Andres Stricker Jonathan Fleiner € binger Stefan Stu Henrik Fleiner Daniel Buser Dieter D. Bosshardt

Ridge preservation after ridge expansion with simultaneous guided bone regeneration: a preclinical study

Authors’ affiliations: Andres Stricker, Henrik Fleiner, Department of Oral and Maxillofacial Surgery, University Hospital of Freiburg, Freiburg, Germany Andres Stricker, Jonathan Fleiner, Center of Implantology, Periodontology and 3D Head- and Neck Diagnostics, Konstanz, Germany Jonathan Fleiner, Oral Imaging Center, Faculty of Medicine, Katholieke Universiteit Leuven, Leuven, Belgium Stefan St€ ubinger, Musculoskeletal Research Unit, Equine Hospital, Vetsuisse Faculty ZH, University of Zurich, Zurich, Switzerland Stefan St€ ubinger, Center of Applied Biotechnology and Molecular Medicine (CABMM), Vetsuisse Faculty, University of Zurich, Zurich, Switzerland Daniel Buser, Dieter D. Bosshardt, Department of Oral Surgery and Stomatology, School of Dental Medicine, University of Bern, Bern, Switzerland Dieter D. Bosshardt, Robert K. Schenk Laboratory of Oral Histology, School of Dental Medicine, University of Bern, Bern, Switzerland Dieter D. Bosshardt, Department of Periodontology, School of Dental Medicine, University of Bern, Bern, Switzerland

Key words: bone splitting/ridge expansion, guided bone regeneration, miniature pig, osseoin-

Corresponding author: Dr. Andres Stricker, Department of Oral and Maxillofacial Surgery, Universityhospital of Freiburg, Hugstetter Strasse 55, 79106 Freiburg, Germany Tel.: +49 761 27049400 Fax: +49 761 27048400 e-mail: [email protected]

group compared to the MF group. Furthermore, buccal bone thickness in the GBR group was 0.93,

tegration, ridge preservation Abstract Objective: To evaluate ridge preservation after ridge splitting with simultaneous implant placement and guided bone regeneration (GBR) in a miniature pig model. Material and Methods: In miniature pigs, the mandibular premolars and first molars were extracted together with removal of the interdental and buccal bone. Three months later, ridge splitting and expansion of the buccal plate were performed with simultaneous placement of two titanium implants per quadrant. On the test side, access by a mucoperiosteal flap followed by GBR with a biphasic calcium phosphate and a collagen membrane was performed. On the contralateral control side, a mucosal flap (MF), leaving the periosteum attached to the buccal bone, was elevated. After healing periods of 6 and 12 weeks, eight and four animals, respectively, were sacrificed for histological and histometric evaluation. Results: In the MF group, all 16 implants were osseointegrated, while in the GBR group, one bone fracture occurred, and six of 16 implants were lost. After 6 weeks, significantly higher bone crest levels were found for the GBR group than for the MF group both buccally and lingually (P < 0.001), and buccal bone thickness was greater in the GBR group than in the MF group (P < 0.001 at the implant shoulder [IS]). After 12 weeks, bone was significantly higher in the GBR 4.5, and 5.94 mm at, and 2 and 4 mm apical to the IS, respectively. The corresponding values in the MF group were greatly reduced (0, 0.21, and 2.56 mm). Bone loss on the buccal side compared to the lingual side was significantly greater only in the MF group. Conclusions: In this ridge expansion model in miniature pigs, the buccal bone volume was significantly better preserved with GBR when compared to a mucosal access flap, provided that soft tissue healing occurred complication free.

