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Table of Contents
CASE REPORT
Year : 2020  |  Volume : 3  |  Issue : 1  |  Page : 27-34

Socket preservation using demineralized tooth graft: A case series report with histological analysis


1 Private Practice, Tijuana, Mexico
2 Department of Dentistry and Oral and Maxillofacial Surgery, School of Medicine, Catholic University of Daegu, Republic of Korea
3 Department of Dentistry and Advanced General Dentistry, Catholic University Hospital of Daegu, Daegu, Republic of Korea
4 Department of Dentistry and Oral and Maxillofacial Surgery, Catholic University Hospital of Daegu, Daegu, Republic of Korea

Date of Submission24-Dec-2019
Date of Acceptance18-Mar-2020
Date of Web Publication16-Apr-2020

Correspondence Address:
Dr. Dong-Seok Sohn
Department of Dentistry and Oral and Maxillofacial Surgery, Catholic University Hospital of Daegu, 3056-6 Daemyung 4-Dong, Nam-Gu, Daegu 705-034
Republic of Korea
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/GFSC.GFSC_16_19

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  Abstract 


Purpose: The aim of the study is to histologically evaluate new bone formation in extraction sockets augmented with autologous demineralized dentin bone (ADDB). Materials and Methods: Teeth were extracted from 19 patients and prepared as ADDB and then grafted in 27 extraction socket sites. The graft was covered with membrane made of concentrated growth factors (membrane) and the wounds were closed without periosteal releasing incision (open-membrane technique). Bone biopsy was performed at each implant site after 4 months of healing. Results: Wound healing was uneventful. Histologically, bone reformation was observed in all the augmented sockets, and ADDB showed favorable integration with newly formed bone. Conclusion: Immediate graft after extraction using ADDB is recommended for socket preservation.

Keywords: Demineralization, dentin bone graft, new bone formation, ridge augmentation, socket preservation


How to cite this article:
Tulio AV, Kang CD, Fabian OA, Kim GE, Kim HG, Sohn DS. Socket preservation using demineralized tooth graft: A case series report with histological analysis. Int J Growth Factors Stem Cells Dent 2020;3:27-34

How to cite this URL:
Tulio AV, Kang CD, Fabian OA, Kim GE, Kim HG, Sohn DS. Socket preservation using demineralized tooth graft: A case series report with histological analysis. Int J Growth Factors Stem Cells Dent [serial online] 2020 [cited 2020 Aug 7];3:27-34. Available from: http://www.cellsindentistry.org/text.asp?2020/3/1/27/282566




  Introduction Top


Following tooth extraction, volumetric changes in the residual ridge is expected. Difficult tooth extraction procedures may also result in additional bone loss due to surgical trauma.[1] According to Botticelli et al.'s research, buccal bone revealed more severe resorption than lingual bone resorption.[2] Preserving the alveolar ridges at the time of the tooth extraction helps minimize difficulties during subsequent implant placement. Bone graft in extraction socket resulted in lesser change of extraction socket.[3] Long-term esthetic success requires vital alveolar bone to support healthy keratinized tissue and dictate soft tissue profile, which follows the underlying bone contour. Various bone substitutes and several surgical techniques have been employed for socket preservation.[4] Autologous bone graft is considered a gold standard because it shows osteoinductivity, osteoconductivity, and osteogenic capacity.[5] However, due to its drawbacks including morbidity of the second surgical site and prolonged surgical time, various bone substitutes have utilized.[6],[7] These bone substitutes are osteoconductive and mainly function as space makers with volume preservation.[8] The protocol of socket preservation is based on guided bone regeneration using osteoconductive bone substitutes and barrier membrane.[9],[10]

Autologous demineralized dentin bone (ADDB) has been introduced for ridge and sinus augmentation as a new alternative to using autogenous bone graft as it is osteoinductive and osteoconductive.[11],[12],[13] An extracted tooth from the patient was grafted as block or particulate form for socket preservation.[14],[15] Resorbable or nonresorbable barrier membrane is commonly used to prevent soft tissue in growth to the graft site. Autologous fibrin membranes, as barrier membrane, have also revealed favorable bone regeneration with ridge augmentation.[16],[17] Sohn et al. also reported successful guided bone generation using a concentrated growth factor (CGF) barrier membrane with primary wound closure.[18] The aim of this retrospective clinical study is to evaluate augmented socket sites using ADDB and CGF barrier membrane according to histological analysis.


