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Materials Science: Biological Aspects

Poliklinik für Zahnerhaltung und Parodontologie, Klinikum der Universität, D-93042 Regensburg, Germany; Gottfried.Schmalz{at}klinik.uni-regensburg.de

KEY WORDS: dental materials • toxicity • biocompatibility • cell culture • standardization

	THE FIRST STEPS

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Biological aspects of dental materials have received scientific interest ever since they were used in patients. However, during the first half of the last century, these aspects were apparently not considered to be very important; e.g., standards (specifications) for dental materials developed in the 1920s to guarantee their quality covered only technical properties, not biological aspects. The same was apparently true for materials used in medical applications (Autian and Brewer, 1958).

However, the number of publications slowly increased until the middle of the last century, but the scope of biological aspects was restricted to the adverse effects of dental materials. Two main streams of interest could be observed. The first one concentrated on local effects—e.g., pulp problems associated with dental filling materials such as silicate cements (Fasoli, 1924), based on the clinical observations of pulp necroses under silicate fillings. In the second stream, the systemic adverse effects of one material—namely, amalgam—was the center of not only scientific (and non-scientific) reports but also of strongly emotional public discussions.

	DEVELOPING NEW TECHNIQUES

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In the 1950s and ’60s, a larger number of experimental studies on adverse effects elicited by dental materials was published in parallel with the development of new resin-based materials and new treatment techniques, such as high-speed air rotors. Again, two main streams in the literature could be distinguished. In one stream, animal studies were conducted, in which the materials were applied as they would be in patients and the tissue reactions were observed by histological techniques. The experimental techniques involving experimental animals, mainly non-human primates, were improved by standardization of the histological techniques and evaluation procedures (Langeland et al., 1966; Baume et al., 1972; Klötzer and Langeland, 1973).

The second main stream concentrated on the evaluation of more basic biological properties, e.g., using cell culture techniques. These methods became available to dental research during the 1950s, and one of the pioneers in this field was H. Kawahara and his team in Japan (Kawahara et al., 1955). In Europe, similar studies were initiated (Maizumi and Sauerwein, 1962).

	MATERIALS SCIENCE TOXICOLOGY

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Supported by stipends from both the University of Tennessee and the German Research Foundation (DFG), I had the chance and the privilege in 1974 to join one of the emerging laboratories devoted to a systematic and comprehensive approach to the study of the adverse effects of dental (and other biomedical) materials, the "Materials Science Toxicology Laboratory (MST)". It was based upon an interfaculty agreement between the Colleges of Dentistry and Pharmacy of the University of Tennessee (Memphis), and it was devoted to the study of the toxicity of biomaterials (dental and medical), their ingredients, their interaction with drugs, and to safety and standardization aspects, thus merging materials science and pharmacology/toxicology.

The Director of and the driving force behind MST was J. Autian. He started his career with a study on the biocompatibility of synthetic high-molecular-weight polymer ("plastic") materials used—e.g., in tubings and syringe hubs (Autian and Brewer, 1958). Analysis of his data suggested that problems may occur when a plastic material is used in a pharmaceutical application without adequate testing being carried out first. In 1960, at the University of Texas, Austin (School of Pharmacy), he continued his work on the toxicity and drug interactions of plastic medical materials. During this time, the field of materials toxicity attracted post-doctorate and graduate students, and test methods were developed such as the agar diffusion test, which today is included in most biological standards. Attention was also paid to the elution and biological effects of plasticizers, e.g., phthalate esters, a topic which has kept its significance even today. In 1967, Autian was appointed a professor in the Colleges of Pharmacy and Dentistry and director of the MST laboratory.

MST was divided into four sections (heads in parenthesis), reflecting its different roots: (1) physical chemistry (L. Nunez), (2) cellular toxicology (E.O. Dillingham), (3) animal toxicology (H. Lawrence), and (4) pathology (J. Turner). Teaching obligations in both Pharmacy and Dentistry were a further basis for intensive interdisciplinary exchange. A graduate program in toxicology and materials science (MS and PhD) had been established. From 1968-1986, basic aspects of toxicity testing of biomaterials were established and different tests developed, refined, or adjusted from general toxicology to be used for materials testing, e.g., the agar diffusion method, the (static) hemolysis test, and the inhibition of cell growth test (all in vitro), or the rabbit intramuscular implantation test and LD₅₀ tests (in vivo). Most of these tests have become classics, and many are still in use today (Schmalz, 1997). Furthermore, the idea of the structure-related activity of chemicals was applied to toxicology with substances used in plastic medical devices, like acrylates and methacrylates (Dillingham et al., 1983). In the 1970s, MST received a contract from FDA to develop tests for the safety of materials to be used in medical applications. A set of tests known as the "MST Primary Acute Toxicity Screening Program" was devised, based on the philosophy that safety testing of materials must not be based on a single test alone, but on a set of tests (Autian, 1977).

