J Periodontol • September 2002

Histocompatible Healing of PeriodontalDefects After Application of an InjectableCalcium Phosphate Bone Cement. APreliminary Study in Dogs*Yoshinori Shirakata, Shigeru Oda, Atsuhiro Kinoshita, Shigenari Kikuchi, Hiroaki Tsuchioka, and Isao Ishikawa

1043

Background: A novel injectable, fast setting calcium phos-phate cement (CPC) is currently used in orthopedic therapy forbone fractures. This study evaluated the possibility of applyingthis cement to healing periodontal defects.

Methods: Fenestrations and 3-walled periodontal defects weresurgically created on bilateral first molars and canines in 5 bea-gle dogs. CPC was applied to the defects on one side of themandible. Untreated defects on the contralateral side served ascontrols. CPC was applied to all defects in the maxilla. Twelveweeks after surgery, the animals were sacrificed and decalcifiedand undecalcified specimens were prepared. Periodontal tissuehealing was evaluated histologically and histometrically undera light microscope.

Results: Healing of periodontal tissues in terms of bone andcementum formation was consistently observed in the CPC-applied sites. CPC was partly replaced by new bone. The resid-ual CPC appeared detached from the denuded root surface. Newcementum and periodontal ligament-like tissue were observedbetween the detached CPC and root surface. No unfavorablereaction was noted in the CPC-applied sites. No statistically sig-nificant difference was noted in the experimental or control sitesunder histometric analysis.

Conclusions: Although there were no statistically significantdifferences between the 2 treatment groups, histological obser-vation indicated that CPC seemed to act as a scaffold for boneformation and provided histocompatible healing of periodontaltissues in this study. This cement might be applicable to peri-odontal therapy; however, further investigations are required.J Periodontol 2002;73:1043-1053.

KEY WORDSPeriodontal diseases/therapy; calcium phosphate cement;animal studies; osteogenesis.

* Division of Periodontology, Department of Hard Tissue Engineering, Graduate School,Tokyo Medical and Dental University, Tokyo, Japan.

Periodontal regeneration is the ultimategoal of periodontal therapy. To accom-plish this, various different procedures

including grafting, guided tissue regenera-tion, root surface conditioning, and appli-cation of growth factors are performed.1

Bone grafts are used to enhance periodon-tal regeneration in periodontal defects.1,2

The autograft procedure has disadvantages,not only because of the limited supply ofgraft material often available, but alsobecause the surgery is time-consuming andinvades intra- or extraoral sites.1,2 Al-though some allografts and xenografts havebeen reported to be osteoinductive/osteo-conductive,1-5 their use is still controversialin some parts of the world. Consequently,the use of synthetic materials has been pro-posed for grafting procedures.1,2 Because oftheir handling characteristics, biologicalactivity,6-8 and ready availability in varioussizes, bioceramics have been developedand utilized for grafting.6-19 The most com-monly used ceramic alloplastic materialsare various forms1,2,18 of hydroxyapatite(HA)15,16 and tricalcium phosphate (TCP).

The application of TCP materials in ani-mal and human clinical studies6-14,17,19 hasdemonstrated a resorbable property andconsequent replacement by natural bone.After application, the TCP particles are oftenencapsulated by connective tissue and along junctional epithelial attachment, ratherthan a connective tissue attachment, isformed on the root surface.12-14,17 A simi-

1286_IPC_AAP_553152 8/23/02 1:33 PM Page 1043

Healing After Calcium Phosphate Bone Cement Application Volume 73 • Number 9

lar phenomenon is observed following application ofHA alloplastic material.15,16

A novel injectable cement material consisting of 2different calcium phosphates, calcium carbonate andsodium phosphate, has been developed for bone repairtherapy.18,20 This calcium phosphate cement (CPC) isinjectable, moldable, fast setting, resorbable, and hashigh compressive strength. Orthopedic and materialstudies20-25 have shown that this calcium phosphatecement is non-exothermic and biocompatible in situ.It cures in vivo to form an osteoconductive-carbon-ated apatite, with chemical and physical characteris-tics similar to the mineral phase of bone, which is con-sequently replaced by natural bone. Since it isstructurally similar to bone, the body treats it as such,gradually resorbing it and replacing it with new bone.20

In the present study, we focused on the unique phys-ical properties of this injectable alloplast material andinvestigated the possibility of applying it to periodon-tal defects in dogs.

MATERIALS AND METHODSAnimalsFive healthy male 1-year-old beagle dogs, weighingbetween 12.5 and 13.5 kg, were used in this study.The protocol design and procedures were approvedby the Animal Research Center of Tokyo Medical andDental University.

