 50 µm; c and insets of e: 10 µm.

“From stem cells to the completely differentiated adult cells we were able to decipher the differentiation pathways of odontoblasts, which give rise to dentine – the hard tissue closest to the pulp – and ameloblasts, which give rise to the enamel,” say the study’s last author Igor Adameyko at the Department of Physiology and Pharmacology, Karolinska Institutet, and co-author Kaj Fried at the Department of Neuroscience, Karolinska Institutet. “We also discovered new cell types and cell layers in teeth that can have a part to play in tooth sensitivity.”

Some of the finds can also explain certain complicated aspects of the immune system in teeth, and others shed new light on the formation of tooth enamel, the hardest tissue in our bodies.

“We hope and believe that our work can form the basis of new approaches to tomorrow’s dentistry. Specifically, it can expedite the fast expanding field of regenerative dentistry, a biological therapy for replacing damaged or lost tissue.”

The results have been made publicly accessible in the form of searchable interactive user-friendly atlases of mouse and human teeth. The researchers believe that they should prove a useful resource not only for dental biologists but also for researchers interested in development and regenerative biology in general.

SourceKarolinska Institutet

Availability – Processed data and interactive views of datasets can be accessed on the authors’ website: [http://pklab.med.harvard.edu/ruslan/dental.atlas.html].

Krivanek J, Soldatov RA, Kastriti ME, Chontorotzea T, Herdina AN, Petersen J, Szarowska B, Landova M, Matejova VK, Holla LI, Kuchler U, Zdrilic IV, Vijaykumar A, Balic A, Marangoni P, Klein OD, Neves VCM, Yianni V, Sharpe PT, Harkany T, Metscher BD, Bajénoff M, Mina M, Fried K, Kharchenko PV, Adameyko I. (2020) Dental cell type atlas reveals stem and differentiated cell types in mouse and human teeth. Nat Commun 11(1):4816. [article]

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