Organoids Developed to Secrete Tooth Enamel Proteins

University of Washington scientists have created organoids that secrete the proteins that form dental enamel, the substance that protects teeth from damage and decay. “This is a critical first step to our long-term goal to develop stem cell-based treatments to repair damaged teeth and regenerate those that are lost,” said Hai Zhang, co-author of the paper describing the research that was published in Developmental Cell.

Tooth enamel, which protects teeth from the mechanical stresses incurred by chewing and helps them resist decay, is made during tooth formation by specialized cells called amelobasts. When tooth formation is complete, these cells die off. Consequently, the body has no way to repair or regenerate damaged enamel, and teeth can become prone to fractures or subject to loss.

To create ameloblasts in the laboratory, the researchers first had to understand the genetic program that drives fetal stem cells to develop into these highly specialized enamel-producing cells. To do this they used single-cell combinatorial indexing RNA sequencing (sci-RNA-seq), which reveals which genes are active at different stages of a cell’s development.

By performing sci-RNA-seq on cells at different stages of human tooth development, the researchers were able to obtain a series of snapshots of gene activation at each stage. They then used a sophisticated computer program, called Monocle, to construct the likely trajectory of gene activities that occur as undifferentiated stem cells develop into fully differentiated ameloblast.

“The computer program predicts how you get from here to there, the roadmap, the blueprint needed to build ameloblasts,” said Ruohola-Baker, senior author on the paper.

With this trajectory mapped out, the researchers, after much trial and error, were able to coax undifferentiated human stem cells into becoming ameloblasts. They did this by exposing the stem cells to chemical signals that were known to activate different genes in a sequence that mimicked the path revealed by the sci-RNA-seq data. In some cases, they used known chemical signals.

While conducting this project, the scientists also identified for the first time another cell type, called a subodontoblast, which they believe is a progenitor of odontoblasts, a cell type crucial for tooth formation.

The researchers found that together these cell types could be induced to form small, three-dimensional, multicellular mini-organs, called organoids. These organized themselves into structures similar to those seen in developing human teeth and secreted three essential enamel proteins: ameloblastin, amelogenin, and enamelin. These proteins would then form a matrix. A mineralization process that is essential for forming enamel with the requisite hardness would follow.

Zhang said the research team now hopes to refine the process to make an enamel comparable in durability to that found in natural teeth and develop ways to use this enamel to restore damaged teeth. One approach would be to create enamel in the laboratory that could then be used to fill cavities and other defects.