Funding opportunities

Regeneration of enamel forming ameloblasts from human non-dental epithelial cells

Funding Type: 
Basic Biology IV
Grant Number: 
RB4-05930
Funds requested: 
$1 380 754
Funding Recommendations: 
Not recommended
Grant approved: 
No
Public Abstract: 
Tooth defect and loss greatly diminishes the quality of individual's life, and bioengineering of teeth is an important goal in regenerative medicine. Tooth regeneration requires both ameloblasts to form enamel, and dental mesenchyme to form dentin/pulp complex. Human ameloblasts are not available in the erupted teeth. We have observed that when human developing tooth mesenchyme is combined with epithelial cells, the tooth mesenchyme can instruct the epithelial cells to differentiate into ameloblasts. We propose to de-differentiate pulp cells from extracted human teeth toward an early developmental stage that is capable of instructing epithelial cells to form ameloblasts. In a parallel effort, we will use key gene transcriptional regulators that we have identified as important for ameloblast differentiation, to directly transdifferentiate accessible human non-dental epithelial cells into an ameloblast fate. Reprogramming accessible human cells toward tooth lineage allows us to further develop cell based strategies for enamel or even whole tooth regeneration. The knowledge we gain from the epithelial-mesenchymal cells interation will benefit the goals for regeneration of other epithelial appendages, such as skin and mammary gland, which use the similar mechanism regulating organogenesis. Finally, the reprogrammed dental pulp cells can be used to regenerate or repair pulp in teeth with pulpal infection or those requiring pulp removal such as in root canal therapy.
Statement of Benefit to California: 
A full and functional dentition and intact tooth structures are critical for the health and well being of all Californians. A survey conducted in 2004 by American Public Health Association shows the rate of tooth loss for population in California is 13.3%, indicating that tooth related diseases and trauma are of significant concern for Californians. The lack of readily available cell sources for ameloblast regeneration constrains the advance of tooth bioengineering. In this proposal, we will explore two strategies to bioengineer ameloblasts. First, we will reprogram pulp cells from either fully formed permanent or primary teeth, to an earlier developmental stage, which can then instruct differentiation of epithelial cells into ameloblasts. Second, we propose to directly reprogram epithelial cells to an ameloblast phenotype, to further enhance the potential for successful tooth regeneration. These studies will allow us to move our current material based dental therapies toward a more biological approach of regenerating tooth tissues. It is also possible that de-differentiated dental pulp cells could be used to regenerate missing or infected tooth pulp cells, and may also be used for regeneration of other neural crest derivatives that comprise the periodontal ligament, alveolar bone, Schwann cells and neurons, further benefiting the health and well being of Californians.
Review Summary: 
Teeth develop from two cellular sources, dental mesenchyme, which gives rise to dentin/pulp, and ameloblasts, an epithelial cell type that produces enamel. The immature dental mesenchyme further serves an inductive role during tooth development, directing immature epithelial cells toward the ameloblast fate. Since ameloblasts are not present in adult human teeth, the applicant proposes to generate such cells from non-dental epithelial cells, thereby enabling future tooth bioengineering efforts. In one approach, the applicant intends to generate immature dental mesenchyme cells with strong ameloblast-inducing ability by reprogramming pulp cells isolated from extracted human teeth. These will then be co-cultured with human non-dental epithelial cells to induce the latter to adopt ameloblast characteristics. In a second approach, the applicant will attempt to directly reprogram human non-dental epithelial cells into ameloblasts. Significance and Innovation - Reviewers were not convinced that there is a pressing need for bioengineered teeth. - If tooth bioengineering is to be achieved, identification of a human source of ameloblasts, as proposed here, is essential. - The applicant does not make a convincing case that the proposed approaches represent an appropriate way forward for dental practice. It is questionable whether a complex and most likely expensive approach based on co-cultures is feasible and will ever be viable as a treatment for tooth disease. - The basic approaches proposed for cell reprogramming are likely innovative within the dental field, but derive directly from work being done in many other cell lineages. Furthermore, others have generated enamel-secreting ameloblasts by recombining human keratinocytes with mouse embryonic dental mesenchyme; the significance of the current project is to achieve the same goal using exclusively cells of human origin. - The experimental system has the potential to be used for very interesting developmental biology studies, but this is not well reflected in the actual proposal. Feasibility and Experimental Design - The preliminary data are not sufficiently compelling to suggest feasibility of de-differentiating pulp cells, since the differences in transcription factor expression levels observed between immature and mature dental mesenchyme are too small to support the notion that their overexpression will promote reprogramming to the desired cell type. - Some of the preliminary data are supportive and clearly presented, but would have benefitted from providing the purity of isolated human dental cell populations and from evidence that a critical resource, a cell type specific promoter, functions properly in the desired cell populations. Principal Investigator (PI) and Research Team - The PI has a solid but not exceptional track record. - The PI has assembled a research team that has the appropriate expertise and seems capable of carrying out the proposed experiments. The collaborations will most likely contribute to the success of this project. Responsiveness to the RFA - The mechanistic work has already been completed as the transcriptional regulators to be tested as tools for cell fate change have already been identified. The proposed work is more akin to early translational studies. - Although the proposed work is somewhat responsive to RFA, it is unlikely to reveal fundamental new basic mechanisms.
Conflicts: 

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