CLINICAL DENTISTRY AND RESEARCH 2015; 39(1): 4-11 Original Research Article

CorrespondenceChong Huat Siar, BDS, MSc, FDSRCPS, FRCPath

Department of Oro-Maxillofacial Surgical and Medical Sciences

Faculty of Dentistry, University of Malaya,

50603 Kuala Lumpur, Malaysia,

Phone: +603 7967 4859,

Fax: +603 7967 4531,

e-mail: siarch@um.edu.my

Chong Huat Siar, BDS, MSc, FDSRCPS, FRCPathProfessor, Department of Oro-Maxillofacial Surgical and

Medical Sciences, Faculty of Dentistry, University of Malaya,

Kuala Lumpur, Malaysia

Chee Keong Pua, BDS, MScPrivate Dental Practitioner

Georgios Romanos, DDS, PhDProfessor, School of Dental Medicine,

Stony Brook University,

New York, United States of America

Chooi Gait Toh, BDS, MSc, FDSRCPS, DRD, FDSRCS Dean and Professor, School of Dentistry,

International Medical University, Kuala

Lumpur, Malaysia

Kok Han Ng, BDS, MSc, FDSRCPS, FRCPath, Former Director and Senior Consultant Oral Pathologist,

Unit of Stomatology, Institute for Medical Research,

Jalan Pahang, Kuala Lumpur, Malaysia

THE EFFECT OF IMPLANT-SUPPORTED FIXED PROSTHESIS ON THE PERIODONTAL LIGAMENT WIDTH AND STRUCTURE IN

OPPOSING NATURAL TEETH OF MACACA FASCICULARIS

ABSTRACT

Background and aims: Although rehabilitation of the partially

dentate jaw with implant-supported fixed prosthesis has achieved

high success rates, little is known regarding their effects on the

opposing natural dentition. The periodontal ligament (PDL) functions

as a dynamic attachment apparatus, and responds to occlusal forces

by widening/narrowing and alterations in collagen fiber density/

orientation. This study aimed to determine the effects of occlusal

load from implant-borne bridgework on PDL of the opposing natural

teeth.

Materials and Methods: Test samples consisted of four healthy

adult male Macaca fascicularis with implant-supported 3-unit

bridgework at the second premolar-second molar regions of their

mandibles – one side for immediate-loading and the other side for

delayed-loading, in a split mouth design. Another pair of monkeys

with natural dentition served as normal controls. After 3 months

of functional loading, the animals were sacrificed. Non-decalcified

sections were prepared for histomorphometric assessment of PDL

structure, cervical and apical widths.

Results: PDL collagen fiber density and orientation were similar in

both groups and no signs of inflammation were present. The width

of PDL of the teeth in the experiment group at the cervical and apical

regions were significantly increased compared to control (p<0.05).

Conclusion: PDL widening might reflect a functional response to

occlusal forces from the ‘rigidly ankylosed’ implant-supported fixed

prosthesis.