Date: Accepted 29 January 2015 To cite this article: Stricker A, Fleiner J, St€ ubinger S, Fleiner H, Buser D, Bosshardt DD. Ridge preservation after ridge expansion with simultaneous guided bone regeneration: a preclinical study. Clin. Oral Impl. Res. 00, 2015; 1–9. doi: 10.1111/clr.12574

To achieve long-term optimal functional and esthetic results with dental implants, one major prerequisite is sufficient bone quantity and quality at the recipient site. However, in many situations, there is a bone deficiency (Braut et al. 2011; Chappuis et al. 2013) indicating a procedure that predictably leads to sufficient bone. Guided bone regeneration (GBR), alveolar distraction osteogenesis, and onlay bone block grafting are augmentation techniques used to enhance horizontally the bone volume (Buser et al. 1999, 2002; Cordaro et al. 2002; Donos et al. 2008; Chiapasco et al. 2009). While onlay grafting implies a reconsolidation time of at least 3 months to enable implant placement in a staged procedure (Buser et al. 1996; Chiapasco et al. 1999;

© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd

McAllister & Haghighat 2007), a GBR procedure can be performed with simultaneous implant placement (Wilson & Buser 1994; Zitzmann et al. 2001; Aghaloo & Moy 2007; Jung et al. 2009). Many studies have demonstrated the success of these surgical approaches, but donor site morbidity, infection, and unexpected bone resorption are among the documented drawbacks of these augmentation techniques (Machtei 2001; Chiapasco et al. 2006; Funaki et al. 2009). To enhance the horizontal bone volume in case of a narrow alveolar ridge, the bonesplitting/ridge expansion technique has been introduced by Nentwig in 1986 (Nentwig 1986). Here, the thin alveolar ridge is crestally opened and subsequently split with special

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osteotomes (Simion et al. 1992; Lustmann & Lewinstein 1995), followed by implant insertion into the expanded space between the medial and buccal bone walls (Koo et al. 2008; Funaki et al. 2009). After 4 months of submerged healing, implant uncovering and loading can be performed (Basa et al. 2004). The ridge-splitting procedure offers the advantage of simultaneous implant placement and avoiding bone graft harvesting from secondary donor sites even in situations with severe atrophies of the alveolar crest (Scipioni et al. 1994, 1999; Sethi & Kaus 2000; Ferrigno & Laureti 2005; Blus & Szmukler-Moncler 2006; Chiapasco et al. 2006; Bravi et al. 2007; Elian et al. 2008; Han et al. 2011). Yet, as buccal bone resorption after ridge expansion is a prevalent long-term sequela, prevention of resorption of the mobilized buccal bone is an issue of great clinical importance (Scipioni et al. 1997; Strietzel et al. 1999; Funaki et al. 2009; Bassetti et al. 2013; Beolchini et al. 2015). Some approaches were proposed to reduce bone resorption (Sethi & Kaus 2000; Vercellotti 2000; Coatoam & Mariotti 2003; Basa et al. 2004; Enislidis et al. 2006; Koo et al. 2008), but evidence for their efficacy is still lacking. Moreover, there is still no clear concept concerning the role of the periosteum attached to the buccal bone in the ridge-splitting technique. While in many studies, the surgical site was accessed by a mucoperiosteal fullthickness flap with complete denudation of the expanded buccal bone (Coatoam & Mariotti 2003; Basa et al. 2004; Ferrigno & Laureti 2005; Blus & Szmukler-Moncler 2006; Lai et al. 2007; Santagata et al. 2008; Danza et al. 2009; Demarosi et al. 2009) , other studies proposed a mucosal split flap leaving the periosteum attached to the buccal bone (Scipioni et al. 1999; Chiapasco et al. 2006; Beolchini et al. 2014). While similar implant success rates have been demonstrated for the full-thickness, the partial-thickness, and the osteoperiosteal flaps in a clinical study (Jensen et al. 2009), a preclinical study demonstrated favorable bone preservation for the partial-thickness flap (Stricker et al. 2014b). Ridge splitting in animals histologically demonstrated marginal bone loss (Han et al. 2011), particularly at buccal sites (Scipioni et al. 1997; Funaki et al. 2009; Stricker et al. 2014a). One possibility to reduce this bone loss is a GBR procedure. Except for one very recently published clinical study (Kolerman et al. 2014), there are no data available on ridge preservation after ridge splitting combined with GBR. Therefore, the aim of this study was to evaluate ridge preservation after

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Clin. Oral Impl. Res. 0, 2015 / 1–9

ridge splitting with simultaneous implant placement and GBR in a recently established miniature pig model.

