Tooth Bioengineering and Regenerative Dentistry

Abstract

Over the past 100 y, tremendous progress has been made in the fields of dental tissue engineering and regenerative dental medicine, collectively known as translational dentistry. Translational dentistry has benefited from the more mature field of tissue engineering and regenerative medicine (TERM), established on the belief that biocompatible scaffolds, cells, and growth factors could be used to create functional, living replacement tissues and organs. TERM, created and pioneered by an interdisciplinary group of clinicians, biomedical engineers, and basic research scientists, works to create bioengineered replacement tissues that provide at least enough function for patients to survive until donor organs are available and, at best, fully functional replacement organs. Ultimately, the goal of both TERM and regenerative dentistry is to bring new and more effective therapies to the clinic to treat those in need. Very recently, the National Institutes of Health/National Institute of Dental and Craniofacial Research invested $24 million over a 3-y period to create dental oral and craniofacial translational resource centers to facilitate the development of more effective therapies to treat edentulism and other dental-related diseases over the next decade. This exciting era in regenerative dentistry, particularly for whole-tooth tissue engineering, builds on many key successes over the past 100 y that have contributed toward our current knowledge and understanding of signaling pathways directing natural tooth and dental tissue development—the foundation for current strategies to engineer functional, living replacement dental tissues and whole teeth. Here we use a historical perspective to present key findings and pivotal advances made in the field of translational dentistry over the past 100 y. We will first describe how this process has evolved over the past 100 y and then hypothesize on what to expect over the next century.

Introduction

A Brief History of Tooth Replacement Therapies

Dentistry is one of the oldest of medical professions, traceable back to Egyptian times in approximately 2600 BC (American Dental Education Association c2015–2019). Archaeologists have unearthed dental fillings in teeth dating back to ~8000 BC, and references to dental decay can be found in Sumerian texts from 5000 BC (Delta Dental of Michigan). Study of tooth decay has continued since the 1700s until today (ADEA c2015–2019). Although Hippocrates and Aristotle, perhaps the earliest “evidence-based” dental clinicians, wrote about decaying teeth in the 4th century BC, the first book entirely devoted to dentistry—The Little Medicinal Book for All Kinds of Diseases and Infirmities of the Teeth—was not published until 1530 (ADEA c2015–2019).

Dentistry as a profession became established in the early 1700s, when the French surgeon Pierre Fauchard published his book titled The Surgeon Dentist, a Treatise on Teeth, introducing key ideas such as the importance of dental hygiene, the proposed use of dental fillings and dental prostheses, and the fact that sugar contributed to tooth decay (Adolfo Patiño 1985). The first dental college, the Baltimore College of Dental Surgery, opened in 1840, nearly 20 y after the American Dental Association (ADA) was formed, and the first university-affiliated dental school was founded, the Harvard University Dental School in 1867 (ADEA c2015–2019).

Soldiers became the driving force of dentistry following World War II when it became evident that significant improvements needed to be made to dental and oral hygiene, as well as to methods used to repair teeth, to maintain a reliable, functional, and healthy army. This realization spearheaded the establishment of the National Institutes of Health (NIH) in 1931 and the National Institute of Dental Research (NIDR) in 1948, later renamed the National Institute for Dental and Craniofacial Research (NIDCR) in 1988 (Sheridan 1988). Since then, efforts to create new and improved methods for tooth replacement therapies have been pursued. Although the field of dental materials research for dental tissue and tooth repair has dramatically expanded over the past 100 y, this review will focus on synthetic, bioengineered, and combination dental therapies that have led to current advancements in whole-tooth tissue engineering.

The Evolution of Clinical Approaches to Treat Tooth Loss

Despite the ancient origins of dentistry, treatments for dental caries, periodontal disease, and edentulism have for the most part remained largely unchanged for many decades and include the standard tooth replacement therapies described below.

