The effect of platelet-rich plasma on bone healing around implants placed in bone defects treated with Bio-Oss: a pilot study in the dog tibia Tae-Min You, DDS,a Byung-Ho Choi, DDS, PhD,b Jingxu Li, DDS,c Jae-Hyung Jung, DDS,a Hyeon-Jung Lee, DDS,a Seoung-Ho Lee, DDS, PhD,d and Seung-Mi Jeong, DDS, PhD,e Seoul, South Korea and Wonju, South Korea YONSEI UNIVERSITY, YONSEI UNIVERSITY WONJU COLLEGE OF MEDICINE, AND EWHA WOMANS UNIVERSITY Objective. The aim of this study was to examine the influence of platelet-rich plasma (PRP) used as an adjunct to BioOss for the repair of bone defects adjacent to titanium dental implants. Study design. In 6 mongrel dogs, 12 screw-shaped titanium dental implants were inserted into the osteotomy sites in the dogs’ tibias. Before implantation, a standardized gap (2.0 mm) was created between the implant surface and the surrounding bony walls. The gaps were filled with either Bio-Oss cancellous granules alone or Bio-Oss cancellous granules mixed with PRP. Results. After 4 months, the Bio-Oss–treated defects revealed a significantly higher percentage of bone-implant contact than the defects treated with Bio-Oss and PRP (60.1% vs. 30.8%; P ⬍ .05). Conclusion. The results indicate that when PRP is used as an adjunct to Bio-Oss in the repair of bone defects adjacent to titanium dental implants, PRP may decrease periimplant bone healing. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2007;103:e8-e12) Bone resorption occurring after tooth extraction reduces the height and width of the alveolar crest, hindering the use of dental implants.1,2 The placement of a dental implant immediately after tooth extraction has been recommended as a means to minimize bone loss and shorten the time of the prosthetic treatment.3,4 Immediate implantation into fresh extraction sockets is often associated with a residual bone defect between the implant neck and the residual bone walls. As has been previously reported,5,6 large gaps may jeopardize the success of immediate implant procedures. Such gaps may cause Supported by grant R13-2003-13 from the Medical Science and Engineering Research Program of the Korean Science & Engineering Foundation. a Graduate Student, Department of Oral & Maxillofacial Surgery, College of Dentistry, Yonsei University. b Professor, Department of Oral and Maxillofacial Surgery, College of Dentistry, Yonsei University. c Research Assistant, Department of Dentistry, Yonsei University Wonju College of Medicine. d Associate Professor, Department of Periodontology, Ewha Womans University. e Assistant Professor, Department of Dentistry, Yonsei University Wonju College of Medicine. Received for publication Aug 17, 2006; returned for revision Oct 18, 2006; accepted for publication Nov 22, 2006. 1079-2104/$ - see front matter © 2007 Mosby, Inc. All rights reserved. doi:10.1016/j.tripleo.2006.11.042 e8 cell migration from the connective and epithelial tissue into the gap, possibly preventing osseointegration. Various techniques, including the use of barrier membranes and grafting material, have been proposed for the management of these defects.7 In particular, grafting material mixed with platelet-rich plasma (PRP) has been reported to enhance bone formation8-11 because it contains large numbers of platelets, which in turn release significant quantities of growth factors known to promote wound healing.12,13 However, contradictory results were reported in a recent animal study by Jensen et al.,14 who investigated the effect of PRP on bone regeneration in an allograft. They demonstrated that the addition of PRP into an allograft has no effect on new bone formation in the graft. The inconsistency of these results prompted this study on the effect of PRP on bone regeneration in a xenograft. This study examined the influence of PRP used as an adjunct to Bio-Oss in the repair of bone defects adjacent to titanium dental implants. MATERIAL AND METHODS Six adult female mongrel dogs, each weighing more than 15 kg, were used in this experiment. Approval was obtained from our animal care committee. Platelet-rich plasma was prepared using a technique described previously.15 Briefly, 20 mL of autologous blood withdrawn from each dog was initially centrifuged at 2400 rpm for 10 minutes to separate the PRP OOOOE Volume 103, Number 4 You et al. e9 Kun Dang Co., Seoul, Korea) was administered 1 hour before surgical procedure and once daily for 2 days following the surgical procedure. Sample preparation Animals were killed 4 months after the surgical procedure, and bone blocks with the implants were excised. Resected bone specimens were fixed in 10% buffered formalin and embedded in methylmethacrylate resin. The blocks were cut longitudinally through the middle plane of the implants. Histological sections (40 ␮m) were prepared using a cutting-grinding method and were stained with toluidine blue. Fig. 1. Experimental design. A, The cortical defect filled with Bio-Oss; B, the cortical defect filled with Bio-Oss and platelet-rich plasma (PRP). and platelet-poor plasma (PPP) portions from the red blood cell fraction. The PRP and PPP portions were again centrifuged at 3600 rpm for 15 minutes to separate the PRP from the PPP. Platelet counts were then performed for each dog, yielding a mean PRP platelet count of 1 380 000 (range: 1 010 000 to 2 230 000). The PRP was activated just before application with a 10% calcium chloride solution and 5000 units of bovine thrombin to form a gel. Surgical procedure All surgical procedures were performed under systemic (5 mg/kg ketamine and 2 mg/kg intramuscular xylazine) and local (2% lidocaine with 1:80 000 epinephrine) anesthesia. The bone surface of the tibia was exposed by an incision made on the internal side of the tibia. Before implantation, corticocancellous bone blocks were removed from 2 implant recipient sites by using a trephine bur of 6.0 mm. Bone defects of 6.1 mm (approximately 5.0 mm in length) were created at each site by enlarging the upper aspect of the osteotomy site by