Grafting MaterialThe grafting material is a powder of monocalciumphosphate monohydrate (Ca[H2PO4]2 ⋅ H2O), α - tri-calcium phosphate (α Ca3 (PO4)2) and calcium car-bonate (CaCO3) mixed with a solution of sodium phos-phate.18,20 The resultant paste can be inserted orinjected into defects or fractures. Once the paste hard-ens (after about 5 minutes), it is chemically identicalto the mineral phase of natural bone.18,20 For thisstudy, the powder and liquid were provided separatelycontained in a sterile capsule. The capsule wasmounted in an amalgam mixer-like apparatus and thecontents blended. The mixed CPC cement† wasinjected into the defects with a specially designed appli-cator.

Surgical ProtocolAll surgical procedures were performed under generaland local anesthesia in sterile conditions. Medetomi-dine hydrochloride‡ was administered intramuscularlyas premedication (0.05 ml/kg). General anesthesiawas achieved using intravenous sodium thiopentalinjection§ (0.005 ml/kg) and spontaneous breathingwas maintained. Additional sodium thiopental wasinjected to maintain anesthesia throughout the surgi-cal procedure. Local anesthesia was performed with 2%lidocaine hydrochloride containing epinephrine� at aconcentration of 1:80,000. Three months prior to the

surgery, all second and third incisors were extracted toprovide adequate space for fabrication of 3-walleddefects. Antibiotics (penicillin G, 200,000 units/day)were administered intramuscularly for 2 days.

Defect Fabrication and TreatmentFenestration defect. Fenestration defects were surgi-cally created on the mesial roots of bilateral mandibu-lar first molars. An intracrevicular incision was madeon the buccal aspect, from distal of the second molarto the mesial of the fourth premolar. Following eleva-tion of the buccal mucoperiosteal flap, a square-shapedfenestration defect, 5 mm wide and 5 mm deep, wasfabricated on the mesial root of the first molar (Fig. 1)using round and fissure burs with sterile saline coolant.Root planing was performed using Gracey curets anda chisel. Cementum was completely removed. CPCwas then applied to fill the defect on the experimen-tal side. Care was taken to avoid overfilling the defectand the material was molded according to the buccalbone morphology (Fig. 2). The contralateral defect,which served as control, was left untreated. Themucoperiosteal flap was repositioned and sutured¶ atthe cemento-enamel junction (CEJ).

Three-walled defect. Three-walled defects were cre-ated on the mesial side of bilateral canines of themandible and maxilla. Intracrevicular incisions weremade along the mesial side of the canine and distalside of the first incisor. A semilunar incision was madefrom the mesial angle of the canine to the distal angleof the first incisor beyond the mucogingival junction.Following elevation of the mucoperiosteal flap, a 3-walled intrabony defect was fabricated on the mesialside of the canine using round and fissure burs withsterile saline coolant (Fig. 3). The defect in themandible was 4 mm wide bucco-lingually and mesio-distally and 8 mm deep, while the defect in the max-illa was 5 mm, 7 mm, and 5 mm, respectively. Rootplaning to completely remove cementum was per-formed using Gracey curets and a chisel.

The 3-walled defects on one side of the mandiblewere completely filled with CPC (Fig. 4), while noth-ing was applied to the contralateral side (control). Inthe maxilla, CPC was applied to all defects. On oneside, 4 holes were made in the hardened CPC extend-ing from the coronal surface to the underlying hostbone using a root canal drill# (Fig. 5) in order toincrease the vascular supply and the surface area ofCPC for faster resorption. The mucoperiosteal flap wasthen repositioned and sutured at the CEJ.

1044

† Norian PDC, Shofu Inc., Kyoto, Japan.‡ Dormitor, Orion Corporation, Espoo, Finland.§ RAVONAL, Tanabe Inc., Osaka, Japan.� Xylocain, Fujisawa Inc., Osaka, Japan.¶ Gore-Tex CV-5 Suture, W.L. Gore and Associates, Inc., Flagstaff, AZ.# Original Beutelrock B2 Reamer 005, Beutelrock Co., Munich, Germany.

1286_IPC_AAP_553152 8/23/02 1:33 PM Page 1044

J Periodontol • September 2002 Shirakata, Oda, Kinoshita, Kikuchi, Tsuchioka, Ishikawa

Postsurgical CareAll dogs received antibiotics (penicillin G, 200,000units) intramuscularly daily for 3 days after CPC appli-cation. The dogs were fed a soft diet** for 2 weeks in

order to reduce potential mechanical interference. Asa plaque control measure, 2% solution of chlorhexi-

1045

Figure 1.The fabricated fenestration defect on the buccal side of the mesialroot of the mandibular first molar.

Figure 2.CPC was applied to fill the fenestration defect.