Dentures

Dentures have been used to treat tooth loss as far back as 700 BC, when Etruscans of northern Italy fabricated dentures out of human or animal teeth (Proskauer 1979). Dentures made of ivory were popular in the 1700s, including sophisticated versions consisting of a denture plate made of carved hippopotamus ivory that supported human, donkey, and horse teeth (Ring 2010). Although porcelain dentures were invented in the mid-1770s, they were found to be too brittle and prone to chip, and human teeth secured to animal ivory remained the preferred therapy (Jordan 2014). In 1802, improved porcelain dentures mounted via springs on gold plates or embedded in the hard rubber material, Vulconite, were used rather than ivory, later to be replaced by acrylic resin and other types of plastic (Ladha and Verma 2011). Removable partial dentures, which in contrast to fixed dentures that could only be placed and/or removed by a clinician, provided a dental prosthetic that could be removed and reinserted by the patients themselves, became popular in the 1950s (Schmidt 1952). Today, dentures have fallen into disfavor because they are uncomfortable, resulting in lack of compliance and leading to bone resorption, even worse fit, and more discomfort for the denture wearer.

Dental Implants

Very early versions of dental implants found in remains from the year 2000 BC consisted of carved bamboo pegs (López-Píriz et al. 2019). The body of an early Egyptian king originating from 1000 BC contained an upper jawbone into which a copper peg had been hammered, either during his lifetime or after his death (Smith 2019). A Celtic grave in France contained a body with an iron false tooth thought to date to 300 BC, also likely to have been placed postmortem. Long before current dental implants were designed, ancient skulls were found to contain rare gems of jade, as well as more common materials such as sea shells (Irish 2004).

The search continued for improved methods to fabricate dental implants that were both durable and functional and that would not be rejected by the host. Then, in 1952, an orthopedic surgeon made the accidental finding that titanium cylinders implanted in a rabbit femur created a very tight fusion, termed ankylosis (Branemark 1983). This discovery led to the creation of titanium dental implants, the first of which was placed in 1965 by the orthopedic surgeon, Branemark (Adell et al. 1981). Since that time, for the past half century, dental implant therapies have remained basically the same, with only subtle improvements such as new surface coatings and textures thought to improve osseointegration. According to the NIDCR, the use of dental implants increased 4-fold from 1983 to 1987 (Garcia 2009), and today for those who can afford it, titanium alloy dental implants are the most popular go-to therapy for tooth loss (Bidra et al. 2016). Approximately 3 million people in the United States currently have implants, and that number is growing by 500,000 each year (Bidra et al. 2016).

Titanium implants have become increasing popular in all specialty fields of dentistry, with up to 10% of all dentists in the United States currently placing implants. Implants are so popular that many clinicians choose not to perform endodontic treatments to prolong the life of a natural tooth but rather will “save time” by going straight to an implant, based on the assumption that root canal–treated teeth will eventually desiccate and crack, eventually requiring implant therapy.

Advantages of Living Replacement Teeth versus Synthetic Dental Implants

A detailed discussion of dental implants, including their perceived strengths and weaknesses, can be found in this issue. Therefore, here we will focus on efforts to create living replacement teeth, based on the anticipated advantages of vital, vascularized, and innervated bioengineered teeth, as compared to synthetic dental implants. Due to the potential complications and associated poor outcomes for synthetic dental implants, laypersons, dental researchers, clinicians, and biomedical engineers have been working to establish biological-based strategies to treat edentulism. These efforts have spearheaded efforts in translational dentistry and whole-tooth tissue engineering.

Here we will discuss the history of efforts to regenerate teeth that closely resemble natural tooth development, as well as the potential advantages of replacement tooth therapies that use bioengineered dental tissues that closely match the physical and functional properties of natural dental tissues (Fig. 1).

Figure 1.

Timeline: characterizing embryonic and adult dental stem cells for tooth regeneration. DPSC, dental pulp stem cell; DSC, dental stem cell; NCC, neural crest cell; PDL, periodontal ligament; SHED, stem cells from human exfoliated deciduous teeth.

Embryonic Tooth Bud–Based Strategies for Tooth Regeneration

Embryonic Tooth Formation

In the early embryo, teeth develop from 2 types of cells, embryonic ectoderm (epithelium) and mesenchyme derived from cranial neural crest cells. Pioneering experiments by Ed Kollar, Andrew Lumsden, and others established the basic principles of how these 2 cell types interact to kickstart the entire developmental program, leading to the formation of functional teeth (Mina and Kollar 1987; Lumsden 1988). Beginning at ~10 d in mouse and 6 wk in humans, the first tooth-inductive signals originate in the oral ectoderm and are received by the underlying mesenchyme. These first inductive signals are transient, and upon receipt by mesenchymal cells, their expression ceases through mechanisms that are poorly understood at the present time. The mesenchymal cells then respond by producing their own inductive signals, which are transmitted to and received by the dental epithelium. This reciprocal crosstalk continues throughout tooth development, resulting in a fully formed tooth. As such, these 2 cell types mutually induce each other to adopt odontogenic fates and participate in the process of tooth development.

