Histological observations on gingival mucosa level determined by fixed partial dentures

Authors: Lucia-Elena Moldoveanu, Ph.D., D.M.D, Assistant Professor, Faculty of Dental Medicine, Ovidius University, Constanta, Romania Mehedinti Teofil, Ph.D., Professor, Faculty of Medicine, Ovidius University, Constanta, Romania Gheju Anca, Ph.D., MD, Anatomical Pathology Laboratory, No.2 Clinic, Municipal Hospital Constanta, Romania Dima Loredana, Ph.D., Lecturer, non-affiliated Corresponding author: Lucia-Elena Moldoveanu, 92 Crinului Street, Constanta, Romania. Phone: +40. 724.871.338; +40.241.630.868 Email address: lucia_moldo@yahoo.com

Key Words: Fixed Partial Dentures, Oral Mucosa, Fibrous chorion, Gingival Epithelium, Papillomatous Lesions.

ABSTRACT Background The study aims to present the results of the microscopic tests obtained by prelevating gingival mucosa from patients with fixed dentures. It used conventional histological techniques for overall examination of tissues. We should mention from the very beginning that this is the first part of a more detailed study which covers a larger number of preparations and more articulated observations. Methods The cytohistopatological study was performed based on the known fact that the interaction between biomaterial and tissue occurs both directly in a very narrow area in the proximity of the prosthetic biomaterial, and indirectly in secondary reactions and interactions situated at a distance and deep in these areas. Cells can interact through molecular groups located on their membranes (membrane receptors, ligands etc.) or through various biochemical substances found in the extracellular matrix (proteoglycans, inorganic substances and so on). Literature information has shown that biomaterials with different chemical compositions trigger different biological responses. Results The results present 18 images of the chorion and the treatment of the gingival mucosa epithelium at the cellular level. It was observed that the biological habitat formed by wearing dentures, as well as different prosthetic materials, can cause various clinical and histopathological changes on gingival mucosa.

Conclusions The cytohistological changes of the gingival mucosa are present in all types of prostheses, but their intensity and variability depends on the biomaterial used in their production. Subjects who used fixed dentures with metal components presented chronic inflammatory reactions, as well as histological reactions depending on the type of metal used in making the denture as follows: fixed dentures with metal part of gaudent revealed histological lesions such as condyloma acuminata, while fixed dentures with metal parts made of wipla predominantly showed papillomatous lesions.

ABSTRAIT Contexte La recherche se propose de prsenter les rsultats des testes microscopiques obtenues travers le prlvement de la muqueuse gingivale des patients qui portent des prothses dentaires fixes. Dans cette recherche on a utilis des techniques histologiques classiques pour lexamen des tissus. Mthode La recherche cisto-pathologique a comme point de dpart la connaissance de linteraction entre le bio-mtal et le tissu, interaction qui se manifeste directement dans une zone trs troite situe prs du bio matriel prothtique o on a faire aussi avec des ractions secondaires et des interactions indirectes distance et au profondeur de ces zones. Les cellules peuvent interactionner comme des regroupements cellulaires aux niveaux de leurs membranes (rcepteurs de membrane et lments de liaisons) ou comme des substances biochimiques qui se trouvent dans la matrice extracellulaire (substances anorganiques). Dans la littrature de spcialit on voit que les bio matrielles ayant de diffrentes compositions chimiques donnent des rponses biologiques diffrentes. Rsultats Les rsultats prsentes 18 images du chorion et du traitement pithlial de la muqueuse gingivale au niveau cellulaire. On a observ que lhabitat biologique form la suite de lutilisation des prothses, comme aussi des divers matriaux prothtiques, peut porter des changements cliniques et histo-pathologiques au niveau de la muqueuse gingivale.

Conclusions Pour toutes les prothses on observe des changements cyto-histologiques de la muqueuse gingivale, mais leur intensit et leur variabilit dpend du bio matriel utilis dans leur fabrication. Ceux qui ont utilis des prothses dentaires fixes ayant du mtal en composition, ont eu des ractions dinflammation chronique, mais aussi des ractions histologiques selon le type du mtal utilis: les prothses fixes ayant en composition du gaudent met en vidences des lsions histologiques type condylome acumin, en tant que les prothses fixes avec du mtal de type wipla ont eu surtout des lsions papillomateuses.