Keywords: Animal Studies, Functional Loading ,Immediate

Loading, Implant, Natural Dentition, Periodontal Ligament,

Submitted for Publication: 11.10.2014

Accepted for Publication : 03.30.2015

4

5

CLINICAL DENTISTRY AND RESEARCH 2015; 39(1): 4-11 Orjinal Araştırma

Sorumlu YazarChong Huat Siar Malezya Üniversitesi

Diş Hekimliği Fakültesi Ağız, Diş ve Çene Cerrahisi Anabilim Dalı

50603 Kuala Lumpur, Malaysia,

Telefon: +603 7967 4859,

Faks: +603 7967 4531,

E-mail: siarch@um.edu.my

Chong Huat Siar, Prof. Dr., Malaya Üniversitesi Diş Hekimliği Fakültesi,

Ağız, Diş ve Çene Cerrahisi Anabilim Dalı,

Kuala Lumpur, Malezya

Chee Keong Pua, Dt., Serbest Diş Hekimi,

Klang, Malaysia

Georgios RomanosProf. Dr., Stony Brook Üniversitesi,

Diş Hekimliği Fakültesi,

New York, Amerika Birleşik Devletleri

Chooi Gait Toh, Prof. Dr., International Medical Üniversitesi,

Kuala Lumpur, Malaysia

Kok Han NgKuala Lumpur Tıp Merkezi, Stomatoloji Bolümü,

Oral Patoloji Direktörü,

Malezya

MACACA FASCICULARİSLERDE İMPLANT DESTEKLİ SABİT PROTEZLERİN KARŞIT ÇENEDEKİ DIŞ YAPISI VE PERİODONTAL

LİGAMENT GENİŞLİĞİNE ETKİSİ

ÖZ

Amaç: Kısmi dişsizlik durumlarında implant destekli sabit protezler

tedavi seçeneği yüksek başarı oranlarıyla uygulanmasına rağmen

bu tedavinin karşıt çenedeki etkileri çok az bilinmektedir. Periodontal

ligament dinamik bir ataçman olarak fonksiyon görür ve kollojen

liflerin sıklığı ve uzanımını değiştirerek okluzal yükler karşısında

kalınlaşıp incelerek yanıt oluşturur. Bu çalışmada implant taşıyıcılı

köprülerin olduğu çenenin oluşturduğu okluzal yükün karşıt çenedeki

doğal dişlerin PDL durumu üzerindeki etkilerinin araştırılması

amaçlanmıştır.

Gereç ve Yöntem: İkinci premolar ikinci molar mandibular bölgeye

implant yerleştirilen 4 adet yetişkin erkek Macaca fascicularis’e

3 üyeli köprüler yapılmıştır. bir yarım çenedeki implantlara erken

yükleme diğer tarafa ise geç yükleme yapılarak iki ayrı tasarım

uygulandı. Doğal dişlenmeye sahip bir çift maymunun ise kontrol

grubu olarak belirlendi. Fonsiyonel yüklemeden üç ay sonra

hayvanlar tıbbi koşullar altında öldürüldü. PDL yapısı,servikal ve apikal

genişliğinin histomorfometrik incelemesi için non-dekalsifiye kesitler

hazırlandı.

Bulgular: PDL kollejen liflerin sıklığı ve uzanımı her iki grupta benzer

bulundu ve enflamasyon bulgularına rastlanmadı. Kontrol grubu ile

karşılaştırıldığında deney grubundaki dişlerin PDL genişliğinin servikal

ve apikal bölgelerde belirgin olarak arttığı belirlendi. (p<0.05)

Sonuçlar: Rijit olarak ankiloze olan implant destekli sabit protezlerin

oluşturdukları okluzal yüklere PDL genişleyerek fonksiyonel bir cevap

verebilir.

Anahtar Kelimeler: Hayvan Çalışmaları, Fonsiyonel Yükleme,

Erken Yükleme, İmplant, Doğal Dişlenme, Periodontal Ligament

Yayın Başvuru Tarihi : 10.11.2014

Yayına Kabul Tarihi : 30.03.2015

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CLINICAL DENTISTRY AND RESEARCH

INTRODUCTION

Nowadays, dental implants have become a widely-accepted therapeutic option for the restoration of the partially dentate jaw.1-5 However, concern has been expressed that with this treatment modality the maximal occlusal bite forces of the ‘rigidly’ ankylosed implant-supported prosthesis has been shown to be significantly greater than tooth-supported prosthesis.1 Moreover, dental implants possess a higher sensory perception threshold than natural teeth.1,6 Although the maximal occlusal forces are greater for the implant in implant-tooth-supported fixed partial prosthesis but this does not adversely affect the time-dependent marginal bone level.7 The periodontium refers to the dental tissue complex that principally functions as an attachment apparatus enabling teeth withstand masticatory forces.8,9 This tissue ensemble comprises the periodontal ligament (PDL), root cementum, alveolar bone and dentogingival junction. Among these components PDL is the most important and is responsible for providing direct support to the tooth, and for maintaining the functional equilibrium/homeostasis of the periodontium. Histologically, PDL is identified as a well-delineated zone of connective tissues that occupy the space between the tooth root and alveolar bone.9,10 Type I collagen is the principal component of the intercellular matrix that makes up the PDL, and collagen synthesis in the PDL plays an important role in the dynamic remodelling processes during functional tooth movement.10,11 The support function of PDL is influenced by the stress-strain and visco-elastic properties of this tissue.10,12,13 Evidence from finite element analysis indicated that the effect of the periodontal ligament on strain distribution and⁄or absorption is restricted locally to the alveolar bone surrounding the teeth and does not affect other regions of the mandible.12

It is hypothesized that natural teeth opposing the implant-supported prosthesis would be associated with plausible changes in the PDL structure and width. The aim of this study was to investigate the effects of implant-supported bridgework on the PDL structure and width of opposing natural teeth following occlusal loading for a period of 3 months.

