science news logo

science service logo


Something to Chew On
Hard facts about tooth enamel

Alexandra Goho

If you're not sitting in the dentist's office reading this article, chances are that your tooth enamel is doing its job, at least for now. This protective ceramic-like veneer, thick as a dime, is the strongest material in the human body. It's your teeth's first line of defense against corrosive bacteria and the constant pounding and stresses that come with chewing.

photo

SHINING ARMOR. Enamel, the protective coating on every tooth, is 95 percent mineral, and it's the strongest material in the human body. Scientists are working to emulate this natural armor to create better and longer-lasting materials for tooth restoration.

PhotoDisc

Strong as it is, enamel is also brittle. What keeps it from shattering is an underlying protein-rich layer of dentin, a softer, more compliant substance that acts like a shock absorber. "If you had enamel sitting on a hard surface, every time you'd bite down on it, it would crack," says Arthur Veis, a molecular biologist at Northwestern University in Evanston, Ill.

Enamel is one of the trickiest dental tissues to mimic, but it's a necessary component for creating synthetic stand-ins or repair kits for teeth. Over the years, a variety of ceramics and metals have been used as fillings, crowns, and other dental implants, but they lack the strength of enamel and often crack, dislodge, or wear down. Dental materials gone bad are a common clinical problem. According to the American Dental Association based in Chicago, dentists spend more than 50 percent of their time restoring damaged or decaying teeth and about two-thirds of these procedures are for replacing dental materials that have failed.

Investigators have made several attempts to grow enamel from the cells that make it within the gums, but "nobody has really demonstrated true enamel," says Pamela Robey of the National Institute of Dental and Craniofacial Research in Bethesda, Md.

Researchers have isolated several of the other cell types involved in tooth development. They have used these cells to regenerate dentin; cementum, which is the hard tissue on root surfaces; and the periodontal ligament that attaches each tooth to the jaw. "The one thing that's missing now is enamel," says Robey.

With several recent discoveries, researchers have gotten the best glimpse yet of enamel in formation. Thanks to advances in materials science as well, they're closer than ever to making synthetic enamel that can compete with nature's version. With that, they hope to reach an even more audacious goal: growing an entire tooth from scratch.

Swift assembly

The primary ingredient of both enamel and dentin is the calcium phosphate mineral called hydroxyapatite. Yet these two dental tissues have dramatically different properties. That's because about 40 percent of dentin consists of protein, whereas enamel is almost devoid of organic material. Like a dinner plate, tooth enamel is almost pure ceramic.

That property has long puzzled researchers. Nearly all biomineralized structures found in nature, from mollusk shells to bone, rely on proteins for their assembly. The proteins serve as scaffolds that guide crystal growth, often resulting in elaborate shapes and structures (SN: 7/17/04, p. 42: Available to subscribers at http://www.sciencenews.org/articles/20040717/bob9.asp). In the case of dentin, long and narrow crystals of hydroxyapatite grow along rod-shaped molecules of collagen, a particularly abundant structural protein in the body. The collagen becomes an integral part of dentin and counters hydroxyapatite's inherent brittleness.

For years, researchers have had good reason to suspect that a protein called amelogenin contributes to the growth of enamel in gums. Studies showed that defective amelogenin proteins lead to the disease amelogenesis imperfecta. In that disorder, tooth enamel doesn't form properly, leaving teeth discolored and vulnerable to damage. What's more, says Janet Moradian-Oldak of the School of Dentistry at the University of Southern California in Los Angeles, "a single mutation in the sequence of the gene that codes for amelogenin can completely destroy enamel structure."

photo

BANDS THAT BOND. During tooth development, a small protein called amelogenin spontaneously assembles into a microribbon structure (left). These protein ribbons serve as templates for the growth of hydroxyapatite crystals (right), the main ingredient in tooth enamel.

Science

At first glance, amelogenin looks all wrong for the job of crystal growth. It's about one-tenth the size of collagen and doesn't have the rodlike shape of the hydroxyapatite crystals in enamel. Researchers such as Moradian-Oldak have tried to study amelogenin in animals, but the protein disappears almost as soon as the cells make it.