Figure 3.The fabricated narrow deep 3-walled defect on the mesial side of themandibular canine.

Figure 4.CPC was applied to fill the 3-walled defect.

** DKM, Oriental Yeast Co. Ltd., Tokyo, Japan.

1286_IPC_AAP_553152 8/23/02 1:33 PM Page 1045

Healing After Calcium Phosphate Bone Cement Application Volume 73 • Number 9

dine gluconate†† was used 4 times a week for 12weeks. Sutures were removed after 2 weeks.

Histological ProcessingTwelve weeks after surgery, the animals were sacrificedwith an overdose of sodium thiopental. All of the defectson the experimental and control sites were dissectedwith the surrounding soft and hard tissue. Block sec-tions were fixed in 10% buffered formalin and histolog-ical preparation was performed as described below.

Decalcified specimens. The blocks with 3-walleddefects were demineralized with Plank-Rychlo solution,trimmed, dehydrated,‡‡ and embedded§§ in paraffin.Serial sections of 6 µm thickness were prepared� � inthe mesio-distal plane. Sections were stained withhematoxylin and eosin at intervals of 30 µm.

Non-decalcified specimens. Blocks with fenestra-tions were dehydrated and embedded in polyesterresin.¶¶ The resin blocks were cut bucco-lingually toa thickness of 100 to 150 µm with a low speed dia-mond saw.## Slides were ground and polished to a finalthickness of 35 to 45 µm using a microgrinding sys-tem*** with non-adhesive abrasive discs††† and thenstained with toluidine blue.

Histometric and Statistical AnalysisAll specimens were analyzed histometrically under amicroscope‡‡‡ equipped with a computerized imagesystem.§§§

Fenestration DefectFor histologic measurements, one histological sectionfrom the most central part of the defect was selectedfrom each fenestration defect. The following parame-ters were measured.

Defect height (DH). Distance between the apicaland the coronal ends of the defect.

New cementum formation (NC). Combined linearlengths of newly formed cementum and a cementum-like deposit at coronal and apical ends and within thedefect.

New bone (NB). Combined linear lengths of newly-formed alveolar bone at coronal and apical ends andwithin the defect.

Means and standard deviations for each parameterwere calculated for experimental and control groups.Differences between the experimental and controlgroups were analyzed using Student t-test for pairedobservations (n = 5).

Three-Walled Mandibular DefectFor histologic measurements, 3 histologic sections ofthe most central area of the 3-walled mandibulardefects, approximately 100 µm apart, were selected.The mean value was calculated from measured valuesin these 3 specimens in each site. The following param-eters were measured.

Defect height (DH). Distance between the apicalextent of root planing and the CEJ.

New cementum formation (NC). Distance betweenthe apical extent of root planing and the coronal extentof newly formed cementum and cementum-like depositon the denuded root surface.

New bone (NB). Distance between the apical extentof root planing and the coronal extent of newly formedalveolar bone along the root surface.

Connective tissue attachment (CT). Distancebetween the apical extent of the junctional epitheliumand the apical extent of root planing.

Means and standard deviations for each parameterwere calculated for experimental and control groups.Differences between the experimental group and con-trol group were analyzed using Student t-test for pairedobservations (n = 5).

1046

Figure 5.Four holes were made to penetrate the hardened CPC from thecoronal surface to the bottom of the defect on one side of the maxilla.

†† Hibitane concentrate, Sumitomo Inc., Osaka, Japan.‡‡ Paraffin tissue processor HMP 110, Microm International GmbH.,

Heidelberg, Germany.§§ AP 300-3, Microm International GmbH.�� Rigolac, Nisshin EM Co. Ltd., Tokyo, Japan.¶¶ Rotary Microtome HM 360, Microm International GmbH.## Microcutting Machine BS-300CL, Exact Apparutebau GmbH,

Norderstedt, Germany.*** Microgrinding Machine MG-400CS, Exact Apparutebau GmbH.††† Mecaprex, Presi, Grenoble, France.‡‡‡ Eclipse E800, Nikon Inc., Tokyo, Japan.§§§ Image-pro Plus Version 3.0.1 for Power Mac, Media Cybernetics, L.P.,

Silver Spring, MD.

1286_IPC_AAP_553152 8/23/02 1:33 PM Page 1046

J Periodontol • September 2002 Shirakata, Oda, Kinoshita, Kikuchi, Tsuchioka, Ishikawa

RESULTSClinical ObservationsHealing occurred uneventfully in all 30 sites followingsurgical treatment. No visible adverse reactions, suchas material exposure, infection, or suppuration wereobserved. The initial inflammation noted immediatelyafter surgery was comparable in both control andexperimental sites.