MST trained people from industry, and visiting scientists from all over the world, including the author of this article, were invited every year, and thus the idea of materials toxicity being a relevant aspect of biomaterials was further spread. In 1986, J. Autian retired from the University of Tennessee after he had been Dean of the School of Pharmacy and—later—Vice-Chancellor (Health Science Center).

Other groups worldwide became active or enlarged their activities in this field. Standing in for these researchers, only a few can be mentioned here. NIOM (Nordisk Odontologisk Materialprøvning) was founded in 1972 by the Nordic countries in Europe. Its first director was I. Mjör, followed in 1995 by A. Hensten-Pettersen. Like MST, a spectrum of biological test methods was applied, but the activities also included technical and chemical testing. NIOM was partially funded by the Nordic States and partially by industry, which was required to have NIOM approval before they could place their materials on the market in these countries. The great influence of NIOM was based on its legal status as well as on its publication activities. Furthermore, together with our group, much effort was placed on the development of new standards and European legislation.

One of the most active university-based groups was in Connecticut, guided by K. Langeland, with the impetus on pulp/endodontic studies in monkeys. Later, this group was joined by L. Spangberg, introducing the Cr⁵¹ method as a cell culture technique into dental materials testing (Spangberg, 1973). H. Stanley at the University of Florida published important studies on pulp reactions to dental filling materials and to pulp capping. The field of biological aspects was now broadened from adverse effects testing to the evaluation of beneficial biological properties, like promoting healing or new tissue formation, an area which today is called tissue engineering.

	SETTING UP STANDARDS

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It had been an essential aim of MST, and others, to promote the development of standards for the biological testing of dental materials, to have an effective safety testing program and to ensure the comparability of the results from different laboratories. In 1972, the ADA published recommendations on standard practices for biological evaluation of dental materials, which, after revision, were issued in 1979 as ANSI/ADA document No. 41 and which was the basis for our further activities. Other national standards in this field were set up, e.g., in Germany (Schmalz and Klötzer, 1986). Discussions arose as to which tests should be performed and whether some of these tests should be mandatory, the latter especially related to the expensive, time-consuming, and publicly discussed (animal protection societies) animal tests. Experience from drug testing showed, however, that a system of mandatory tests with defined criteria for failed/accepted (as is traditional in technical testing standards) could not be transferred to the area of biocompatibility evaluation, because this approach does not account for the diversity of the reactions observed and the different methods used for evaluation. This has been recognized by the newest standards (ISO 10 993-series), where the final choice of the relevant tests and the interpretation of the test results fall to the responsibility of an expert (Schmalz, 1997).

In 1980, the FDI’s "Recommended standard practices for biological evaluation of dental materials" was published under the leadership of K. Langeland. Under the wise guidance of H. Stanley, finally, in 1997, a full standard on the "Preclinical evaluation of biocompatibility of medical devices used in dentistry—Test methods for dental materials (ISO 7405)" was finalized (International Organization for Standardization, 1997). Interestingly, international standards on the biocompatibility of biomaterials in general were based to a large extent on the experiences of scientists from dentistry. The result of this work was the ISO 10993 series, which by now is transferred to European standards (EN 30 993-series) having—within the EU—an almost "law-like" status. ISO 10993 and ISO 7405 complement one another.

	LEGAL REGULATIONS AND PUBLIC DISCUSSIONS

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Scientific reports on biological hazards from (dental) biomaterials have resulted in legal regulations for the safety of the consumer (patient), of the dental team/technician, and for the protection of the environment (Arenholt-Bindslev, 1998). To the knowledge of this author, the first legal regulations were the "Medical Device Amendments", which passed the US Congress in 1976. According to this regulation, an important aspect is safety, and the FDA is in charge of installing and enforcing corresponding regulations. Within the European Union, legislation on biomaterials resulted in the Medical Device Directive (MDD, 93/42 EWG), which became effective in 1998. During the formulation of this directive, it was discussed whether dental materials should be regarded as devices or as drugs. Experiences in Germany, where at that time (1980s-90s) most dental materials were legally regarded as drugs, were not encouraging, because most aspects of drugs (dose, solubility, metabolism) were difficult if not impossible to apply reasonably to dental materials as they were applied to drugs. Therefore, researchers from NIOM, together with the author of this article and other colleagues, favored the inclusion of dental materials in the medical device legislation.

