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Azerbaycan Saytlari

 »  Home  »  Endodontic Articles 3  »  Guided bone regeneration (GBR) using membranes and calcium sulphate after apicectomy: a comparative histomorphometrical study
Guided bone regeneration (GBR) using membranes and calcium sulphate after apicectomy: a comparative histomorphometrical study
Introduction - Materials and methods.

G. Yoshikawa, Y. Murashima, R. Wadachi, N. Sawada & H. Suda
Department of Restorative Sciences, Graduate School, Tokyo Medical and Dental University, Tokyo, Japan.

Osseous defects following apicectomy can be large and complicated in cases with substantial apical lesions, such as combined endodontic-periodontal lesions and through-and-through osseous defects. Andreasen & Rud (1972) divided modes of healing after periapical surgery into two types:

  1. healing by periodontal tissue regeneration, and
  2. healing by fibrous scar tissue.

The most desirable healing after apicectomy is regeneration by bone tissue without scar tissue formation, comparable to healing by periodontal tissue regeneration.
Nyman et al. (1982) first reported the application of guided tissue regeneration (GTR) to periodontal defects in humans. The biological principle of GTR is to exclude dentogingival epithelium and gingival connective tissue proliferation into the wound area adjacent to the root surfaces and, simultaneously, to create a space to give preference to periodontal ligament cells for coronal migration (Nyman 1991). Guided bone regeneration (GBR) is the application of the concept of GTR to osseous defects. Several studies have shown that GBR using expanded polytetraflouroethylene (e-PTFE) membrane was successfully applied to osseous defects in conjunction with apicectomy (Dahlin et al. 1990, Pecora et al. 1995). However, non-resorbable membranes, such as e-PTFE membrane, must be removed some time later following the initial surgery, whilst follow-up surgery is not necessary for resorbable membranes. Several types of resorbable membranes, including the polylactic acidco- glycolic acid (PLGA) membrane and collagen membrane, have been introduced and applied to GTR (Tanner et al. 1988, Minabe et al. 1989, Magnusson et al. 1990, Kon et al. 1991, Caffesse et al. 1994, Gottlow et al. 1994, Cortellini et al. 1996). However, it has not been determined whether the application of resorbable membranes is effective for GBR or not (Simion et al. 1996, Uchin 1996, Ito et al. 1998, Maguire et al. 1998, Zahedi et al. 1998, Bohning et al. 1999).
As an alternative, calcium sulphate has recently been applied to osseous defects during apicectomy as a substitute for membranes (Pecora et al. 1997b). Membranes are difficult to apply in cases with no cervical cortical bone, and in cases with a through-and-through osseous defect, whilst the application of calcium sulphate to GBR is easier.
Although many studies have reported on resorbable and non-resorbable membranes, and calcium sulphate in periodontal treatment (Radentz & Collings 1965, Tanner et al. 1988, Minabe et al. 1989, Magnusson et al. 1990, Kon et al. 1991, Caffesse et al. 1994, Gottlow et al. 1994, Cortellini et al. 1996, Andreana 1998, Kim et al. 1998a,b), only a few studies have been made on bone regeneration in endodontics (Dahlin et al. 1990, Kellert et al. 1994, Pecora et al. 1995, 1997b, Uchin 1996, Maguire et al. 1998).
The purpose of the present study was to evaluate histomorphometrically the effects of resorbable and non-resorbable membranes, and calcium sulphate on bone regeneration in osseous defects in conjunction with apicectomy.