Histologic ObservationsHistologic observations were performed in all sites in5 animals. No acute inflammation or foreign body giantcell reaction was observed in any specimens.

Fenestration defects. The healing pattern varied inthe animals. In experimental sites, periodontal tissuehealing in terms of bone and cementum formation wasconsistently observed. CPC still remained in thedefects. In all animals, the residual CPC appeared to

be detached from the denuded root surface (Fig. 6Aand 6B). New cementum and highly vascular peri-odontal ligament-like tissue were observed in the nar-row space between the detached CPC and root surface.A thin layer of bone-like tissue was observed on thesurface of CPC facing this space. No ankylosis orextensive root resorption was noted (Fig.7). Thick newbone was observed on the buccal surface of CPC inanimals that had a thick host bone plate at the defectsite (Fig. 6A). In animals with a thin host bone plate,only a thin bone-like tissue was found on the buccalsurface of the CPC (Fig. 6B).

In the control sites, the amount of new bone for-mation varied (Fig. 6C and 6D). Two animals that hada thick host bone plate at the defect site exhibitedcomplete closure of the fenestration defect with newbone (Fig. 6C), whereas new bone formation was poorin the animals with a thin host bone plate (Fig. 6D).

1047

Figure 6.A and B: Photomicrographs of a fenestration defect in the CPC-applied group. CPC remained in the defects.The residual CPC was detached from thedenuded root surface.Thick new bone (NB) was observed on the buccal surface of the CPC in some animals that had a thick host bone plate at thedefect site (A). In the other animals with a thin host bone plate, only a thin bone-like tissue (BLT) was found on the buccal surface of the CPC (B).C and D: Photomicrographs of a fenestration defect in the control group.The animals that had a thick host bone plate at the defect site exhibitedcomplete closure of the fenestration defect with new bone (NB) (C). New bone (NB) formation was poor in the animals with a thin host bone plate(D). D: root dentin (bar: 1 mm; original magnification ×0.6; toluidine blue stain).

1286_IPC_AAP_553152 8/23/02 1:33 PM Page 1047

Healing After Calcium Phosphate Bone Cement Application Volume 73 • Number 9

In the defects that completely closed, a thin layer ofnew cementum was formed on the denuded root sur-face and periodontal ligament-like tissue was observedbetween the root surface and new bone (Fig. 8A). Inthe animals with poor bone formation, cementum for-mation on the denuded root surface was limited to theapical and coronal ends of the defect. In these defects,the denuded root surface was predominantly coveredby connective tissue (Fig. 8B). No ankylosis wasobserved.

Three-walled defect in the mandible. In the exper-imental sites, the defect was almost completely filledwith new bone and residual CPC fragments. Newlyformed thick trabecular bone and bone marrow merg-ing with the host bone were observed on the bone sideof the defect. Near the root surface, residual CPC frag-ments were encapsulated by thin newly formed bone-like tissue. Cells infiltrated into the outer portion of the

fragments. Connective tissue was observed betweenCPC fragments with the bone-like tissue on the surface.A continuous layer of new cellular cementum withfunctionally oriented Sharpey’s fibers was observed onthe denuded root surface, extending from the bottomof the defect up to the same height of new bone for-mation (Figs. 9 and 10). Epithelial downgrowth ter-minated slightly above the coronal extent of the newcementum. Neither extensive root resorption nor anky-losis was observed (Fig. 9).

In the control sites, the defect was almost com-pletely filled with thick new trabecular bone and bonemarrow merging with the host bone. The features ofnew cementum and epithelium were comparable tothose observed in the experimental sites; however, thecementum was rather thin. Neither extensive rootresorption nor ankylosis was observed (Fig. 11).

Three-walled defect in the maxilla. In the sites with-out holes in the CPC, the new bone along the root wasalmost at the same level as the intact host bone. CPCappeared as a mass rather than fragments in the

1048

Figure 8.Higher magnification of the framed area in Figure 6C (A) and Figure 6D(B), respectively. A.Thin new cementum (NC) was formed on thedenuded root surface and periodontal ligament-like tissue (PDL) wasobserved between the root surface and the new bone (NB). B. Most ofthe denuded root surface was covered by connective tissue. D: rootdentin (bar: 0.1 mm; original magnification ×12; toluidine blue stain).

Figure 7.Higher magnification of the framed area in Figure 6B. New cementum(NC) and periodontal ligament-like tissue (PDL) were observed in thespace between the detached CPC and root surface.Thin bone-like tissue(BLT) was observed on the surface of the CPC facing this space. D: rootdentin (bar: 0.1 mm; original magnification ×20; toluidine blue stain).