The transformation of clinical observations and scientific reports into legislation sounds like a real success story of science, directly demonstrating the beneficial impact of research on the quality of life of the population. However, a few dental materials recently being marketed showed biocompatibility problems, e.g., cases of severe pulpal pain, which needed revision of the restoration. This demonstrates the limits of legislation, of pre-clinical testing, and the importance of clinical testing over a given period of time. It also demonstrates that this sort of legislation needs constant revision.

In this context, the biological aspects of dental materials have attracted considerable public concern which cannot be dealt with in detail here, but which is specific to this area of research. Many researchers were involved in public discussions and were members of national and international committees set up by politicians/administrations to define a basis for political decisions, e.g., to forbid, or to restrict, the use of a material by legal regulations. A main problem in this context, which this and other authors observed during these public discussions, is that it is apparently difficult for the lay press to put data from toxicity tests into the right perspective and to critically analyze the data with respect to the health risk of the population. This is considered to be the task of a scientist, and therefore the involvement of scientists in these discussions is necessary.

	NEW STRATEGIES

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Based on the MST concept, we installed in the new Dental School at the University of Regensburg an interdisciplinary research group (dentistry, biology, mathematics, chemistry). Similar approaches were chosen elsewhere. As a new strategy, research did not only concentrate on the phenomena, like pulp damage, but it also focused on mechanisms, to be able to improve materials and techniques. One example is the theory of bacteria instead of materials being the cause of pulp reactions (Brannstrøm and Nyborg, 1971; Bergenholtz et al., 1982). This theory has been widely accepted today as one possible mechanism, and it has resulted in the claim for bacteria/toxin tight fillings.

In line with the attempts to elucidate the mechanisms, research proceeded from cellular toxic to subtoxic effects. One example is mutagenicity; apparently, components of dental materials, such as TEGDMA or BADGE, may interfere with DNA, a cause mutation which can be transferred to future cell generations (Schweikl et al., 1998; Schweikl and Schmalz, 1999; Geurtsen and Leyhausen, 2001). Other research concentrated on the influence of dental materials and their constituents on inflammation mediators (Schmalz et al., 1998, 2000). This approach seems very interesting to us, because it may show a direct biochemical link between the parameters measured in vitro and clinical effects (inflammation) in vivo. Other groups concentrated on phospholipids and glutathione (Engelmann et al., 2001), on estrogenic effects (Wataha et al., 1999; Hashimoto and Nakamura, 2000), or on heat-shock proteins (Noda et al., 2002). Thus, the influence of materials on cell metabolism has become a topic of research. Close contact with basic science groups, such as pulp and cell biology, are of utmost importance for further development—e.g., using microarray techniques.

Another trend is the increasing tendency to replace animal tests by in vitro cell culture methods. Apparent discrepancies between results from in vitro tests and those from animal or human studies may be overcome by the new design of test methods, taking the in vivo anatomical situation into account in the in vitro situation. One example is the Dentin Barrier test (Tyas, 1977; Meryon et al., 1983; Hanks et al., 1988; Hume, 1988). Through genetic engineering, cells can be constructed that may meet the requirements for a standardized test and which, on the other hand, better represent the in vivo target tissue (Thonemann and Schmalz, 2000). Standardizable test methods are today based on commercially available equipment, which can be combined with three-dimensional cell cultures with constant medium perfusion simulation blood flow (Schuster et al., 2001; Schmalz et al., 2002).

	CONCLUSIONS

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A look into current textbooks of dental materials shows that the biological aspects have become an indispensable part of materials science during the last century, with impact on legal regulations and scientists being involved in political discussions. The scope of biological aspects of dental materials will further be widened. Activities will no longer be restricted to adverse effects, but will extend to "positive" interactions with the living tissue. An example is the incorporation of signal molecules into materials to stimulate dentin apposition or bone growth (Decup et al., 2000). The concept of a biomaterial, which in every application must be inert, is and will in the future further be abandoned in lieu of an active interaction with cell metabolism.

Research on biological aspects of dental materials was possible only as a result of the interdisciplinary approaches of dentists with disciplines like pharmaceutical science, toxicology, chemistry, and biology. Even more, it was shown that the basic problems, strategies for their solution, and the single methods are very similar for biomaterials with both dental and medical applications, which again shows that dentistry is an integral part of the medical scene.

Received June 24, 2002; Accepted July 22, 2002

	REFERENCES

TOP THE FIRST STEPS DEVELOPING NEW TECHNIQUES MATERIALS SCIENCE TOXICOLOGY SETTING UP STANDARDS LEGAL REGULATIONS AND PUBLIC... NEW STRATEGIES CONCLUSIONS REFERENCES

 
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