  Materials and Methods Top


Patients selection

A total of 27 extraction sockets from 19 patients requiring extraction of hopeless teeth were selected. All extraction socket sites were augmented with ADDB and covered with CGF barrier. In three cases, the mandibular 3rd molar was extracted for socket preservation. The average age of this group was 47.8 years old (ranging 19–74). All patients had medical condition with American Society of Anesthesiologists 1 and 2 were selected (healthy patients or patients with controlled systematic diseases). All patients were informed and documented consent obtained for treatment procedure, histological analysis, and restoration procedure [Table 1].
Table 1: Patient's information

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Preparation of autologous demineralized dentin bone

Prophylactic oral antibiotics (Ampliron Dúo, Siegfried Rhein Co., México) were provided to patients, intake starting 1 day before the procedure and continued for 7 days. Chlorhexidine mouthwash (0.2%) was used by patients. Atraumatic extraction was performed under local anesthesia using 2% lidocaine HLC (1:100,000 epinephrine). Preparation of ADDB using extracted teeth was performed chairside. Old restorations, caries, calculus, and soft tissues attached to the extracted teeth were removed with sharp manual instruments and rotary instrument with copious coolant. A surgical mallet was used to particulate the teeth into sizes between 0.8 mm and 1.0 mm. Accelerated demineralization and sterilization of the grafts were achieved using a vacuum-ultrasonic device (VacuaSonic System, CosmoBioMedicare Co., Seoul, Korea) with fifteen minutes of demineralization using 0.6 N hydrochloride and 10 min of sterilization using sterilization reagents (peracetic acid ethanol solution). The demineralized particulate tooth bone was washed and neutralized using phosphate-buffered saline [Figure 1] and [Figure 2].
Figure 1: Untreated dentin reveals blocked dentinal tubules with mineralized tissue (scanning electron microscope image × 5000)

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Figure 2: Note sufficient exposure of collagen fibers and enlarged dentinal tubule after demineralization treatment. Diverse growth factors in peridential tubule are known to be released though enlarged dentinal tubules (Scanning electron microscope image × 5000)

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Preparation of sticky autologous demineralized dentin bone and concentrated growth factor barrier membrane

Autologous fibrin glue (AFG) was mixed with ADDB for “sticky ADDB” and CGF barrier was prepared according to Sohn et al.'s protocol.[18] 30–50 cc of patient's venous blood was withdrawn from the patient's forearm, one contained in a noncoated Vacutainer to obtain AFG, and two to four glass-coated Vacutainers. These Vacutainers were centrifuged at 2400–2700 rpm using specific centrifuge (Medifuge, Silfradent srl, Sofia, Italy) for 12 min. ADDB was mixed with AFG – collected from the upper level of the noncoated Vacutainer after centrifugation and allowed for polymerization with ADDB for 5–10 min. The sticky ADDB graft does not separate or migrate due to its interlinked fibrin networks and reduces the healing period. The CGF clot is collected from the middle layer after the glass-coated Vacutainer is centrifuged. The dense clot contains polymerized fibrin and is compressed down flat in a metal box (CGF box) with the cover to be used as a membrane barrier.

Surgical procedure

Any soft tissue or inflammatory tissue in the extraction socket was completely removed by manual curettage. Sticky ADDB was grafted in the socket by gentle compaction and 2 CGF membranes were placed over the graft. Wound was closed with chromic catgut 4-0 suture without periosteal releasing incision (open-membrane technique). After 4 months of healing, full-thickness flap was raised to expose the augmented socket site and 3-mm wide trephine drill was used to collect bone for biopsy and to prepare the osteotomy for implant placement. Implant was placed after consecutive drilling and wound closure using catgut 4-0 suture.

Histological analysis

Collected bone for biopsy was placed in a container of formaldehyde and delivered to the oral pathologist for histological studies. The specimens were fixed for 24 h with neutral buffered formalin, then washed with 0.1 M phosphate buffer solution, and decalcified with 10% formic acid for 5 days. The specimen was embedded in paraffin (Paraplast; Oxford, USA) and sliced coronally into serial sections 5 μm thick. The specimens were stained with hematoxylin and eosin (H and E) and Masson's trichrome (MT) stains, and examined under light microscopy.

Case report 1: Socket preservation and sinus lift lateral approach

A 68-year-old female patient presented with a chief complaint of toothache when biting and mobility Grade III on teeth #13, #14, #15. The patient requested for an implant-supported fixed restoration. Three teeth were removed under local anesthesia using 2% lidocaine (1:100,000 epinephrine) with atraumatic extraction technique also using a Piezotome device to maintain the crestal bone. Manual curettage of the extraction site was performed to remove granulation tissue. The extracted teeth were prepared as particulates and decalcified and sterilized according to the manufacturer's instructions (VacuaSonic®; Cosmobiomedicare, Seoul, Korea). Sticky ADDB was then grafted into the socket with CGF membrane coverage. The wound was closed with chromic catgut 4-0 suture without periosteal releasing incision.

Bone graft site was re-entered after 4 months of healing. Full-thickness flap was elevated to expose the augmented socket and favorable ridge augmentation was seen at the site of the extraction socket. A 3-mm wide trephine drill was used as the initial drill for preparing the osteotomy site for implant placement and provided collection of bone for biopsy.