MATERIALS AND METHODS

Animals and experimental procedures

The animal experimental design and schedule were approved by the University of Malaya Medical Ethics Review

Figure 1. A-D: Clinical view of test sites with 3-unit splinted metal superstructures in the premolar-molar mandibular region and natural teeth in the maxillary arch. A. immediate-loading (IL) mandibular implant-borne bridgework; B. delayed-loading (DL) mandibular implant-borne bridgework; C, undecalcified semi-thin section of right maxillary molar teeth; and D. undecalcified semi-thin section of left maxillary molar teeth.

Board on Animal Use [Ethics Ref. No. U1/10/6/96/TCG (R)-1]. Animals were randomly assigned to the test and control groups. Test sample comprised four healthy adult male monkeys (Macaca fascicularis) with a mean age above 7 years, mean body weight of 5738.45 g (+ 84.13) and with fully erupted mature dentition. These monkeys had implant-supported 3-unit bridgework placement in the second premolar-second molar regions of their mandibles – one side for immediate-loading (IL) and the other side for delayed-loading (DL), in a split mouth design (Figure 1). The implant data have been previously published.14,15 Control group consisted of maxillary second premolar-second molar jaw segments from another pair of monkeys with natural teeth and no restorations. Briefly, the monkeys were fed standard laboratory chow as recommended for M. fascicularis and water ad libitum. A rigorous oral hygiene program consisting of mechanical cleaning 3 times a week supplemented by chemical plaque control once a week, was carried out during the entire period of study.

Histologic processing and analysis

After 3 months of functional loading, the animals were sacrificed with an intravenous overdose of pentobarbital sodium. Specimens of maxilla were harvested, fixed in 4% neutral buffered formaldehyde and hemisected in a mesio-

7

Implant functIonal loadIng and pdl

distal direction. One half of the bisected specimen was decalcified in 10% EDTA at pH 6.9 and two representative sections were prepared and stained with haematoxylin and eosin. The other half was processed by the Exakt Cutting-Grinding technique (Exakt Apparatebau, Nordestedt, Germany). Briefly, the specimens were dehydrated in an ascending series of alcohol rinses, cleared in xylene, infiltrated and embedded in glycolmethacrylate resin (Technovit 7200 VCL resin, Kulzer, Wehrheim, Germany). Upon completion of polymerization, these specimens were sectioned with a diamond saw to about 200mm and ground down to about 10mm (Figs. 2C, D). At least 2 representative sections were obtained and these were stained in 1% toluidine blue. Both the decalcified and nondecalcified sections were subjected to descriptive and quantitative analysis.Histologic and histomorphometric analyses were performed by two investigators (C.H.S. and C.K.P.) blinded to the clinical and surgical phases of the experiment. A light microscope (Reichert-Microstar I Leica, Cambridge, UK) with a charge coupled devised camera, captured selected images, and transferred them to an Image Analyzer system (Leica, Cambridge, UK) equipped with a morphometry software package (Leica Quantimet Q500 MC, Leica, Cambridge, UK). All sections were examined under several magnifications of light microscopy to identify specific anatomical landmarks. The cervical (mesial and distal) and apical PDL widths were measured, their criteria being the widest zone of PDL in the cervical third and apical third regions of each tooth respectively. Areas showing resorption of root cementum and alveolar bone as well as histologic artifacts were avoided. A minimum of 4 readings per tooth were obtained for each variable and their averages were calculated.

Statistical Analysis

The results of a pilot study with 36 implants, and applying the same protocols were used for sample size calculation. G Power sample size calculating process indicated that 44 samples would be needed to perform a non-parametric statistical comparison. Descriptive statistics were expressed as mean values, and standard deviations. In this study, as there were three independent groups, repeated measures of ANOVA were used, and for significant results pairwise comparison was conducted using Bonferroni adjustment. Values of p< 0.05 were accepted as statistically significant.

RESULTS

Clinical findings

During the 3 months of functional loading, all implants, maxillary premolars and molars in test (Figure 1) and control were clinically healthy and no complications were observed.