Material and methods Study design

On each side of the mandible in miniature pigs, ridge expansion was performed and two titanium implants were placed in the created bone gap. There were three experimental groups with two healing periods each. A randomization schedule was applied. The two implants installed in the same hemimandible received the same treatment. In the first group, a mucoperiosteal flap (MFP) with denudation of the buccal bone was elevated. In the second group, access was made by a mucosal flap (MF) leaving the periosteum attached to the buccal bone. In the third group, a MFP was raised, and after implant installation, GBR was applied with a synthetic bone substitute and a collagen barrier membrane. The data of the first group, which have recently been published (Stricker et al. 2014b), will not be further discussed here. Animal model and management

Study approval was obtained from the Ethical Committee of the University of LundMalm€ o, Sweden. The study design was in accordance with internationally accepted guidelines for animal trials testing of biomaterials and the related evaluation of their efficacy as stated by Dard (2012). Twelve adult female G€ ottingen miniature pigs (Ellegaard, Special Diet Service, Dalmose, Denmark) with an average body weight of 40 kg were housed in standard cages and were fed on a soft diet for miniature pigs (Special Diet Service, UK). Prior to both surgical procedures, all animals were fasted overnight to prevent vomiting. On the day of surgery, all animals were premedicated with an intramuscular injection of atropine (Atropinum sulfuricum, 0.05 mg/kg IM). All surgical procedures were performed under general anesthesia and aseptic conditions in a dedicated animal surgical clinic (Malm€ o, Sweden). All animals were anesthetized according to the following procedure: 10 ml of ketamine (Ketalar Vet; Pfizer AB, Sollentuna, Sweden, 50 mg/ml) was mixed with 3 ml midazolam (Dormicumâ 5 mg/ml: Roche, Basel, Switzerland). During surgery, 10 ml of ketamine had been injected when needed. All miniature pigs received 10 ml of ketamine every 30 min and, if needed, 1.5 ml of midazolam. An additional local anesthesia (Xylocain Dental adrenalin

20 mg/ml + 12.5 mg/ml; Astra AB, S€ odert€alje, Sweden) was given to reduce the dosage of the systemic anesthetic as well as to reduce the bleeding during surgery and to alleviate pain after surgery. Postsurgical treatment with systemic antibiotics (Streptocillin vet.â; Boehringer Ingelheim, Copenhagen, Denmark) was given for 7 days to avoid infections. Within the first days after surgery, all animals were monitored routinely and further analgesia was given if necessary. The whole study was accompanied and monitored by a veterinarian and researchers with extensive experience performed all surgical procedures. Surgical phase

The two surgical procedures were performed under aseptic conditions in an animal operating theater under general anesthesia. In the first procedure, three mandibular premolars and the first mandibular molar were carefully extracted on each side followed by the removal of the buccal bone wall and the interradicular septa by a round bur. Single-tooth film X-rays were taken to control complete extraction of all remaining fractured roots. After 3 months of healing of the atrophic sites, the second intervention consisted of the ridge-splitting procedure. According to the randomization schedule of the study design, the crestal incision was followed by preparation of a mucoperiosteal full-thickness flap with complete denudation of the buccal bone at the test group, while in the control group, a mucosal split flap leaving the periosteum attached to the buccal wall was reflected. The narrow ridge was cut crestally 30 mm in the mesio-distal direction and 6 mm deep with a piezo instrument (Piezosurgeryâ; Mectron s.p.a., Carasco, Italy) (Fig. 1a). Mesially and distally, a buccal release cut of 6 mm was performed and the mobilized buccal bone plate was displaced in the buccal direction with the help of a special osteotome (Ergoplantâ Aesculap AG, Tuttlingen, Germany). Thereafter, the expanded gap was kept open with retraction inserts (Ergoplant), and two SLActive implants (Straumannâ Bone Level Implant, NC Ø 3.3 mm/8 mm, Institut Straumann, Basel, Switzerland), which achieved primary stability in the deeper portion of the nonfractured gap, were installed per quadrant in such a way that the implant shoulder (IS) was flush with the level of the lingual and buccal bone crest (BC) (Fig. 1b). In the test group, a GBR procedure consisting of a lateral augmentation of the buccal bone plate with granules consisting of an alloplastic biphasic calcium phosphate (BCP) biomaterial

© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd

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(Straumannâ BoneCeramic, 400–700 lm; Institut Straumann) mixed with blood (Fig. 1c) and a barrier membrane consisting of xenogenic collagen (Fig. 1d) (BioGideâ; Geistlich, Wolhusen, Switzerland) was applied. In the control group, no GBR procedure was applied. For tension-free soft tissue closure, periosteal release incisions were applied in the test group (GBR), while mobilization by the mucosal flap was sufficient in the control group (MF). Interruptive suturing of the flaps with a resorbable material (Vicryl 4-0; Ethicon Norderstadt, Germany) was followed by postoperative X-ray control from each side. Terminal procedure

Eight animals were sacrificed 6 weeks after implant installation, and four animals were sacrificed after 12 weeks. The termination was conducted by inducing cardiac arrest with an intracardiac injection of a 20% solution of pentobarbital (Pentobarbitalnatrium; Apoteket AB, Stockholm, Sweden, 60 mg/ ml). Block resections of the implant sites were performed using an oscillating autopsy saw to keep the soft tissue intact. The removed block sections were fixed by immersion in 4% buffered formalin for 2 weeks. Histology

The specimens were left undecalcified and dehydrated in increasing concentrations of ethanol and embedded in methylmethacrylate. Serial sections of ~500 lm in thickness were cut in a bucco-lingual direction using a

low-speed diamond saw with coolant (Varicutâ VC-50; Leco, Munich, Germany). In addition, sagittal sections of the gap region between implants were produced. After mounting the sections onto acrylic glass slabs, they were ground and polished to a final thickness of about 100 lm (KnuthRotor-3; Struers, Rodovre/Copenhagen, Denmark). The sections were stained with toluidine blue and basic fuchsin, and the two most central ground sections per implant and per interimplant region were used for qualitative analysis. Digital photography was performed using a ProgResâ C5 digital camera (Jenoptik Laser; Optik Systeme GmbH, Jena, Germany) connected to a Zeiss Axioplan microscope (Carl Zeiss, G€ ottingen, Germany). Position and osseointegration of the implants, new bone formation, and soft tissue condition were assessed directly in the microscope. Histomorphometry

Histomorphometric analysis (Fig. 2) was applied to evaluate (i) the level of the bone crest, that is, the distance (IS-BC) between the IS and the BC; (ii) the level of the most coronal bone-to-implant contact, that is, the distance (IS-cBIC) between the IS and the coronal-most bone on the implant surface (cBIC); (iii) the thickness of the buccal bone at the IS (W0) and 2 mm (W2) and 4 mm (W4) apical to the IS; (iv) the maximum thickness of the buccal bone(Wmax); and (v) the percentage of bone-to-implant contact (BIC).

(a)

(b)

(c)

(d)

Fig. 1. Intra-operative buccal views illustrating (a) bone splitting of the narrow ridge and (b) implant installation in the control group with mucosal flap. In the test group with guided bone regeneration, (c) the particulate bone substitute was placed buccally and (d) covered with a collagen membrane.

© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd

Statistical analysis

All analyses were performed with the statistical Software R, version 2.15.1 (http://www.Rproject.org, Vienna, Austria, 2008). To analyze the 6-week data, the nonparametric methods of Brunner et al. (2002), in particular the LD-F1 model, were used to determine the impact of the factor group (with classes GBR and MF) and the factor side (with classes lingual and buccal) on IS-BC, IS-cBIC, W0, W2, and W4. All response values were measured on both anterior and posterior position. Statistical analysis was then performed on the average of these two positions. The 12-week data were analyzed descriptively. P-values