Materials and methods.
Twelve beagle dogs were used in this study with the approval of the Institutional Animal Care and Use Committee of Tokyo Medical and Dental University. The animals were sedated with an intramuscular injection of ketamine hydrochloride (0.3 mg kg –1 , Sankyo Co. Ltd., Tokyo, Japan), and subsequently anaesthetized with sodium-pentobarbital (12.5 mg kg –1 , Tanabe Seiyaku Co. Ltd., Osaka, Japan) given intravenously and added when necessary to maintain deep anaesthesia. In addition, local anaesthesia was performed with an infiltrative injection of 2% lidocaine with 1 : 80 000 epinephrine (Astra Japan Ltd., Osaka, Japan) to control haemorrhage.
Following the general and local anaesthesia, pulpectomies were performed on the mandibular third and fourth premolars on both sides. The root canals were instrumented with K-files (sizes 10–50) and Gates Glidden burs (sizes 2–4), and alternately irrigated with 6% sodium hypochlorite and 3% hydrogen peroxide. After the canals were dried with paper points, they were obturated with gutta-percha and root canal sealer by the lateral condensation method. The access cavities were sealed with glass-ionomer cement.
After the root canal treatments, full mucoperiosteal triangular flaps on both sides were reflected from the distal region of the first molar (vertical releasing incision) to the mesial region of the second premolar (intrasulcular incision). Cortical bone was removed using a trephine bur (4.8 mm in diameter) under water cooling, and the roots of the third and fourth premolars were resected with a fissure bur, also under water cooling. Subsequently, retrograde cavities were made using an ultrasonic tip (Osada Electric Co. Ltd., Tokyo, Japan). Following haemostasis of the osseous defects with 0.1% epinephrine pellets and drying the retrograde cavities with air, Super- EBA® (Harry J. Bosworth, Skokie, IL, USA) was placed as a retrofill. The osseous defects were randomly divided into five groups. In groups A, B, and C the osseous defects were covered with e-PTFE membranes (W. L. Gore & Associates, Inc., Flagstaff, AZ, USA), PLGA membranes (GC Corp., Tokyo, Japan), and collagen membranes (Koken Co. Ltd., Tokyo, Japan), respectively. In group D the defects were filled with medical grade calcium sulphate (Class Implant, Rome, Italy). In group E, they received no further treatment and served as controls. All mucoperiosteal flaps were repositioned and sutured.

Morphometrical parameters
Table 1. Morphometrical parameters.

The dogs were divided into three groups of four animals each, and they were allowed healing periods of 4, 8, and 16 weeks after surgery, respectively. During those periods, bone labelling with fluorescent dyes was completed as follows: tetracycline (20 mg kg –1 , Sigma, St. Louis, MO, USA) and calcein (8 mg kg –1 , Fluka, Buchs, Switzerland) were injected subcutaneously at 15 days and 2 days before sacrifice, respectively. The animals were sacrificed by an overdose of sodium-pentobarbital. Mandibular blocks including the third and fourth premolars were removed, sectioned into small segments containing a single root each, and immediately fixed with 8% formalin and 2.5% glutarardehyde solution for 1 week at 4 C. After fixation, each segment was dehydrated in a series of graded ethanol concentrations and embedded in polyester resin (Nisshin EM, Tokyo, Japan). Undemineralized semiserial sections in the buccolingual direction were obtained from the central portion of the root using a diamond saw (Exact-Apparatebau, Hamburg, Germany) and ground to approximately 50 m in thickness. The specimens were stained with toluidine blue stain or Villanueva’s bone stain. Each section was evaluated both histologically and morphometrically under a light microscope and a fluorescence microscope.

Histological evaluation.
Tissue sections were evaluated histologically with respect to:

  1. presence of the membrane or calcium sulphate at each period;
  2. tissue reaction to the materials used;
  3. inflammation in the osseous defect area;
  4. bone regeneration into the osseous defect;
  5. deposition of cementum-like structure, root resorption, and ankylosis on the resected root surface.

Morphometrical evaluation.
An image analysing system (KS400®, Carl Zeiss Co. Ltd., Göttingen, Germany) was used to measure the morphometric parameters on the image, which was captured by computer software (Adobe Photoshop®, Adobe Systems Inc., San Jose, CA, USA). The images were numbered randomly to blind the experimental groups, and another examiner performed all the morphometrical evaluations to avoid measurement error.

Bone volume is the area marked by asterisks
Figure 1. Diagram of morphometrical measurements. Trapeziform measuring area is surrounded by the broken lines. Bone volume is the area marked by asterisks. Dotted line, concavity of new cortical bone; a, b, c, d, outer and inner edges of original cortical bone; a-b, reference line.

The measured and calculated parameters are shown in Table 1. Four reference points (a, b, c, d in Fig. 1), which were represented by the outer and inner edges of the original cortical bone, clearly created the measuring area representing tissue volume (TV). Bone volume (BV) was defined as the total bone areas within the measuring area. BV/TV was calculated from the percentage of bone volume in the measuring area. The reference line (a–b) was drawn between the outer-top and outer-bottom edges of the original cortical bone (Fig. 1). Concavity of new cortical bone at 16 weeks was obtained by measuring the longest distance from the reference line to the most concave point of new cortical bone (Fig. 1). For statistical analysis, the one-way anova , and two-way anova with Fisher’s posthoc test were performed.