Histologic observations

In 4 monkeys, 22 out of 24 posterior maxillary teeth, which were in function for a period of 3 months opposing implant-borne bridgework, were available for analysis as experimental test sample. The remaining 2 teeth were omitted because of histological artifacts. In the control group, 10 out of 12 maxillary posterior teeth opposing natural dentition, served as control. The remaining 2 teeth were also excluded for the same technical reasons.Histologic examination revealed that both the test (IL and DL) and control samples did not show any measurable differences in the PDL morphology in terms of cellularity, collagen fiber density and orientation. Within the same tooth, the cellular components and collagen fibers were differentially distributed in different parts of PDL. The most common pattern was that the cells were evenly distributed in the PDL, and well-formed collagen fibers were observed to run in the apico-occlusal direction (Figure 2). The alveolar bone and cementum surfaces facing the PDL were relatively smooth. A less frequent pattern was observed, with the cells being sparsely distributed in PDL (Figure 3). The collagen fibers formed delicate bundles with intervening loose myxoid connective tissues containing dilated endothelial-lined spaces. These collagen fiber bundles were observed to be running in mesio-distal direction. The alveolar bone and cementum surfaces facing the PDL were relatively irregular (Figure 3). Alveolar bone and root cementum resorption were evident but no osteoclasts were observed. In all samples examined inflammation was absent.

Histomorphometric analysis

Mean and range for cervical and apical PDL width in test and control are summarized in Table 1. Statistically significant differences in mean cervical and apical PDL width among the various teeth in test and control are shown in Figure 4 (p< 0.05).

DISCUSSION

In the present study, the most clinically relevant finding was significant widening of the cervical and apical periodontal

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CLINICAL DENTISTRY AND RESEARCH

ligament of the natural teeth that were opposing functional occlusal forces from an implant-supported superstructure for 3 months. The most plausible explanation for this may be the timing of mandibular tooth extractions and implant installation protocols. In the current study, two surgical implant placement protocols were performed: conventional

2-stage DL implant in 1 quadrant and the one-stage IL implant in the contralateral quadrant. The absence of normal occlusal functional contact for a period of 3 months (IL) and 6 months (DL) may have an effect on its natural dentition and supporting apparatus.16,17 Compagnon & Woda18 in their study on human maxillary first molar in the absence of the opposing antagonist observed that in the first 12 months following the loss of the opposing antagonist, the supra-eruption of maxillary first molar is mainly due to growth of periodontal tissues in an occlusal direction. Later tooth supra-eruption may be due to active tooth eruption.18-20 In the present study, supra-eruption in the experimental test samples opposing the one-stage IL implants was observed to a lesser degree compared to those opposing the conventional 2-stage DL implants.Occlusion is a critical factor for longevity of implants.21 Teeth and implants present in the same arch or even in opposing contact occlusal function, should be evaluated with different perspectives.22-25 The supporting apparatus of the teeth responds to load with an immediate deformation, as well as with a long-term reorganization among PDL, cementum and alveolar bone. The osseointegrated implant does not have this dynamic response, and therefore tends to be the major load carrier when present with natural teeth in the same quadrant.26 The supporting system and stress management for the tooth and implant are different, and this may induce certain changes to the natural dentition opposing implant-borne prosthesis relationship.26 In the natural dentition, the PDL has the capacity to absorb stress or allow for tooth movement, but the bone-implant interface does not have this build-in capacity. Therefore, when implant-supported superstructure is opposing the natural dentition, the higher magnitude and more “rigid” impulse-like forces applied and occlusal stress may be transferred to the natural dentition. In the present study, the increase in periodontal width and root resorption in the experiment test samples may be the result of this effect. Another consideration is that the implant-borne superstructure being without the normal tooth morphology, but simply presented as flat occlusal table. This will again reduce the total surfaces of occlusal contact which in turn might cause the implant-supported prosthesis to introduce a higher occlusal impulse-like force onto their opposing natural dentition.23,24,27-29 Besides increase in mobility, tooth may show clinical signs of increased stress such as enamel wear facets. The implant-borne superstructure, on the other hand, may undergo fatigue fracture and exhibit clinical signs

Figure 2. A-D: Representative sections of a natural tooth from the test sample (IL) showing the periodontal ligament, adjacent root and alveolar bone. A. normal PDL with evenly distributed collagen fibres that appear wavy and relaxed; B. details of Sharpey’s fibres C. apical periodontal ligament; and D. cell rests of Malassez (toluidine blue stain; original magnification C x 40; A x100; B,D x 400).