About a decade ago, she and her colleagues came upon a clue to how amelogenin might work in the formation of enamel. While observing the protein under a microscope, the researchers watched it form spontaneously into tiny spheres about 20 nanometers in diameter. Each sphere consisted of a cluster of some two dozen individual protein molecules. Intriguing as the finding was, it seemed unlikely that these spheres could have much to do with enamel's long, narrow crystals.

Reporting in the March 4 Science, Moradian-Oldak and Guiseppe Falini at the University of Bologna in Italy appear to have finally uncovered amelogenin's mineral-forming properties.

First, when the researchers added the protein to a solution that closely matched the physiological conditions in gums, the proteins spontaneously assembled into nanospheres like those Moradian-Oldak had seen years ago. That's not all that happened.

The nanospheres joined to form chains. Then, the chains aligned themselves to form microribbons several hundred microns in length and about 30 microns wide. Using both a transmission electron microscope and an atomic-force microscope, the researchers mapped out the fine details of this protein-assembly process.

Since the overall shape of the ribbons matched that of enamel's constituent crystals, Moradian-Oldak could see the pieces of the puzzle coming together. So when she and her colleagues immersed the amelogenin microribbons in a solution of calcium phosphate, they were not entirely surprised to see well-ordered apatite crystals form along the length of the ribbons.

In addition to demonstrating how these highly organized amelogenin structures serve as scaffolds for enamel formation, the findings suggest that the same protein structures might initiate, or nucleate, the crystallization process, the researchers say.

Each amelogenin protein has a negatively charged tail that is attracted to water. So, when the proteins form nanospheres, the tails orient themselves along the surface of the spheres. Positively charged calcium ions from a calcium phosphate solution might then attach to the negatively charged protein tails, thereby initiating crystal growth simultaneously at multiple points along the ribbon's surface.

"People have speculated for years that these structures form," says Veis. Now, Moradian-Oldak and her crew have shown for the first time not only that amelogenin assumes these shapes but also that these structures guide crystal growth, Veis says.

Crack attack

Anyone who takes a close look at his or her teeth in the mirror will likely notice several vertical cracks in the enamel. These are not flaws needing a dentist's attention. Rather, they are physical records of the enamel doing its job. When a crack propagates through the enamel from the surface toward the interior of the tooth, it stops right at the junction between the enamel and the dentin. Without this built-in stopping mechanism, teeth would fracture in no time.

photo

TRACKING CRACKS. To study how cracks move through teeth, scientists created a series of microscopic dents in the enamel of an extracted human molar (top). These dents initiated artificial cracks in the tooth's hard coating, and the cracks migrated toward the junction between the enamel and the softer underlayer of dentin (scalloped line in top inset). Once the main crack passed the junction (bottom), smaller cracks created regions of unbroken material called uncracked ligament bridges (arrows) that brought the main crack to a halt.

Ritchie

Just how the dental-enamel junction (DEJ), which is less than a micron wide, serves as a stop sign for cracks has remained unclear. The junction is made of a material that keeps the enamel from peeling off the dentin. "Normally, the interface between two dissimilar materials is the weakest link in a structure," says Grayson Marshall of the Department of Preventive and Restorative Medicine at the University of California, San Francisco.

Originally, researchers assumed that the DEJ was tougher than dentin and, therefore, could absorb enough of the stress of a propagating crack to keep it from penetrating the dentin. However, a collaboration of dental researchers and materials scientists has found a different crack-stopping mechanism at work.

In the March Nature Materials, Grayson, Robert Ritchie of the University of California, Berkeley, and their colleagues describe artificial cracks that they created in more than a dozen extracted human molars. The researchers used an indenter—a materials-testing instrument that resembles a miniature plunger with a fine, diamond tip—to create microscopic pits on the surface of each molar. The pits, in turn, initiated cracks in the enamel. The researchers then used a scanning electron microscope to determine how the cracks move through the different layers of dental tissue.