INTRODUCTION The human gingival mucosa is a variety of oral mucosa that forms a unitary and integrated morphofunctional complex in the stomatognathic system, the most exposed to external influences. The gingival mucosa changes caused by fixed partial dentures aggression can determine clinical and histopathological manifestations and provide important data on the oral cavity tissue tolerance to materials used in making dentures. Different prosthetic parts can cause changes of gingival mucosa, some featuring adjustment and disappearing after the adjustment process, while others can cause histological changes with inflammatory character, both in the areas of direct contact with prosthetic material and perilesional, in the proximity or small distance from it. OBJECTIVE: The study of microscopic preparations obtained by prelevating gingival mucosa from patients with fixed dentures (who have agreed on that) was conducted using conventional histological techniques for overall examination of tissues. This is the first part of a more detailed study which covers a larger number of preparations.

MATERIAL

Histological investigations were performed on a total of 83 fragments of human gingival mucosa prelevated from patients wearing unidental and pluridental fixed dentures, during 20022008. The dentures were made of Gaudent copper-based alloys, Wipla, Remanium and acrylate. The study of microscopic preparations was conducted using common colorations to examine the overall tissue architecture. For the comparative analysis a control group was used, with normal clinical aspect, belonging to the same subject. Patients included in the study covered both genders, aged 31 - 74. The selection criteria considered patients with no affections associated to oral mucosa or registered with general affections. The patients came from the geographical environment of Dobrogea, both urban and rural areas. Out of the 45 patients wearing fixed prostheses, 15 patients had pluridental fixed partial dentures, while 30 patients had unidental fixed prostheses. The patients had worn fixed dentures for at least 10 years, and the material used in making dental prosthesis consisted of Wipla, Gaudent, Remanium and acrylate.

No. Topography of prelevated edentulous ridge 1. crt. 2. 3. 4. 5. 6. Agrressed maxillary edentulous ridge Maxillary edentulous ridge to fixed prostheses with normal clinical aspect Agrressed mandibular edentulous ridge Mandibular edentulous ridge with normal clinical aspect Agrressed marginal periodontium Total of prelevated fragments of gingival mucosa

No.of fragments 14 10 19 10 30 83

Table 1. Topographical distribution of prelevated fragments of gingival mucosa

The size of the prelevated fragments of gingival mucosa was 3-4 mm in surface, and in depth the excision was done up to the periosteum of maxillary bone.

METHODS The cyto-histological and histometrical study was based on the histologic technique of paraffin inclusion used in the Laboratory of Cytohistology of the Faculty of Medicine Constana (Professor. Dr. Teofil Mehedini). The histopathologic exam necessary for the speciality diagnosis was performed in the Anatomical Pathology Laboratory, No.2 Clinic, Municipal Hospital Constanta (dr.Gheju Anca). The histologic sections were stained both in common dyes with hematoxylin and eosin, as well as in Massons trichrom for overall structures.

RESULTS During optical microscopy there were observed acute inflammatory lesions of the perilesional tissue with vascular dilatation and inflammatory exudate (by plasma extravasation, of figurative elements, etc.) (Fig.1)

Fig.1:

Chorion

with

lymphocyte-plasmocyte-granulocyte

polymorphous

inflammatory infiltrate, hematic hyperemiated and extravasated capillaries. Col. Massons trichrome, ob X 40 In chronic inflammations of perilesional mucosa there were found proliferative lesions in the lesional focal area. The infiltrated chorion appears to be thickened with a conjunctivevascular tissue of neoformation and limfoplasmocitar infiltrate with diffuse or nodular appearance formed by reaction cells (lymphocytes, macrophages). (Fig.2)

Fig.2: chorion with granulomatous infiltration with nodular disposition. Col H.E., obx10 The congestion of chorion was found at the level of periprosthetic gingival cuff. In some patients there was found a gingival mucosa with no inflammatory lesions, but there are degenerative lesions both in the epithelium level (cytoplasmic vacuolation) and on the chorion level. (Fig.3)

Fig.3: Epithelial cytoplasmatic vacuolation. Col.H.E., obX40

There were also observed the widening of the intercellular space and desmolysis lesions, especially within the suprabasal layer. (Fig.4)