1286_IPC_AAP_553152 8/23/02 1:34 PM Page 1048

J Periodontol • September 2002 Shirakata, Oda, Kinoshita, Kikuchi, Tsuchioka, Ishikawa

defects without holes. New cementum with Sharpey’sfibers and highly vascular periodontal ligament-liketissue were observed along the root. Epithelial down-growth terminated slightly coronal to the coronal extentof the new cementum. Neither extensive root resorp-tion nor ankylosis was observed (Fig. 13).

Histometric AnalysisHistometric analysis was performed in the fenestra-tion and 3-walled intrabony defects of the mandible.Results from histometric analysis are shown in Table1. There was no statistically significant differencebetween any of the parameters in the experimentaland control sites, either in the fenestration or 3-walleddefect models.

1049

defect. CPC mass was almost completely covered bynewly formed bone or bone-like tissue. Connective tis-sue infiltrated into the cracks of the CPC mass. Newcementum and periodontal ligament tissue wereobserved along the root surface. The epithelial down-growth terminated slightly coronal to the coronal extentof the new cementum. There was no extensive rootresorption or ankylosis (Fig. 12).

In sites with holes in CPC, new bone had extendedinto the holes. In the region near the host bone in thedefect, the CPC had resorbed and was replaced bynew bone, which was comprised of bone trabeculaeand bone marrow merging with host bone. The CPCappeared as a mass adjacent to the root surface inthe defect, almost entirely enveloped by newly formedbone or bone-like tissue. Connective tissue infiltratedinto the cracks of the CPC mass, as observed in the

Figure 9.Photomicrograph of a 3-walled mandibular defect in the CPC group.The defect was essentially filled with new bone (NB) and fragments ofresidual CPC. Epithelial downgrowth terminated slightly coronal to thecoronal extent of the new cementum (NC). D: root dentin; *calciumphosphate cement; PDL: periodontal ligament-like tissue; JE: junctionalepithelium (bar: 1 mm; original magnification ×0.6; hematoxylin andeosin stain).

Figure 10.Higher magnification of the area near the root surface in thespecimen shown in Figure 9.The residual CPC fragments weresurrounded by thin newly formed bone-like tissue (BLT). Cells infiltratedinto the outer portion of the fragments. Connective tissue wasobserved between CPC fragments with the bone-like tissue (BLT) onthe surface. New cellular cementum (NC) with functionally orientedSharpey’s fibers was observed on the denuded root surface. D: rootdentin; PDL: periodontal ligament-like tissue (original magnification×60; hematoxylin and eosin stain).

1286_IPC_AAP_553152 8/23/02 1:34 PM Page 1049

Healing After Calcium Phosphate Bone Cement Application Volume 73 • Number 9

DISCUSSIONThe objectives of the present study were to investigatethe possibility of applying an injectable calcium phos-phate cement in surgically created periodontal defectsand to verify the nature and extent of periodontal heal-ing in beagle dogs.

At 12 weeks after surgery, CPC appeared to bedetached from the root surface and periodontal liga-ment-like tissue and cementum had formed betweenCPC and the root surface. These findings suggest thatthe CPC mass had detached from the root surface inthe early phase of healing. The space thus acquiredhad been maintained for periodontal tissue healing.Although not statistically significant, there tended to bea greater amount of new cementum in the fenestrationdefect (P = 0.083) in the experimental sites than inthe control sites. Therefore, CPC may act in a similar

manner to the barrier membrane26,27 in guided tissueregeneration therapy,27-29 which inhibits gingival con-nective tissue migration, maintains the stability of thewound while healing,26,30 and aids the migration ofperiodontal ligament cells.

Isidor et al. observed that new attachment failed toform when the coronal growth of periodontal ligamenttissue was prevented, and suggested that the repopu-lation of a root surface by cells from the periodontalligament was a prerequisite for new attachment for-mation.31 Since the CPC was detached from the rootsurface, it would not have prevented the coronal growthof periodontal ligament tissue in the present study.

A previous human study19 that evaluated the effi-cacy of another calcium phosphate cement in intra-bony periodontal defects reported that all or most of

1050

Figure 11.Photomicrograph of a 3-walled mandibular defect in the control group.The defect was almost completely filled with thick new trabecularbone (NB) and bone marrow continuous with the host bone. D: rootdentin; PDL: periodontal ligament-like tissue; NC: new cementum; JE:junctional epithelium (bar: 1 mm; original magnification ×0.6;hematoxylin and eosin stain).