Although the DEJ appears under a light microscope as a thin, straight line, higher magnifications reveal a scalloped structure that increases the contact surface between dentin and enamel, explains Grayson. When a crack reaches the DEJ from the enamel side of the tooth, the scallops can deflect the trajectory of the propagating crack. This decreases the likelihood that the crack's tip will hit the junction head-on and with full force.

Even at the dentin, "the crack doesn't have smooth sailing from then on," says Grayson. "Another mechanism kicks in."

As the propagating crack begins to pry open the dentin structure, bridges made of so-called uncracked ligaments form. By the time the crack has traveled less than a cell's thickness into the dentin, these bridges bring the crack to a stop.

The bridges form as a consequence of small cracks that develop ahead of that main crack's tip. Because the daughter cracks don't always line up perfectly with the mother crack, they create regions of unbroken tissue, or bridges, between themselves and the primary crack tip. The researchers observed several of these smaller cracks, which dissipate some of the energy driving the main crack.

Ritchie argues that empty tubules, left behind by dentin-forming cells after their job in tooth development is done, serve as seeds for the formation of these daughter cracks. "The presence of these tubules is critical," says Ritchie.

And since there are no such tubules in enamel, he adds, this crack-thwarting mechanism occurs only in dentin.

Ritchie and his colleagues have found that as teeth age, the tubules fill up with hydroxyapatite. "When they're plugged up, they can't [initiate] these microcracks quite the same way as they did before," he says. As a result, fewer uncracked ligament bridges form, and the aging teeth become more brittle.

Nibbling

The revelations about how cracks move through teeth should spur the development of next-generation materials for dental restoration, such as crowns, caps, and cavity fillings, Grayson says. A clearer picture of how the dental-enamel junction connects two dissimilar materials could also help researchers design longer-lasting orthopedic implants, such as hip replacements, that stick better to patients' bones. "The DEJ embodies a whole bunch of secrets that we can mimic," says Grayson.

Scientists in Japan are already showing what might be possible. Knowing that enamel is almost entirely mineral, the researchers aimed to create a similar dental material. Past attempts at all-mineral materials have fallen short of the real thing.

Kazue Yamagishi of the FAP Dental Institute in Tokyo and her colleagues used a super-saturated solution of calcium and phosphate ions to make synthetic enamel that's 100 percent hydroxyapatite. The material starts off as a paste and hardens within 15 minutes of application into a tooth cavity.

Like current fillings do, the synthetic enamel seals the cavity and prevents acid-secreting bacteria from further eroding the tooth. However, unlike conventional treatments, the material doesn't require drilling of the damaged area of the enamel. Dentists often have to remove some of the healthy tooth because conventional fillings such as polymer resins and metal alloys, don't stick properly to small cavities or damaged surfaces.

In the Feb. 24 Nature, the researchers describe experiments on an extracted human molar bearing a new cavity. Shortly after the scientists applied the paste, a minute amount of the molar's enamel dissolved, but it was quickly replaced by synthetic enamel, which filled the cavity and integrated with the natural material.

When viewed under a transmission-electron microscope, the microstructure of the filling was virtually indistinguishable from that of real enamel, and there was no obvious gap between the synthetic material and the natural enamel surrounding the cavity, says Yamagishi. That's important, she says, because existing filling materials have crystal structures different from that of enamel and therefore rarely adhere seamlessly to the natural tissue.

Yamagishi says that her group is gearing up to begin clinical trials. If all goes well, the dental paste could find its way into the dentist's office in the next 2 to 3 years, she says. So far, the material works only for small, early-stage cavities, not the deeper cavities of later-stage tooth decay.

Even so, such advances might ultimately help researchers build whole teeth from scratch to replace diseased or damaged teeth. To that end, the National Institute of Dental and Craniofacial Research plans to establish a center within the next 2 years, where a team of biologists, engineers, materials scientists, and clinicians will bring their expertise to bear on the problem. The replacement teeth that they envision would last longer than the dental implants available today.