The specimens were stained with hematoxylin and eosin and cut longitudinally to a thickness of 2–3 microns. Then, magnifications of ×100 were made to determine the rate of bone regeneration. Four months after implant placement and healing, an impression was taken, and three-unit zirconia bridge was delivered for the patient [Figure 3],[Figure 4],[Figure 5],[Figure 6],[Figure 7,[Figure 8].
Figure 3: Extraction socket revealed bony defect

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Figure 4: Sticky demineralized tooth bone graft

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Figure 5: Placing the bone inside the sockets

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Figure 6: Concentrated growth factors as a membrane to cover the graft

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Figure 7: Clinical view of the bone 4 months after socket preservation

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Figure 8: The examined specimen presents fragments of dentin and predentin with normal aspect dentinal tubules interspersed with well-vascularized irregular dense fibrous connective tissue. There are few lymphocyte predominant inflammatory cells (H and E, ×100)

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Case report 2: Socket preservation and sinus lift crestal approach with Lisa technique

A 50-year-old male patient presented with root canal treated teeth #26 and #27 requiring removal due to vertical fractures. The patient requested for implant-supported fixed restoration.

Under local anesthesia using 2% lidocaine (1:100,000 epinephrine), atraumatic extraction of both teeth was performed. Sticky ADDB is grafted to extraction socket [Figure 9] and [Figure 10]. Then, CGF membrane was applied to cover the bone graft. The wound was closed with chromic catgut 4-0 suture without periosteal releasing incision. Subsequent to 4 months of healing, full-thickness flap was elevated to expose the augmented socket and 3-mm wide trephine drill was used as 1st osteotomy drill at implant site to take bone biopsy. The specimens were stained with H and E and MT under light microscopy. Then, magnifications of × 40 and × 100 were made to determine the rate of bone regeneration. After 4 months of healing, screwed retain restoration made from a structure of Pekkton and layering composite as final restoration [Figure 11] and [Figure 12].
Figure 9: Sockets after extractions

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Figure 10: Sockets grafted with sticky autologous demineralized dentin bone

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Figure 11: The examined specimen presents mature and vital lamellar bone with vesicular osteocyte gaps surrounded by loose, well-vascularized fibrous connective tissue. (H and E, ×40)

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Figure 12: Present a fragment of dentin surrounded by connective tissue and bone previously described is also observed; together with a chronic, mild inflammatory infiltrate with histiocytes generating resorption of said dentin (H and E, ×40)

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Case report 3: Alveolar three-dimensional ridge reconstruction with autologous tooth block and particulate bone

A 74-year-old patient presented with a fistula on #37. The patient requested for implant-supported fixed restoration. Under local anesthesia using 2% lidocaine (1:100,000 epinephrine), atraumatic extraction was performed. ADDB and demineralzied tooth block was grafted on the defect and CGF membrane was applied to cover the extraction socket on the same day. After 4 months of healing period, the patient returned with exposure of graft, but was unaware of the situation and asymptomatic. A full-thickness flap was elevated to expose the graft and a 3-mm wide trephine drill was used to collect bone before implant placement. The specimen was stained with H and E and MT under light microscopy. Then, magnifications of × 40 and × 100 were made to determine the rate of bone regeneration. After 4 months of healing, the patient came back for open tray impression and zirconia screw-retained restoration was delivered. The cross-sectional image of cone-beam computed tomogram shows stable bone augmentation over the implant [Figure 13], [Figure 14], [Figure 15], [Figure 16].
Figure 13: An extracted lower left first molar before preparing the root part as a block graft

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Figure 14: Immediate placement of prepared demineralized tooth block

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Figure 15: Graft exposure after 4 months due to sharp edge

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Figure 16: New vital bone surrounded by auto tooth graft. Masson's trichrome stain × 100

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  Results Top


Clinical results

From a clinical perspective, all cases healed uneventfully except for one case showing exposure of tooth block graft. Four months after initial graft using ADDB, all socket sites resulted in good bone regeneration for implant placement. Favorable ridge augmentation was achieved for implant engagement with initial stability of more than 25 Ncm.

Histological analysis

ADDB was lightly stained and well differentiated from surrounding tissues in MT stain. Four months after socket preservation procedure, no signs of inflammation were found in the MT stains under light microscopy. Bone reformation was observed in all biopsy samples and newly formed bone showed integration with the ADDB surface. The examined specimen presents fragments of dentin and normal aspect dentinal tubules interspersed with well-vascularized irregular dense fibrous connective tissue. The examined specimens present with mature and vital lamellar bone with vesicular osteocyte lacunae.