Bioengineered Tooth-Based Developmental Signaling Cascades

Based purely on these early and relatively straightforward principles of dental tissue interactions, and without prior knowledge of the nature of the inductive signals themselves, studies to fabricate bioengineered teeth from embryonic tooth bud sources were conducted (Fig. 2). A landmark study published in the Journal of Dental Research in 2004 by Ohazama et al. showed that inductive oral ectoderm could be combined with mesenchymal cells obtained from an adult nondental source, such as bone marrow, to initiate tooth development (Ohazama et al. 2004). The bone marrow mesenchymal stem cells were able to receive and respond to tooth inductive signals and send the appropriate signals back to the dental epithelium. In a reciprocal series of experiments, this group was able to induce tooth formation in adult epithelial cells isolated from gingiva by combining them with embryonic tooth mesenchyme and implanting the resulting constructs in a renal capsule (Angelova Volponi et al. 2013). These studies confirmed that an entire tooth organ could be formed outside the embryo by applying the concepts of developmental biology. In both cases, adult cells were rendered odontogenic and fully participated in tooth development after receiving appropriate signals from an embryonic inductive source. The key to the success of these experiments was the use of very large numbers of embryonic inductive cells.

Figure 2.

Embryonic dental stem cells for tooth regeneration. Red arrows indicate tooth primordia. Figure courtesy of Dr. Ana Angelova Volponi (parts of figure created with BioRender.com).

Lesot and colleagues next established the minimum cell numbers required for tooth initiation (Hu et al. 2006). In mice, a range of 4.0 × 10⁴ to 4.0 × 10⁵ cells was obtained by combining cells harvested from multiple embryos, an approach that is obviously not possible in humans. Unfortunately, this study found that in vitro cultured embryonic cells lose their inductive capacity within 24 to 48 h (Zheng, Cai, et al. 2016). These results suggested that in vitro expansion of embryonic inductive dental cells using standard culture methods may not be possible and therefore would not be a feasible approach for human replacement tooth strategies. A fundamental challenge, therefore, is to discover ways to expand populations of embryonic tooth cells while maintaining their inductive capacity. As such, this conundrum has become a roadblock for the use of embryonic tooth bud cells for tooth regeneration strategies and the main reason why further efforts using this approach have slowed.

Several efforts to identify how and why in vitro cultured embryonic cells lose their inductive capacity focused on elucidating roles for cell-cell contact and signaling molecules. Cell-cell contact was thought to be important since inductive dental mesenchymal cells in the embryonic tooth primordium are tightly packed or condensed and as such experience significant cell-cell contact. In contrast, mesenchymal cell condensation is lost in monolayer in vitro tissue culture, resulting in much reduced cell-cell contacts. Methods to promote mesenchymal cell condensation and increase cell-cell contacts included growing cells in hanging drops, a 3-dimensional (3D) cell culture method that has proved successful for growing functional dermal papilla hair cells, although the large numbers of cells needed for tooth induction makes this method inappropriate for regenerative applications in humans (Higgins et al. 2013). To address cell signaling in tooth regeneration, gene expression screens were used to identify signaling factors whose expression was lost in 2-dimensional in vitro cell culture (Zheng, Jia, et al. 2016). A concern with this approach, however, is how best to distinguish between genes whose expression is altered due to changes in inductive odontogenic signaling from those associated with effects of in vitro monolayer culture and loss of cell contact. Similar screens were successfully performed on inductive hair cells using the hanging drop method (Kalabusheva et al. 2017), allowing for comparison of gene expression in primary inductive cells, in vitro cultured cells that have lost inductive signals, and cells in which inductive signals were restored in hanging drops (Kalabusheva et al. 2017). Using this approach, Yang et al. (2017) investigated the phenomenon of the “cell community” effect to determine if the presence of inductive cells could restore inductive capacity in in vitro cultured dental cells (Yang et al. 2017). Using genetically labeled cells and mixtures of inductive and noninductive mesenchymal cells, they showed that inductive capacity could be restored by what they called a “cell community effect,” providing the possible means to identify inductive signals that are lost and then restored. If successful, this approach could facilitate human bioengineered replacement tooth therapies.