Fig.4: Desmolysis in the epithelial suprabasal layer. Col.H.E., obX40 The examination of the basal shell of periprosthetic gingival epithelium showed significant changes at the level of the junctional systems, both intercellular systems and between the basal pole of epithelial cells and the basal membrane, changes manifested by partial dislocations of the epithelial stratum from the subjacent chorion. (Fig.5)

Fig.5: Partial dislocations of the epithelial stratum from the subjacent chorion. Col.H.E., obX40

The papillary chorion looks to be low in cells and richer in fibers of collagen type I, and it also presents vasodilatation and edematous infiltrate. (Fig.6)

Fig.6: Fibered chorion witn numerous capillaries. Col.H.E., obX40 The loss of connections between epithelial cells leads to the formation of cleavages, blisters or bubbles (acantholysis). At the cellular level, the gingival mucosa epithelium showed both nuclear changes, such as picnosis, cariorexis or karyolysis, and changes of the tinctorial reaction and the number of nuclei or nuclear area. (Fig.7)

Fig.7: Diminishment of the epithelial nuclei number. Col.H.E., obX40

The microscopic examination of gingival mucosa epithelium also revealed modifications of cell cytoplasm, manifested by vacuolation or replacement of these cells with koilocytes, cells

modified by prosthetic stress, changes accompanied by nuclear involutions. The presence of these cells was found throughout the thickness of the epithelium. (Fig.8)

Fig.8: Koilocytes. Col.H.E., obX40

Cells showing small core, double or multiple, with chromatin in the form of islands of different shape and size, without nuclear membrane. The core is surrounded by an optically clear perinuclear cavity, bordered by an intense eosinophilic cell membrane. The stratified squamous epithelium shows acantholysis type changes; increased epithelial thickness by squamous cell hyperplasia with hypergranulosis of stratum corneum (Fig.9)

Fig.9: Squamous epithelial hyperplasia with acantholysis, Col.HE, obX10

Superficial chorion papillae appear elongated and epithelial ridges deepen toward the profound layer of the chorion, having the appearance of papillomatosis. Ob.20 analyzed sections and stained with Masson method highlights collagen fibers arranged in thick fuxinophil bundles, targeted at different incidences. In some regions, there are observed deposits of hyaline dystrophy. In some cases there were noted, at the level of paraprosthetic gingival mucosa, papillomatosis type lesions, in some areas the interruption of the continuity of junction epithelium - lamina propria (basement membrane). These lesions have facilitated disruption of selective biological control of basement membrane upon the tissue fluid content in the chorion extracellular matrix. In this way, epithelial-mesenchymal interactions, established already in the embryonary life, are changed, getting other meanings and other parameters. On some sections of the same batch there were found acute inflammatory processes with the predominance of polymorphonuclear neutrophils, phenomena of increased leukocyte margination, as well as phenomena of angiogenesis, with risk of neoformation arranged as islands. There were also observed areas of necrobiosis, both at the level of papillary chorion and at the level of the profound chorion. Fibers lose their individuality, fibril structure and fundamental substance, creating a unique mass. Fibrinous degeneration appears to be homogenous with eosinophilic appearance. The collagen fibrins break apart into fragments, decompose into fibrils and finally dissolve with fibrinoid appearance. In other areas, at the level of lamina propria, a disorganized structure of lax type, with an edematous infiltration, was noted.

The microscopic examination of the lamina propria, together with reaction cells characteristic for acute or chronic inflammatory processes, the hypertrophy of the nerve fascicles is observed, some of them presenting phenomena of reactive neuritis adjacent to the necrobiotic area. On microscopic examination of histological sections obtained from fragments of human gingival mucosa prelevated from patients with fixed dentures with metal part of Gaudent S, there were detected condylomatous lesions with changes such as acanthosis and parakeratosis type, with keratinization of both the superficial layer with microscopic appearance of squames and corneous globes. Parakeratotic elements have rests of nuclei, no granular layer. There were also been highlighted fibersclerosed areas with network disposition. At the junction epithelium - lamina propria there are observed infiltrates with reaction cells (monocytes, lymphocytes, plasma cells) scattered diffusely. In the distal areas of the interpapillar epithelial ridges there are observed cell areas showing cariolisis and cariopicnosis. Superficial areas of the gingival epithelium present numerous cells with vacuolated cytoplasm. The cavitary alteration of the cell occurs as an early cellular edema within the malpighiene epithelial cells. In some cases there are aspects of reticular degeneration by bursting of the cells and creation of multinucleated blisters. In the deep chorion, the connective fibers appear dissociated from edematous infiltration; there is also noted the hyperplasia of nervous fillets and reactive neuritis. The contact surfaces of the prosthesis with the gingival mucosa show small ulcerative lesions and below them there is a fibersclerosed edematous connective tissue.