  Discussion Top


Significant changes in bone and soft tissue occur in the alveolar bone after extraction.[19],[20] The overall resorption of horizontal alveolar ridge has been reported as 50%.[21] Prevention of alveolar bone resorption after extraction is important to reduce the amount of bone augmentation that may be required with future implant placement. Socket preservation is a surgical procedure aimed to preserve the alveolar bone after tooth extraction to eliminate or minimize the need for bone augmentation during implant rehabilitations.[22] Socket preservation technique includes the use of bone or bone substitute grafts with or without membranes.[23]

Bone graft materials used in the field of dental and oral surgery can be classified into autografts, allografts, alloplasts, and xenografts.[24] Autografts are highly favored due to its osteoinductivity, osteoconductivity, and osteogenicity.[25] However, there are factors to consider such as limited availability or the need of a secondary donor site, resorption of the graft, and high cost.[26] Other graft materials such as bovine bone, allograft, or synthetic bone have been used as an alternative for sinus augmentation. However, the materials are only osteoconductive and adequate as space makers. To overcome the limitations of autografts and allograft or synthetic bone, researches on ADDBs are drawing much attention.[27] Teeth are composed of inorganic substances such as hydroxy apatite and calcium phosphate, and organic substances such as collagen. The composition of teeth is very similar to the bone,[13] especially the dentin which contains 65% inorganic substances, 35% organic substances, and alveolar bone composed of 65% inorganic substances 35% organic substances.[28] Type-1 collagen and various growth factors such as bone morphogenic proteins (BMP) are contained in dentin. BMPs play a role in differentiating perivascular mesenchymal stem cells into cartilage and bone tissue. Various studies have showed the osteoinductivity of BMP.[29],[30],[31] While Type-1 collagen accounts for nearly 90% of the organic substances, other substances such as noncollagenous proteins (NCP) account for the rest. The Type-1 collagen molecule is a triple-helix and has been proved to show biocompatibility and interfere in bone formation at bone graft recipient sites or dental implant sites. NCPs are known to contribute to bone mineralization by facilitating bone resorption and by inducing bone generation process.[32],[33],[34],[35] Seventy percent of the inorganic substances of dentin are hydroxyapatite, which are low-crystalline apatite. Bones are also composed of low-crystalline apatite.[36] In a study by Xu et al., ADDB could be an excellent alternative to autogenous bone graft, since it has osteoinductivity and osteoconductivity and shows bone regeneration.[37] Favorable clinical results were obtained, and optimal osteoconductivity and osteoinductivity of graft was made when teeth were demineralized for 15 min using 0.6 N hydrochloride.[12],[38] Several researches on histology showed that ADDB resorbed with time and resulted in good bone regeneration or new bone formation.[26],[37],[39],[40],[41] ADDB is not only good in providing scaffolds for bone formation but also highly biocompatible, and when compared with other bone substitutes, ADDB has low antigenicity due to its structural similarity with autologous bone. It would also eliminate any concerns of possible risk of cross-infection that may result from using animals or other human sources.[11],[42] Furthermore, ADDB has osteoinductive properties with high potential for bone remodeling and usually does not require additional grafts such as autologous block bone.[43],[44],[45],[46]

The use of autologous bone or bone graft with membrane enhances to the alveolar healing process using biological factors. Choukron's platelet-rich fibrin and Sacco's CGF membrane are recently developed fibrin membrane with platelet aggregation.[47],[48] CGF membranes have successfully substituted allogenic or alloplastic barrier membranes, with excellent clinical results in both soft and hard tissue healing.[48] Using CGF membranes, successful wound healing at bone augmentation site is shown, even without primary suturing. Furthermore, complications due to early exposure of allogenic or alloplastic membranes may be prevented.[49] Mineral tooth block graft showed an increased risk of dehiscence ranging from 12.96% to 34.38%.[50]

To prevent early exposure of tooth block, microperforations and demineralization process are essential to allow blood infiltration through the tooth graft.[46] In addition, sharp edge of tooth block should be smoothened before placing it in the bony defect. The clinical cases demonstrate how ADDB with CGF membrane can be used for socket preservation resulting in favorable ridge augmentation and less ridge resorption.

According to the histological analysis, significant new bone formation was revealed at 4 months of healing. ADDB showed gradual resorption by osteoclastic activity and volume of tooth graft reduced over time during the bone remodeling process. Furthermore, more mature lamella bone was revealed with time and dense and high connection of tooth bone and newly formed bone was evident.


  Conclusion Top


According to the present study, using histological analysis, it can be concluded that ADDB is a bone graft material with favorable clinical and histological outcomes and should be considered as a noble bone grafts for ridge preservation or augmentation.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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    Figures

  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8], [Figure 9], [Figure 10], [Figure 11], [Figure 12], [Figure 13], [Figure 14], [Figure 15], [Figure 16]
 
 
    Tables

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Abstract
Introduction
Materials and Me...
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