Functional Bioengineered Tooth Development

A fundamental requirement for dental cell recombination approaches to tooth bioengineering is the ability for the bioengineered tooth primordia to develop into fully functional teeth, in the mouth. In fact, bioengineered tooth primordia created from embryonic tooth cells will readily form mineralized teeth when transplanted and grown under the renal capsule and when transplanted into the mouth (Ohazama et al. 2004). Similarly, bioengineered tooth primordia also formed mineralized teeth following surgical transplantation into the mouth (Nakao et al. 2007; Ikeda et al. 2009). Surprisingly, root formation, tooth eruption, and bone formation, all of which occur postnatally in natural tooth development, occurred in these transplants after a few weeks.

Postnatal Dental Cells for Bioengineered Whole-Tooth Tissue Engineering

Elegant studies of mouse embryonic tooth development, as well as the use of embryonic mouse tooth bud cells for bioengineered tooth formation, have contributed to a much greater understanding of the signaling cascades and in vitro culture conditions important for tooth development and regeneration. But embryonic tooth bud cells are not ideal for clinically relevant tooth replacement therapies, due to the drawbacks mentioned above and because human embryonic tooth bud cells are not readily available, if they are available at all. Therefore, considerable efforts have been made to define the regenerative capacity of adult dental cells for applications in bioengineered replacement tooth formation. Adult dental stem cells harvested from deciduous (baby or primary) or adult exfoliated teeth are ideal sources for applications in tooth regeneration because they are readily available, can be harvested and banked for future use, and are otherwise routinely discarded, making use of an otherwise wasted human autologous tissue source for regenerative dentistry (Fig. 3).

Figure 3.

Adult dental stem cells for tooth regeneration. (A) Dental stem cell sources. (B) Dental pulp of a harvested human tooth. DPSC differentiated to adipogenic (C–E), osteogenic (F–H) and neuronal (I–K) cell fates. PDL, periodontal ligament.

Postnatal Dental Stem Cells for Bioengineered Tooth Formation

Postnatal Dental Stem Cells

Publications proposing the use of adult, postnatal dental stem cells for whole-tooth tissue engineering applications first appeared in the early 2000s. Exciting work by a team at the NIH/NIDCR was the first to characterize a clonogenic and rapidly proliferative population of cells harvested from adult human dental pulp, called dental pulp stem cells (DPSCs) (Gronthos et al. 2000). These cells were distinct from bone marrow stromal cells (BMSCs) in that they only sporadically formed densely calcified nodules in in vitro culture, did not differentiate into adipocytes, and could form a dentin/pulp-like complex when implanted in vivo (Gronthos et al. 2000). This team went on to characterize these cells at the molecular level, demonstrating shared and differential gene expression patterns of BMSCs and DPSCs (Shi et al. 2001). This group further characterized the self-renewal capability of clonogenic DPSCs, demonstrating variability of clones with respect to their ability to generate ectopic dentin when implanted in vivo (Gronthos et al. 2002), and also determined that both DPSC and BMSCs resided in the perivascular niche of dental pulp and bone marrow, respectively (Shi and Gronthos 2003).

Stem Cells from Human Exfoliated Deciduous Teeth

One of the first studies using nonembryonic dental cell sources for tooth regeneration used dental pulp stem cells harvested from human deciduous (baby) teeth, called stem cells from human exfoliated deciduous teeth, or SHED (Miura et al. 2003). SHED are highly proliferative, capable of differentiating into a variety of cell types, including neural cells and odontoblast-like cells. When implanted in vivo, SHED formed bone and dentin, as well as survived and expressed neuronal cell markers when implanted in the mouse brain (Miura et al. 2003). This exciting report identified baby teeth as a potential and unexpected cell source for bioengineered tooth replacement therapies.

Around the same time, another group was taking a different approach, using adult dental mesenchymal (DM) and dental epithelial (DE) cells harvested from unerupted porcine tooth buds from discarded 6-mo-old pig jaws obtained from local abattoirs. Based on the exciting progress being made in the fields of regenerative medicine and tissue engineering, this team decided to determine whether tissue engineering strategies being used to regenerate heart, bone, and other tissues could also be used to engineer teeth. Porcine DE and DM cells were expanded in culture and seeded onto available biodegradable scaffolds such as polyglycolic acid (PGA)/poly(lactic-co-glycolic) acid (PLGA) and either characterized in vitro or implanted in vivo (Young et al. 2002). Excitingly, results from these studies demonstrated that this approach could be used to generate small, anatomically correct tooth crowns, consisting of dental pulp, dentin, and enamel (Young et al. 2002).