The institute's strategy most likely will combine materials science-based approaches for making synthetic enamel and cell-based methods for growing other dental tissues. With a biomaterial that mimics the properties of natural enamel, researchers could bypass the need for enamel-producing cells. For instance, scientists might create a crown out of synthetic enamel and use it as a mold, says Robey. Inside the mold, researchers would then place all the necessary cells for regrowing the rest of the tooth, she says.

That 's the vision. It may take more than a decade before researchers realize such a goal. However, if successful, it would represent a major feat in bioengineering—not to mention a major boon for patients, who won't have to run back to the dentist so often for repairs on false teeth.

Dental science has certainly come a long way since the days of George Washington, whose sets of false teeth were fashioned not from wood, as legend has it, but from gold, hippo ivory, and horse teeth.

********

References:

Du, C., G. Falini … and J. Moradian-Oldak. 2005. Supramolecular assembly of amelogenin nanospheres into birefringent microribbons. Science 307(March 4):1450–1454. Abstract available at http://dx.doi.org/10.1126/science.1105675.

Imbeni, V. … G.W. Marshall … and R.O. Ritchie. 2005. The dentin-enamel junction and the fracture of human teeth. Nature Materials 4(March):229–232. Abstract available at http://dx.doi.org/10.1038/nmat1323.

Kinney, J.H. … and R.O. Ritchie. 2005. Age-related transparent root dentin: Mineral concentration, crystallite size, and mechanical properties. Biomaterials 26(June):3363–3376. Abstract available at http://dx.doi.org/10.1016/j.biomaterials.2004.09.004.

Veis, A. 2005. A window on biomineralization. Science 307(March 4):1419–1420. Summary available at http://dx.doi.org/10.1126/science.1109440.

Yamagishi, K., et al. 2005. Materials chemistry: A synthetic enamel for rapid tooth repair. Nature 433(Feb. 24):819. Abstract available at http://dx.doi.org/10.1038/433819a.

Further Readings:

Habelitz, S. … G.W. Marshall, et al. 2004. Amelogenin-guided crystal growth on fluoroapatite glass-ceramics. Journal of Dental Research 83(September):698–702. Abstract available at http://jdr.iadrjournals.org/cgi/content/abstract/83/9/698.

Goho, A. 2004. Diatom menagerie. Science News 166(July 17):42–44. Available to subscribers at http://www.sciencenews.org/articles/20040717/bob9.asp.

Nalla, R.K. … and R.O. Ritchie. 2005. Effects of polar solvents on the fracture resistance of dentin: Role of water hydration. Acta Biomaterialia 1(January):31–43. Abstract available at http://dx.doi.org/10.1016/j.actbio.2004.08.002.

For more information on the National Institute of Dental and Craniofacial Research's "Building A Tooth" effort, go to http://www.nidcr.nih.gov/Funding/CurrentFundingOpportunities/RecentlyCleared/BuildingATooth.htm.

Sources:

Grayson W. Marshall
Department of Preventive and Restorative Dental Sciences
University of California, San Francisco
707 Parnassus Avenue, Box 0758
San Francisco, CA 94143-0758

Janet Moradian-Oldak
Center for Craniofacial Molecular Biology
School of Dentistry
University of Southern California
2250 Alcazar Street, SCA 103
Los Angeles, CA 90033

Robert O. Ritchie
Materials Sciences Division
Lawrence Berkeley National Laboratory
Department of Materials Science and Engineering
University of California, Berkeley
Berkeley, CA 94720

Pamela G. Robey
National Institute of Dental and Craniofacial Research
National Institutes of Health
Building 30, Room 228
30 Convent Drive, MSC 4320
Bethesda, MD 20892-4320

Arthur Veis
Department of Cell and Molecular Biology
Northwestern University
Chicago, IL 60611

Kazue Yamagishi
FAP Dental Institute
3-2-1 Kakinokizaka
502 Meguro-ku
Tokyo 152-0022
Japan


From Science News, Volume 167, No. 20, May 14, 2005, p. 312.