DISCUSSIONS The citohistological study of gingival mucosa lesions belonging to the prosthetic field is of great importance. It was observed that the biological habitat formed by wearing dentures, as well as different prosthetic materials, can cause various clinical and histopathological changes on gingival mucosa. During the first period of use of the prosthesis may occur citohistological changes that are adaptive, they disappear after a period of adjustment, in other cases due to distress (chronic stress) at the gingival mucosa level in contact with denture may occur changes of the citohistological picture which may indicate the presence of inflammatory or proliferative chronic lesions. The human body is perfectly adapted to certain conditions such as gravity, but it is also versatile, able to adapt to different conditions in order to maintain body homeostasis. Any change in the external environment determines the human body to adapt to annihilate the harmful action and lead to maintaining or restoring the homeostasis. The excessive action of some environmental factors (pressure, postural demands, biological and chemical factors of the environment etc.) can affect the homeostasis and create a pathogenic potential for various diseases. From the microscopic analysis of the histological preparations there was found that the cytohistological changes of the periprosthetic gingival mucosa are present in all types of prostheses (unidental or pluridental), but their intensity and variability depend on the biomaterial used in their manufacture. In 1994 Steflik published a detailed study regarding the necessary conditions for a biomatwerial to ensure the two sequences of biocompatibility:

- Primary sequence, which considers the short-term reaction to achieve gingival mucosaprosthesis interface; - Secondary sequence, on long term, where the direct interface is subject to functional demands called "functional biocompatibility. There is experimental data in literature showing that biomaterials with different chemical compositions trigger various biological responses. In this respect, the chemical composition of the contact surfaces is mainly determined by the action of metal oxide (Kasemo, 1983). Metal oxides influence the cellular connection type of the interface prosthesis-periodontium. There were reported dynamic disorders of blood circulation, with altered vascular permeability and inflammatory exudate formation, either diffuse or nodular. The gingival mucosa showed inflammatory and degenerative lesions both in the epithelium and in the chorion. At the cellular level changes occur in all cellular components (cytoplasmic membrane), but especially at the nuclear level. These changes are detectable microscopically both directly and indirectly through quantitative methods of biometric type. It is known that, of all the constituent components of the cell, the nucleus is the one having the most comprehensive participation, both in physiological processes, and in pathological ones, although such participation is not always reflected by morphopathological changes. That is why we have included in our study, in addition to the microscopic study exploring the visible microcosm, a biometric study that explores the unseen interior of the microcosm. The nucleus is involved in almost every activity of response to the stress apllied upon the cell. The altered function of the nucleus might be the cause of some cytoplasmic dystrophic lesions. During the stage of eustress generated by the action of the dentures, some lesions that

appear may be transitional, since the cell lesion is reversible as long as the nucleus is still unaffected. Morphological lesions of the nucleus are relatively poor and nonspecific. The correlation of the morphological data with the genetic, quantitative and cytological translates into determining changes in the structure of the nucleus, so if the cell is chronically injuried, there is an activation of the nuclear chromatin and the nucleus becomes heterochromatic, the basophilia of the nucleus is erased. This process is described as pyknosis, a process that is largely due to the permeability of cellular and nuclear membranes for calcium ions. It blocks the negative phosphate groups of nucleic acids causing changes of nuclear basophilia and chromatin aggregation. The nucleus lysis - karyolysis - may not be preceded by the appearance of pyknosis if the nuclear membrane is affected and lysosomal enzymes are activated early. Hyperplasia occurs in response to excessive stimuli. It may precede benign or malignant neoplasms. Our study reveals results compared to those obtained in similar studies such as the ones cited below. A research conducted by Bergendal and Isacsson (1983) has shown, at the level of periprosthetic gingival mucosa, the existence of nonspecific inflammatory reactions

characterized by the presence of a stratified malpighian epithelium with parakeratosis, marked acanthosis and spongyosis, and a mononuclear inflammatory infiltrate and capillary vasodilatation at the level of the subjacent connective tissue. Schroder (1996) considered that chronic inflammation of the gingival mucosa might lead to epithelial dysplasia or metaplastic changes.