The results of this pioneering group were groundbreaking and seminal, in that they demonstrated that adult, mammalian tooth bud–derived dental cell sources and tissue engineering strategies could be used to regenerate teeth and, importantly, that in vitro cultured adult DE and DM cells could be expanded in in vitro culture for up to at least 8 passages while still retaining their capacity to form dental tissues, including dental pulp, dentin, enamel, and whole tooth crowns. Based on their findings, this team went on to show that other mammalian dental cell sources, including cells derived from porcine, rat, and human teeth, all could be used to create bioengineered tooth structures (Duailibi et al. 2004; Young, Abukawa, et al. 2005; Young, Kim, et al. 2005; Duailibi et al. 2011).

Induced Pluripotent Stem Cells for Dental Tissue Regeneration

The potential to create any type of autologous cell from cells easily and painlessly obtained from any individual was provided by Dr. Shinya Yamanaka’s laboratory in 2006, when they published methods used to generate induced pluripotent stem cells (iPSCs) (Takahashi and Yamanaka 2006; Yamanaka and Takahashi 2006). These publications and those that have followed have generated significant excitement in the fields of regenerative medicine and dentistry, for which autologous cell sources are quite limited to begin with, and appropriate autologous cell sources may be impossible to obtain because harvesting would create additional damage to an individual. With respect to the use of iPSCs in dental tissue and whole-tooth tissue engineering, a number of groups are working to improve this technology to bring iPSC therapies to clinical relevance. Efforts are ongoing to regenerate dental pulp (Goldberg 2011), periodontal ligaments (PDLs) (Cho et al. 2019), odontoblasts (Xie et al. 2018), and ameloblasts (Cai et al. 2013; Abdullah et al. 2019). Although promising, dental therapies using iPSCs are still relegated to the distant future, to allow for thorough validation of these approaches.

Scaffold Materials for Whole-Tooth Tissue Engineering

To further efforts for regenerative medicine and dentistry, many investigators have been searching for optimal biodegradable scaffold materials and designs that will support the formation of the desired organ and/or tissues of interest, and of particular and specified size and shape. With respect to tooth regeneration, a wide range of scaffolds has been used, including hydrogel scaffolds such as gelatin methacrylate (GelMA) hydrogels (Smith et al. 2016; Khayat et al. 2017; Smith et al. 2017; Smith et al. 2018; Smith and Yelick 2019), scaffold-free dental cell sheets grown on thermosensitive tissue culture plates (Monteiro et al. 2016; Monteiro and Yelick 2017), silk-based hydrogel and porous scaffolds (Xu et al. 2008; Zhang et al. 2011), and natural decellularized tooth bud scaffolds (Traphagen and Yelick 2009; Traphagen et al. 2012; Zhang et al. 2017), to name a few.

To date, using adult dental cell sources and decellularized natural tooth bud extracellular matrix (ECM) scaffolds, close to full-sized teeth have been bioengineered and grown in the jaws of mini pig hosts, bringing us closer than ever before to our goal (Fig. 4) (Zhang et al. 2017). As more sophisticated methods are developed to create scaffolds that more closely resemble natural tissue scaffolds, the field is sure to progress toward the goal of tissue regeneration, including functional tooth regeneration.

Figure 4.

Adult dental stem cell–derived bioengineered tooth. Two different bioengineered teeth (top, bottom). (A) Micro–computed tomography (CT) image of well-developed 6-mo implanted nTB (box). (B) H&E-stained coronal histological section of nTB crown (black box). (C) High-magnification image of the boxed area in B. (D) High-magnification image of the boxed area in C. (E) Micro-CT image of a 6-mo recell-dTB implant. (F) Tooth root structure (white box). (G, H) High-magnification images of dentin and periodontal ligament tissues. C, canine; d, dentin; dTB, decellularized tooth bud; e, enamel; H&E, hematoxylin and eosin; I, incisor; nTB, natural tooth bud; PM, premolar; POL, polarized light image. Scale bar: A, E = 10 mm; B, F = 2 mm; C, D, G, H = 20 microns. (Zhang et al. 2017)

Current Challenges for Whole-Tooth Tissue Engineering

Despite tremendous progress in the field of translational dentistry and medicine, certain obstacles remain, as itemized and discussed below.