Figure 12.Photomicrograph of a 3-walled maxillary defect without holes in CPC.The coronal extent of the new bone (NB) along the root was almostat the same level as the intact host bone. CPC appeared to be a massrather than fragments in the defect.The whole surface of the CPCmass was covered by newly formed bone (NB) or bone-like tissue(BLT). Connective tissue infiltrated into the cracks of the CPC mass.New cementum (NC) and periodontal ligament-like tissue (PDL) wereobserved along the root surface. Epithelial downgrowth terminatedslightly coronal to the coronal extent of the new cementum. D: rootdentin; NC: new cementum; JE: junctional epithelium (bar: 1 mm;original magnification ×0.6; hematoxylin and eosin stain).

1286_IPC_AAP_553152 8/23/02 1:34 PM Page 1050

J Periodontol • September 2002 Shirakata, Oda, Kinoshita, Kikuchi, Tsuchioka, Ishikawa

the implanted material exfoliated from the host boneand narrow radiolucent gaps were observed by 1month. Other studies reported that tricalcium phos-phate cement was encapsulated by fibrous connectivetissue.10,13,14 In our study, the osteoconductive prop-erty of the CPC was histologically revealed by the factthat the new bone merged with the host bone andresidual CPC at the region near the host bone in bothfenestration and 3-walled defects. Furthermore, CPCmight have served as a scaffold for new bone forma-tion because the residual CPC mass and fragmentswere almost completely covered by new bone or bone-like tissue. The potential for replacement with naturalbone has been reported originally in skeletal repair.20

Although CPC was partly resorbed and replaced bynew bone, substantial amounts of CPC were still notedin the defects 12 weeks after application. There wasno exposure of CPC to the oral cavity and no unfa-vorable inflammatory reactions were observed. Fasterresorption of the material would be desirable to avoidthe risk of any infection of the residual material dur-ing periodontal healing.

Histological results from the larger 3-walled defectsin the maxilla suggest that the holes in the CPC wouldhasten the resorption of the CPC mass by increasingthe surface area of the material and by allowingincreased vascular supply. It has been proposed thatperiodontal regeneration requires adequate vascularsupply to receive progenitor cells and signaling factorsindispensable for the healing process.32-34

CPC is moldable at application, stiff during the heal-ing period, osteoconductive, bioabsorbable, and pro-vides space for periodontal ligament tissue formation.Considering the unique properties of this injectablecalcium phosphate cement, it could also be utilized inother situations, such as bone filling extraction sock-ets,35 ridge augmentation, and sinus floor augmenta-tion. In addition, it might prove to be a suitable car-rier of various growth factors for tissue engineering.

In order to observe the periodontal tissue responseto the CPC material without the influence of the epithe-lial downgrowth or infection risk,36 a fenestration-typedefect36-39 was employed in this study. Narrow-deep 3-walled defects are considered to heal with high pre-dictability.27,40,41 The considerable healing of the peri-odontal tissues achieved in the control sites of bothfenestration and 3-walled mandibular defects may beattributed to the better wound stability, uninfected sur-gically created environment of the defects, and the exis-

1051

Figure 13.Photomicrograph of a 3-walled maxillary defect with holes in the CPC.New bone (NB) formed in the holes. In the region near the host bonein the defect, CPC had been resorbed and replaced by NB, which wascomprised of bone trabeculae and bone marrow continuous with thehost bone.The CPC appeared as a mass in the other region near theroot surface in the defect, and almost the entire surface of the CPCmass was covered by newly formed bone or bone-like tissue (BLT).Connective tissue infiltrated into the cracks of the CPC mass. Newcementum (NC) with Sharpey’s fibers and periodontal ligament-liketissue (PDL) were observed along the root. Epithelial downgrowthterminated slightly coronal to the coronal extent of the newcementum. D: root dentin; JE: junctional epithelium (bar: 1 mm; originalmagnification ×0.6; hematoxylin and eosin stain).

Table 1.

Histometric Analyses of Periodontal TissueFormation in Fenestration Defects and inMandibular 3-Walled Defects (mean � SDin mm; Student t test for pairedobservations)

CPC Control P Value

FenestrationDefect height 5.06 ± 0.26 4.84 ± 0.46 0.51New bone 2.98 ± 1.26 3.33 ± 1.74 0.53New cementum 2.80 ± 0.90 1.17 ± 1.83 0.083

3-wall defectDefect height 6.68 ± 0.91 7.54 ± 0.83 0.12New bone 5.80 ± 1.16 5.79 ± 1.48 0.99New cementum 5.88 ± 0.90 6.04 ± 1.25 0.82Connective tissue 6.26 ± 1.04 6.70 ± 1.06 0.54attachment

1286_IPC_AAP_553152 8/23/02 1:34 PM Page 1051

Healing After Calcium Phosphate Bone Cement Application Volume 73 • Number 9

tence of intact periodontal ligament tissue surroundingthe defect area. Histometric analysis revealed no sta-tistically significant difference between the 2 treatmentgroups. Our results were obtained by acute wound heal-ing in dogs, thus caution must be taken against extrap-olation of the results to chronic defects in humans. Inorder to verify the efficacy of the material, further stud-ies using critical size or chronically infected defects ina larger number of animals are necessary.