Ursache (1996) cites histopathological changes of the gingival mucosa in acrylic denture wearers, manifested by hyperplastic epithelium, with filiform or digitiform elongated ridges, with ortokeratosis or parakeratosis with inflammatory phenomena, atrophic epithelium with acanthosis and discheratosis, and at the chorion level with collagen bundles with edematous reaction, the basement membrane with uneven thickness and trajectory, with interruption of continuity. Costin et al (1998) observed, in patients with gaudent prostheses, subacute and chronic inflammatory changes. At the level of lamina propria, the authors noted the presence of dilated vessels, the limfoplasmocitar perivascular inflammatory infiltrate, and hemosiderin granules. A study by Minic et al (1991) on the oral mucosa necropsied from individuals who had worn dentures showed epithelial calcification sources, chronic inflammatory infiltrate in the gingival stroma, presence of granulation tissue, connective tissue myxo degeneration. Tovaru (1999), Staniceanu (2001) emphasized the role of HPV (Human Papilloma Virus) in condylomatous lesions in the oral area. This can cause both benign and malignant tumors. The koilocytes described by Koss (1976) as a squamous cell of superficial or intermediate type, with the nucleus (with HPV parasites) being single, double or multiple, present chromatin of variable shapes and sizes, without apparent nuclear membrane, without nucleoli. The nucleus is always surrounded by an optically clear perinuclear cavity, margined at the periphery by a well-defined cytoplasmic membrane, with eosinophilic, cianofil or policromatofil stain (Meisels et al, 1992). Perinuclear vacuole usually contains usually cyanophilic remains.

In our study, in the mucosae with papillomatous lesions we have shown koilocytes in the superficial layers accompanied by nuclear involutions of cariorexis or karyolysis type. We have met the presence of koilocytes on the entire thickness of the epithelium. Sajin et al (1999) stated that the citomorphologic diagnosis of the condylomatous lesions is based on two pathognomonic (specific) elements: specific koilocytes and dyskeratosis. Trusal et al (1975), Simirad et al (1986), Rihet et al (2000) have continued their studies in this direction.

CONCLUSIONS The biomaterial used in making unidental or pluridental fixed dentures can interact with the components of periodontium and determine clinical and histopathological changes. Some of these are adaptive changes which disappear after a period of time, others are changes associated to physical or biochemical distress leading to inflammatory or proliferative chronic lesions. During the microscopic examination of histological preparations there was found that the citohistological changes of the gingival mucosa are present in all types of prostheses (unior pluridental), but their intensity and variability depends on the biomaterial used in their production. In subjects who used fixed dentures with metal part of gaudent there were present, besides chronic inflammatory reactions, histological lesions such as condyloma acuminata. The microscopic study of gingival mucosa fragments obtained from patients with fixed dentures with metal parts made of wipla predominantly showed papillomatous lesions. On the paraprosthetic condylomatous gingival mucosa sections there was found, on the entire thickness of the epithelium, the presence of koilocyte shape cells, cell vacuolation, picnotic nucleus pushed to the periphery with involutive lesions involution, cariorexis and karyolysis. The nuclear involutive lesions (the nucleus being the vital center of the cell) induced changes of the whole cell metabolism, which favored dysmetabolic or metaplastic processes. The pattern of citohistological changes of gingival mucosal from patients who have worn dentures with wipla type metal component had papillomatous changes.

BIBLIOGRAPHY

1. Banita M., Tissue histology course. The tissues, vol.1, Sitech Ed., Craiova, 2000 2. Bath Balogh Mary, Fenerbach J. Margaret, Dental Embryology, Histology and

Anatomy, Ed. Elsevier Saunders, 2006

3. Bodnar V., Bodnar T., Drafta S., New aspects of oral galvanism, National Dentistry

Magazine, 1998, I12: 36-39

4. Bratu D., Uram-uculescu S., Leretter M., Romnu M., Based-copper Alloys in Dentistry

(Part I), The National Journal of Dentistry, National Ed., 1998, vol. I: 3, 6-14
5. Ibid., Based-copper Alloys in Dentistry (Part II), The National Journal of Dentistry,