How to Create Functional, Full-Sized Bioengineered Teeth of Predetermined Size and Shape?

The ability to create bioengineered teeth that adopt the size and shape of the original tooth is a highly coveted and necessary goal for the field. Size and shape are extremely important, especially for teeth, due to their close occlusion with opposing teeth, where less than millimeter differences can result in jaw muscle and nerve pain, temporomandibular joint dysfunction, and reduced aesthetics. The ability to faithfully fabricate bioengineered tooth replacement models using computer-aided design/computer-aided manufacturing imaging and 3D printing technologies is certainly helping the field move closer to attaining this goal. The size and shape challenge is also closely linked to that of proper vascularization and innervation, in that bioengineered full-sized teeth will need proper vascularization and innervation to grow and survive.

Another important consideration is how best to regrow teeth in a timely and highly functional manner and in a way that is integrated with host-supporting tissues such as alveolar jaw bone and PDL. New and promising strategies for tissue engineering and regenerative medicine include the coordinated simultaneous engineering of tissues—such as muscle, bone nerve, and vasculature—all at once, to facilitate the successful creation of functional bioengineered tissues. Such approaches are currently being applied to the creation of jaw bone and tooth constructs, which can grow together along with supporting vasculature and nervous system, to create functional teeth.

Vascularization and Innervation

Natural tissues and organs are highly vascularized to provide tissues with proper nutrients and to remove unwanted cellular waste products. In addition, and of particular importance with respect to teeth, the presence of a nervous system to sense and modulate pain, such as that experienced by clenching, is very important for the health and longevity of these tissues. Since synthetic dental implants are not innervated, it is not uncommon for implants and/or occluding teeth to be fractured due to excessive force while chewing. The ability to regenerate teeth that are highly vascularized and innervated would be a significant improvement over currently available tooth replacement therapies and is therefore a highly coveted goal for the field. Progress has been made toward both understanding and manipulating vascularization and innervation in bioengineered teeth. Addition of exogenous agents to promote innervation, such as semaphoring 3A receptor inhibitors, promotes dental pulp innervation, immunomodulation by agents such as cyclosporin A, and addition of cells in the form of bone marrow stromal cells (Kökten et al. 2014; Strub et al. 2018). Vascularization occurs readily in bioengineered teeth implanted into ectopic sites and in the mouth (Young et al. 2002; Duailibi et al. 2004; Nait Lechguer et al. 2008; Xu et al. 2008; Abukawa et al. 2009; Zhang et al. 2011; Monteiro et al. 2016; Khayat et al. 2017; Smith et al. 2017; Zhang et al. 2017; Sharpe lab, unpublished observations).

Fighting Infection and Maintaining Oral Health

Another important consideration for both natural and bioengineered replacement teeth is how best to maintain a healthy oral environment, with appropriate and balanced microbiota and natural salivary defenses (Lynge Pedersen and Belstrøm 2019). This continues to be a challenge for dental clinicians as well as for the medical field due to the close association of oral health and systemic health. In particular, since tooth loss is commonly caused by inflammation and disease caused by caries and periodontal disease, reducing unwanted bacterial load in the oral cavity and maintaining a healthy oral environment are key to natural, synthetic implant, and bioengineered tooth survival. Improved knowledge of healthy versus unhealthy oral microbiota, as well as of new methods to create and maintain a healthy oral environment, will certainly facilitate greater longevity of natural, synthetic, and bioengineered teeth.

Complex Dental Tissue Regeneration

The ability to combine synthetic and bioengineered dental tissues is also a promising approach for the field. For example, the ability to augment titanium implants with bioengineered PDL tissues that can elaborate cementum on and attach to the implant surface, as well as synthesize bone at the PDL-bone interface, would improve current implant technology, making dental implants more similar to natural teeth. Such “liga-plants” and other combinations of synthetic materials and living tissue constructs are likely to be a way of the future, blending the advantages of both into a more functional tooth replacement therapy. Finally, cell-free approaches for tissue regeneration are also being explored to avoid the need for autologous cell sources and the anticipated immune rejection response from using allogeneic cell sources. Secreted exosomes and microvesicles harvested from in vitro cultured DE and DM cell tissue culture media could potentially be used to “functionalize” hydrogel and other types of scaffolds, to direct dental cell differentiation into hard and soft dental tissue without the need for seeded cells (Huleihel et al. 2016). In addition, matrix-bound vesicles (MBVs) present on decellularized tooth bud scaffolds may also prove to be instructive for applications in regenerative dentistry (van der Merwe et al. 2019).