ACKNOWLEDGMENTSThe authors thank our colleagues Drs. Akihiko Murata,Kanta Ito, Noriko Kawakatsu, and Daiji Furuse for theirassistance. This study was also supported by ShoufuInc., Kyoto, Japan, which provided the Norian PDC.

REFERENCES1. Karring T, Lindhe J, Cortellini P. Regenerative periodontal

therapy. In: Lindhe J, Karring T, Lang NP, eds. ClinicalPeriodontology and Implant Dentistry. Copenhagen:Munksgaard; 1998:597-638.

2. Brunsvold MA, Mellonig JT. Bone grafts and periodon-tal regeneration. Periodontol 2000 1993;1:80-91.

3. Mellonig JT, Bowers GM, Cotton WR. Comparison ofbone graft materials. Part II. New bone formation withautografts and allografts: A histological evaluation. JPeriodontol 1981;52:297-302.

4. Harakas NK. Demineralized bone-matrix-induced osteo-genesis. Clin Orthop 1984;188:239-251.

5. Gross BD, Nevins A, Laporta R. Bone-induction poten-tial of mineralized collagen gel xenografts. Oral SurgOral Med Oral Pathol 1980;49:21-26.

6. Hentrich RL, Graves CA, Stein HG, Bajpai PK. An eval-uation of inert and resorbable ceramics for future clini-cal orthopedic applications. J Biomed Mater Res 1971;5:25-51.

7. Strub JR, Gaberthüel TW, Firestone AR. Comparison oftricalcium phosphate and frozen allogenic bone implantsin man. J Periodontol 1979;50:624-629.

8. Snyder AJ, Levin MP, Cutright DE. Alloplastic implantsof tricalcium phosphate ceramic in human periodontalosseous defects. J Periodontol 1984;55:273-277.

9. Hashimoto-Uoshima M, Ishikawa I, Kinoshita A, WengHT, Oda S. Clinical and histologic observation of replace-ment of biphasic calcium phosphate by bone tissue inmonkeys. Int J Periodontics Restorative Dent 1995;15:205-213.

10. Baldock WT, Hutchens LH, McFall WT, Simpson DM.An evaluation of tricalcium phosphate implant in humanperiodontal osseous defects of two patients. J Periodontol1985;56:1-7.

11. Bowers GM, Vargo JW, Levy B, Emerson JR, BergquistJJ. Histologic observations following the placement oftricalcium phosphate implants in human intrabonydefects. J Periodontol 1986;57:286-287.

12. Caton J, Nyman S, Zander H. Histometric evaluation ofperiodontal surgery. II. Connective tissue attachment lev-els after four regenerative procedures. J Clin Periodon-tol 1980;7:224-231.

13. Stahl SS, Froum S. Histological evaluation of humanintraosseous healing responses to the placement of tri-calcium phosphate ceramic implants. I. Three to eightmonths. J Periodontol 1986;57:211-217.

14. Froum S, Stahl SS. Human intraosseous healing re-

sponses to the placement of tricalcium phosphateceramic implants. II. 13 to 18 months. J Periodontol1987;58:103-109.

15. Froum SJ, Kushner L, Scopp IW, Stahl SS. Human clin-ical and histologic responses to durapatite implants inintraosseous lesions. J Periodontol 1982;53:719-725.

16. Ganeles J, Listgarten MA, Evian CI. Ultrastructure ofdurapatite-periodontal tissue interface in human intra-bony defects. J Periodontol 1986;57:133-140.

17. Cameron HU, Macnab I, Pilliar RM. Evaluation ofbiodegradable ceramic. J Biomed Mater Res 1977;11:179-186.

18. Hollinger JO, Brekke J, Gruskin E, Lee D. Role of bonesubstitutes. Clin Orthop 1996;324:55-65.

19. Brown GD, Mealey BL, Nummikoski PV, Bifano SL, Wal-drop TC. Hydroxyapatite cement implant for regenera-tion of periodontal osseous defects in humans. J Peri-odontol 1988;69:146-157.

20. Constantz BR, Ison IC, Fulmer MT, et al. Skeletal repairby in situ formation of the mineral phase of bone. Science1995;267:1796-1799.

21. Jupiter JB, Winters S, Sigman S, et al. Repair of fivedistal radius fractures with an investigational cancellousbone cement: A preliminary report. J Orthop Trauma1997;11:110-116.