using a round burr. Implants 15 mm in length and 4.1 mm in diameter were then placed through both the defect and the lower cortical bone, so that a standardized gap of 2.0 mm was created between the bony walls and the implant neck. In all, 12 implants (Osstem, Seoul, Korea) were inserted, 2 in each tibia. All the implants were stable at the time of insertion. Subsequently, the bone gaps were randomly treated with 1 of the following 2 treatment modalities: (1) grafting with Bio-Oss (Geistlich Biomaterials, Wolhuser, Switzerland) cancellous granules alone or (2) grafting with Bio-Oss cancellous granules mixed with PRP (Fig. 1). All experimental areas were covered with the soft tissue flap after removing the periosteum. Cefazolin (Choing Histomorphometry A morphometric study using an image analysis system (IBAS, Contron, Erching, Germany) was used to quantify the newly formed bone around the implants. The bone-to-implant contact, defined as the length of bone surface border in direct contact with the implant perimeter (⫻100%) starting from the most coronal thread down to the fifth thread, was then calculated. The bone-to-implant contact was measured at the upper cortical and medullary levels. Statistical analysis The Wilcoxon signed rank test was used to calculate statistical differences between the 2 active treatments. P values less than .05 were considered significant. RESULTS No postoperative infections or loose implants were observed during the follow-up period. In the Bio-Oss group, the newly formed bone was in contact with the implant surface. New bone was formed largely at the implant interface in the upper cortical portions (Fig. 2, A, B). In the Bio-Oss and PRP group, a fibrous membrane with fibers parallel to the implant surface was found in contact with the implant surface (Fig. 3, A, B). Compared with the Bio-Oss and PRP group, the BioOss group showed more newly formed trabeculae around the implants in the upper cortical and medullary portions. The mean percentages of direct bone-implant contact in the 2 groups are shown in Table I. The quantitative morphometric analysis showed significantly more bone-implant contact in the Bio-Oss group. The boneto-implant contact was significantly higher (P ⬍ .05) in the Bio-Oss group (60.1 ⫾ 10.0%) than the Bio-Oss and PRP group (30.8 ⫾ 6.3%). DISCUSSION The present study showed that when PRP was used as an adjunct to Bio-Oss in the repair of bone defects e10 You et al. Fig. 2. Section from the Bio-Oss group. A, Original magnification ⫻5. B, Original magnification ⫻100. The newly formed bone contacts the implant surface. adjacent to titanium dental implants, periimplant bone healing was influenced by the concomitant use of PRP. The percentage of bone-implant contact in the defects treated with the biomaterial alone was 60.1% at 4 months, whereas in the defects treated with Bio-Oss mixed with PRP, the percentage was only 30.8%. With respect to the biologic effect of PRP on bone regeneration in a graft, the present results contradict the findings of previous studies.8,10,16 Marx et al.16 found that a combination of PRP and autogenous bone graft can increase the rate of osteogenesis and qualitatively enhance bone formation. Furthermore, Kim et al.8 reported that PRP in combination with bovine cancellous OOOOE April 2007 Fig. 3. Section from the Bio-Oss and PRP group. A, Original magnification ⫻5. B, Original magnification ⫻100. A fibrous membrane (arrows) surrounds the implant surface. bone allografts increased bone formation in calvarial defects in rabbits. Trisi et al.10 reported that PRP, added to a mixture of autogenous bone and Biogran, could improve the new bone formation, with a reduction in the time needed for graft healing and a greater amount of bone formed after only 5 to 6 months. The observations made in the present study are in agreement with findings from previous animal experiments in which there was no effect of PRP on new bone formation in the PRP-treated bone graft.14,17 It is not quite clear why the PRP-treated grafts exhibited decreased bone formation when compared with the non-PRP treated grafts. The explanation may be OOOOE Volume 103, Number 4 You et al. e11 Table I. Bone-to-implant contact in the examined dog tibia Dog number 1 2 3 4 5 6 Mean Bio-Oss group % Bio-Oss ⫹ PRP group % 54.6 72.9 56.9 62.9 68.3 45.2 60.1 ⫾ 10.0 33.1 29.2 28.3 34.7 20.7 38.8 30.8 ⫾ 6.3 PRP, platelet-rich plasma. related to the concentration of PRP within the grafts. Variations in platelet concentration are known to influence bone healing.18 Weibrich et al.18 reported that certain platelet concentrations in PRP may inhibit periimplant bone regeneration. A review of the literature reveals that a variety of PRP volumes have been mixed with bone graft materials. Marx et al.16 used approximately 70 mL of PRP, which was derived from 400 to 450 mL of autologous whole blood, in cancellous marrow graft reconstructions of mandibular continuity defects 5 cm or greater in size. Shanaman et al.19 used 50 mL of PRP in localized alveolar ridge defects. Kassolis et al.20 used 50 to 150 mL of PRP for sinus elevation. One might expect that when a small amount of bone graft is mixed with a large volume of PRP, the bone cells either residing in the adjacent tissues or transferred in the autograft, which are the primary boneregenerating cells, would be exposed to high PRP concentrations. Another factor that could influence the PRP concentrations might be bleeding from the bone. Since bleeding from the gap might dilute the PRP, it is hard to predict the final concentration of platelets in the gap. In addition, as bleeding from the bone can supply the graft with platelets when it is packed in the gap, the bone graft used without PRP will be mixed with some platelets. More basic research into the optimal concentration of PRP within grafts is necessary to adequately capitalize on the ability of platelet growth factors to enhance bone formation in a graft. 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