Figure 3. A-D: Representative sections of a natural tooth from the test sample (DL) showing the periodontal ligament, adjacent root and alveolar bone. A. PDL with sparsely distributed collagen fibres and delicate intervening connective tissues. Note the irregular cementum and alveolar bone surface; B. details of collagen fibres running in an apico-occlusal direction. C. apical periodontal ligament; and D. loose collagen fibres in the apical region (toluidine blue stain; original magnification A x 40; B, C x100; D x 400).

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Implant functIonal loadIng and pdl

including crestal bone changes.30 The loss of natural teeth results in the complete loss of the sensory input that has been provided by the PDL. The tactile receptors of the PDL provide a protective function by the inhibitory reflex, which prevent the over-loading. Sensation differs noticeably for natural teeth and implant, as the latter lacks the periodontal apparatus and its mechanoreceptors essential for the touch sensation. This may have an impact on oral sensory feedback and occlusal awareness in the subjects restored with osseointegrated implants, and this in turn would have an impact on occlusal stress onto the opposing natural dentition. 31-33 It is likely that the natural teeth in this study were also subjected to occlusal overloading from its antagonists (i.e. the implant-supported prosthesis) and this would have led to the PDL tissue responses encountered.There is evidence that under loading, PDL is subjected to both tension and compression which operating in different regions of the ligament concurrently.10,22,34 Histological studies revealed that the principal fibres of the periodontal ligament are relaxed and waxy when the jaws are in resting position but become elongated when force is applied to the tooth.10, 34,35 At times, in the zones of compression the collagen fibres were suspending the tooth and acting as a mattress.8,11 In the current study, morphological evaluation of the principle fibres in the PDL did not demonstrate any measurable differences between test and control samples. A monkey was selected as an experimental model here because its oral anatomy, dental formula and bone metabolism are a good representation of the human oral anatomy, dentition and bone physiology.36,37 A setback

Table 1. Cervical and apical periodontal ligament width in control, immediate loading- and delayed-loading groups

Parameter Animal groups p value

PDL width (mm) Control IL DL

Cervical

Mean 159.5 + 5.81 226.5 + 4.36 223.6 + 5.74 0.000***

Range 116.1 – 174.6 171.6 – 299.6 163.9 – 295.3

Apical

Mean 154.5 + 5.67 221.4 + 12.06 238.8 + 7.89 0.022*

Range 145.8 – 161.8 102.4 – 322.5 127.2 – 358.9

Data are shown as means + standard deviation.Abbreviations: IL, immediate-loading; DL, delayed-loading; PDL, periodontal ligament.* p < 0.05; ***p < 0.001 (one-way ANOVA test).

Figure 4. A-B: Barchart comparing test and control samples A. mean cervical PDL width and B. mean apical PDL width of maxillary second premolars, maxillary first and second molars.

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CLINICAL DENTISTRY AND RESEARCH

here was the small sample size as only a few animals could be used. This was because we took into considerations the implications of the “Rs” principles, namely reduction in the number of animals, reduction of animal suffering and possible replacement of an animal study itself.37 Within the limits, it is plausible to surmise that our results would be comparable and extrapolable to those performed in human. However there are two main limitations in our study. In the current experiment, the biologic impact of implant-borne prosthesis on the opposing dentition was evaluated after 3 months of loading. Hence, dimensional alterations and their outcomes that may occur in PDL following longer time phases of functional occlusal loading remain undetermined. Secondly, current PDL findings were based on natural teeth in functional antagonistic contact with the flat occlusal table of the implant-supported superstructure. It would be interesting to know how the PDL and other related support structures would respond during occlusal loading with a superstructure that recapitulate the occlusal morphology of natural teeth.In conclusion, this study attempted to determine the influence of loading from an implant-supported fixed prosthesis on the PDL of antagonistic natural teeth. Our results suggest that PDL widening observed in the opposing natural teeth might represent a functional response to occlusal forces from the ‘rigidly ankylosed’ implant-supported prosthesis in the opposing arch. CONFLICT OF INTEREST: The authors declare that they have no conflict of interest.

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