22. Cohen MS, Whitman K. Calcium phosphate bonecement—the Norian skeletal repair system in orthope-dic surgery. AORN J 1997;65:958-962.

23. Goodman SB, Bauer TW, Carter D, et al. Norian SRScement augmentation in hip fracture treatment. Labo-ratory and initial clinical results. Clin Orthop 1998;348:42-50.

24. Constantz BR, Barr BM, Ison IC, et al. Histological, chem-ical, and crystallographic analysis of four calcium phos-phate cements in different rabbit osseous sites. J BiomedMater Res 1998;43:451-461.

25. Elder S, Frankenburg E, Goulet J, Yetkinler D, Poser R,Goldstein S. Biomechnical evaluation of calcium phos-phate cement-augmented fixation of unstableintertrochanteric fractures. J Orthop Trauma 2000;14:386-393.

26. Haney JM, Nilveus RE, McMillan PJ, Wikesjö UM. Peri-odontal repair in dogs: Expanded polytetrafluoroethyl-ene barrier membranes support wound stabilization andenhance bone regeneration. J Periodontol 1993;64:883-890.

27. Kim CK, Kim HY, Chai JK, et al. Effect of a calciumsulfate implant with calcium sulfate barrier on periodontalhealing in 3-wall intrabony defects in dogs. J Periodon-tol 1998;69:982-988.

28. Melcher AH. On the repair potential of periodontal tis-sue. J Periodontol 1976;47:256-260.

29. Karring T, Nyman S, Gottlow J, Laurel L. Developmentof the biological concept of guided tissue regenera-tion—animal and human studies. Periodontol 2000 1993;1:26-35.

30. Wikesjö UM, Nilveus R. Periodontal repair in dogs: Effectof wound stabilization on healing. J Periodontol 1990;61:719-724.

31. Isidor F, Karring T, Nyman S, Lindhe J. The significanceof coronal growth of periodontal ligament tissue for newattachment formation. J Clin Periodontol 1986;13:145-150.

32. Alberius P, Gordh M, Lindberg L, Johnell O. Effect ofcortical perforations of both graft and host bed on onlayincorporation to the rat skull. Eur J Oral Sci 1996;104:554-561.

1052

1286_IPC_AAP_553152 8/23/02 1:34 PM Page 1052

J Periodontol • September 2002 Shirakata, Oda, Kinoshita, Kikuchi, Tsuchioka, Ishikawa

33. Majzoub Z, Berengo M, Giardino R, Aldini NN, CordioliG. Role of intramarrow penetration in osseous repair: Apilot study in the rabbit calvaria. J Periodontol 1999;70:1501-1510.

34. Lynch SE. The role of growth factors in periodontal repairand regeneration. In: Polson A, ed. Periodontal Regen-eration: Current Status and Directions. Chicago: Quin-tessence Publishing Co, Inc; 1994:179-198.

35. Gauthier O, Boix D, Grimandi G, et al. A new injectablecalcium phosphate biomaterial for immediate bone fill-ing of extraction sockets: A preliminary study in dogs.J Periodontol 1999;70:375-383.

36. Bye FL, Krause ME, Regezi JA, Caffesse RG. Histologicevaluation of periodontal implants in a biologically“closed” model. J Periodontol 1987;58:110-114.

37. Nyman S, Karring T. Regeneration of surgically removedbuccal alveolar bone in dogs. J Periodont Res 1979;14:86-92.

38. Nyman S, Gottlow J, Karring T, Lindhe J. The regener-ative potential of the periodontal ligament. An experi-mental study in the monkey. J Clin Periodontol 1982;9:257-265.

39. Caplanis N, Lee MB, Zimmerman GJ, Selvig KA, WikesjöUM. Effect of allogeneic freeze-dried demineralized bonematrix on regeneration of alveolar bone and periodontalattachment in dogs. J Clin Periodontol 1998;25:801-806.

40. Ellegaard B, Löe H. New attachment of periodontal tis-sue after treatment of intrabony lesions. J Periodontol1971;42:648-652.

41. Selvig KA, Kersten BG, Wikesjö UM. Surgical treatmentof intrabony periodontal defects using expanded poly-tetrafluoroethylene barrier membranes. Influence ofdefect configulation on healing response. J Periodontol1993;64:730-733.

Correspondence: Dr. Yoshinori Shirakata, Division of Peri-odontology, Department of Hard Tissue Engineering, Grad-uate School, Tokyo Medical and Dental University, 1-5-45Yushima, Bunkyo-ku, Tokyo 113-8549, Japan. E-mail:yoshi.peri@tmd.ac.jp.

Accepted for publication April 5, 2002.

1053

1286_IPC_AAP_553152 8/23/02 1:34 PM Page 1053