Conclusions

The Future: New and Promising Opportunities for Dental Tissue and Whole-Tooth Tissue Engineering—Where To Next?

We anticipate that continued advances in the fields of tissue engineering and regenerative medicine and dentistry will facilitate the development of improved dental repair therapies, including whole-tooth tissue engineering. At the present time, the overall concept of tooth bioengineering has been proven in principle. Combinations of adult and embryonic cells from mice and humans have been shown to form tooth primordia in vitro. And surgical transplantation of these constructs into the mouth was shown to provide a suitable environment for their development into fully functional, erupted teeth. With the longstanding roadblock of how to maintain inductive capacity in in vitro cultured embryonic dental cells perhaps close to being solved, as well as the successful manipulation of adult cell sources to form bioengineered dental tissues, clinical therapies to create human bioengineered teeth may be a realistic prospect in the not so distant future (Fig. 5).

Figure 5.

The future of translational dentistry: whole-tooth tissue engineering (Monteiro and Yelick 2017).

There remain, however, a number of issues that must first be resolved. One is the significant concern of how to control bioengineered tooth crown size and shape. How can the precise size and shape of a tooth crown be directed by bioengineered tooth primordia? A second consideration is that full human enamel mineralization takes approximately 1 y, while freshly erupted bioengineered teeth are likely to be substantially undermineralized, relatively. In truth, however, such issues may be less important if one focuses instead, for the time being at least, on perfecting methods to bioengineer functional tooth roots. The challenge with current replacement tooth therapies, whether synthetic or bioengineered, is how best to secure the replacement in the jaw. Currently used metallic dental implants are essentially artificial tooth roots. If bioengineering approaches could be used to create fully functional tooth roots, containing dentin and pulp and supporting cementum, PDL, and alveolar bone tissues, the size, shape, and mineral quality of the crown may be less important, as long as the bioengineered tooth root can support an accurately 3D printed tooth crown.

Although very optimistic about the future for whole-tooth bioengineering, we are also cognizant of the significant obstacles that must first be overcome, which will require considerable financial investment, time, and a certain amount of luck. It is also important to consider the likely cost of a human bioengineered tooth therapy. If a bioengineered tooth costs $10,000, for example, will it be a commercially viable product that can be supported by dental insurance? This is an important consideration, based on the fact that the field of regenerative medicine is littered with products that have failed due to prohibitive costs. Some regenerative medicine and dentistry approaches, including those using iPSCs, would be highly advantageous but raise the concern of whether iPSC-based therapies will be commercially viable in the high-volume, low-cost world of dental treatments. Although we are certain that whole-tooth bioengineering will eventually be possible, it will need to be cost competitive to serve as a mainstream clinical therapy. As such, ongoing and future research efforts for bioengineered tooth tissue engineering therapies should include cost considerations.

The Current Market and Anticipated Need

The current market for tooth replacement therapies is bright. Various sources quote that the global dental implant market size was valued at US$3.77 billion in 2016 and growing at a compound annual growth rate of 7.7% over the forecast period 2014–2016 (Grand View Research 2018). The World Health Organization (WHO) reported that injuries sustained by approximately 10 million people per year require tooth replacement therapies (Day et al. 2019), and the American Academy for Implant Dentistry stated that over 15 million people per year undergo bridge and crown replacement therapies for missing teeth (Bassir et al. 2019). The American Dental Association reported that 5 million implants are placed per year in the United States (Block 2018), and this number is anticipated to continue to increase. The Market Report from iData Research states that the entire US dental market is currently worth over US$17 billion, with a whopping $10,300 million in dental prosthetics, $1,400 million in dental implants, and another $1,000 million in dentures, bridges, and dental bone graft substitutes (Listl et al. 2015). These figures are partially driven by the increasing aged population in the United States and world, with the population over 65 y of age in the United States anticipated to reach 20% by 2050. All of these forecasts emphasize the need for better and more effective therapies for bioengineered dental tissues and whole teeth.

Author Contributions

P.C. Yelick, contributed to conception, design, and data acquisition, drafted and critically revised the manuscript; P.T. Sharpe, contributed to conception and data interpretation, drafted and critically revised the manuscript. Both authors gave final approval and agree to be accountable for all aspects of the work.