National Ed., 1998, vol. I, no. 4-5: 5-24

6. Bratu D., Nussbaum R., The chemical basis of fixed prosthetics, Diagnosis, Observation

Sheet in Fixed Prosthetics, Sigmata Ed., Timioara, 2001

7. Costin G., lonescu Gh., Ursache Maria, Considerations on the biocompatibility of

conjunctive prosthetic dentures made of Al-Cu-Ni alloys, Dental Medicine, 1998, vol. 2, no. 4: 20-24
8. Costin G., Ursache M., Gheorghiu R., Modification of interfacial pressure of dental

biomaterials, Dental Medicine, Vol. 3, Nr. 3, 1999

9. Donlan RM, Costerton JW., Biofilms: survival mechanisms of clinically relevant

microorganisms, Clin Microbiol Rev 2002; 15: 167-193

10. Forna Norina, Assessment oh health status affected by edentation, Ed. Demiurg, 2007 11. Gartner P. Leslie, Hiatt L. James, Color Atlas of Histology. Ed. Lippincott Williams &

Wilkins, February 2009

12. Geurtsen W., Toxicology of Dental Materials and Clinical Experience, J DENT RES

July 2003 82: 500

13. Ibid., Biocompatibility of dental casting alloys, CROBM, January 2002; vol. 13, 1: pp.

71-84
14. Gladwin M, Bagby M., Clinical Aspects of Dental Materials, Baltimore, Md: Lippincott

Williams & Wilkins; 2004

15. Haulica I., Human Physiology, second edition, Ed. Medicala, Bucureti, 2007 16. Holdich R., Starov V. M., Prokopovich P., Njobuenwu D. O., Rubio R., Zhdanov S.,

Velarde M. G., Spreading of Liquid Drops from a Liquid Source, Colloids and Surfaces A: Physicochemical Engineering Aspects, 282283, p. 247-255, 2006
17. Humphrey Sue P., Williamson Russell T., A review of saliva: Normal composition, flow,

and function. J Prosth. Dent, 2001, 85(2):162-169

18. Khasawneh S., Al Wahadni A., Control of denture plaque and mucosal inflammation

in denture wearers. J. Ir. Dent Assoc, 48(4): 132 8, 2002

19. Lockhart S. R., Joly S., Vargas K., Swails Wenger J., Enger L., Soll D. R., Natural

defenses against Candida Colonization breakdown in the oral cavities of the elderly. Journal of Dental Research, Vol 78, 857 868
20. Mehedinti T., Mehedinti, R., Hincu M. Moyle, Histology of the Orofacial Biosystem,

Dunarea de Jos University Foundation, Galati, 2006

21. Mogoanta L., Hincu M., Guide to Techniques of Histology,

Cytology and

Immunohistochemistry, Medical University Ed., Craiova, 2003

22. Mogoanta L., Hincu M., Mehedinti T., Medical Histology, Aius Ed., Craiova, 2004

23. Murray P.R., Baron E.J., Jorgensen J.H. et al, Manual of clinical microbiology, 8th ed,

ASM Press, Washington, 2003

24. Oprian Alexandra, Elements of Oral Pathology: Anatomy and Histology of Oral Cavity,

Medical Publishing House, Bucharest, 2001

25. Sadighpour L., Geramipanah F., Raeesi B., In Vitro Mechanical Tests for Modern Dental

Ceramics. Journal of Dentistry, Tehran University of Medical Sciences, Vol. 3, No. 3, pg 143-152, 2006
26. Scrieciu Monica, Crioiu M., Influences of fixed dentures on gingival mucosa, Medical

Craiova Journal, 2002, 4: 41-44

27. Tovaru Serban, Medical Dental Pathology, Editura Cerma, Bucharest, 1999: 23-47 28. Turssi, C.P., Ferracane J.L., Vogel K., Filler features and their effects on wear and

degree of conversion of particulate dental resin composites. Biomaterials, 26: 49324937, 2005
29. Tyas Martin John, Clinical evaluation of glass-ionomer cement restorations. J. Appl.

Oral Sci. vol.14 no.spe Bauru, 2006

30. Ursache Maria, Paraprosthetic Stomatopathy, Ed. Ankarom, Iasi, 1999 31. Van Noort R, Gjerdet N, Schedle A, Bjorkman L, Bergland A., An overview of the

current status of national reporting systems for adverse reactions to dental materials. J Dent 32(5): 351-358, 2004.