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Objectives: To evaluate the effect of immortalized hypertrophic chondrocytes extracellular matrix (HCM) with or without the use of guided bone regeneration (GBR) on the healing of critical-size calvarial defects., Material and Methods: In 42 rats, 5 mm critical-size calvarial defects were surgically created. The animals were randomly allocated to six groups of seven rats each: Group A1: one defect was left untreated (control), while the contralateral defect was covered by a double non-resorbable membrane (GBR). Group B1: one defect was filled with calcium phosphate cement (CP), while the contralateral defect was treated with GBR and CP. Group C1: one defect was filled with a mixture of CP and HCM, while the contralateral defect was treated with GBR and CP+HCM. The healing period for all three groups was 30 days. The remaining three groups were treated in a similar manner but the healing period was 60 days. Five animals from each group were evaluated by maceration and two animals were analysed histologically., Results: At 30 days, all the control-treated defects did not present complete closure. When GBR was applied alone or combined with CP, 3/5 and 5/5 defects, respectively, presented complete closure. At 60 days, one defect from the control group presented complete closure. All the defects treated with GBR alone presented complete closure, whereas the combined use of GBR with CP or CP+HCM resulted in 4/5 and 3/5 defects with complete closure, respectively. The only treatment modality that did not present any specimen with defect closure at both 30 and 60 days was the combination of CP+HCM. The histological analysis indicated that when GBR was not used alone, the healing consisted of an amorphous acellular structure and loose granulation tissue, which, even though clinically resembled hard tissue, did not demonstrate the histological characteristics of bone., Conclusion: The predictability of bone formation in critical-size defects depends mainly on the presence or absence of barrier membranes. The combined use of GBR with calcium phosphate alone or in combination with immortalized human HCM did not enhance the potential for osseous healing provided by the GBR procedure., (© 2011 John Wiley & Sons A/S.)

Objectives: To evaluate the effect of guided bone regeneration (GBR) in combination with or without deproteinized bovine bone mineral (DBBM) and/or an enamel matrix derivative (EMD) on the healing of critical-size calvarial defects., Material and Methods: Forty rats were used. In all animals, a standardized critical-size calvarial defect was created surgically. The animals were randomly allocated into 4 groups of 10 animals each. Group A: One calvarial defect was left untreated, while the galeal and the cerebral aspect of the contralateral defect were covered with a bioresorbable membrane (GBR). Group B: One calvarial defect was filled with EMD, while the contralateral defect was treated with GBR and EMD. Group C: One defect was filled with DBBM, while the contralateral defect was treated with combination of GBR and DBBM. Group D: One defect was filled with DBBM combined with EMD, while the contralateral defect was treated with combination of GBR, DBBM and EMD. The healing period was 4 months. Five specimens from each group were macerated and the length, the width and the vertical dimension (thickness) of the remaining defect were evaluated by a stereomicroscope. The remaining specimens in each group were analyzed histologically., Results: The defects of the macerated specimens that were left untreated or were treated only by EMD, DBBM and combination of EMD and DBBM did not present predictably complete healing of the defects. All the defects where GBR was applied alone or combined with DBBM and/or EMD presented always complete healing (P<0.05). The combined use of GBR with EMD and/or DBBM did not offer any significant advantage above GBR alone in terms of healing of the length and the width of the defect. However, the vertical dimension of the defect was significantly higher (P<0.05) in the GBR-treated specimens of Groups C and D. The histological analysis supported these findings., Conclusion: The predictability of bone formation in critical-size defects depends mainly on the presence or absence of barrier membranes (GBR). The combined use with deproteinized bovine bone mineral and/or enamel matrix proteins did not significantly enhance the potential for complete healing provided by the GBR procedure.

Objectives: The aim of the study was to examine whether bone produced by guided tissue regeneration (GTR) in combination with Bio-Oss or Biogran is stable on a long-term basis., Material and Methods: Fifty-four, 3-month-old Wistar rats were divided into three groups and rigid, hemispherical, teflon capsules were placed with their open part facing the lateral surface of the exposed mandibular ramus (one capsule per animal). In the first group, the capsules were loosely packed with a standardized quantity of Bio-Oss, in the second group with Biogran, and in the last group were left empty. After 1 year of healing, the capsules were removed. Six animals from each of the 3 experimental groups were killed immediately after capsule removal (baseline), or 3 or 6 months after re-entry. The volume of (1) newly formed bone, (2) remaining biomaterial particles, and (3) soft connective tissue in the space originally created by the capsule was estimated by a point-counting technique in three to four histological sections, taken by uniformly random sampling., Results: While considerable bone formation had occurred in the empty control capsules, only limited bone formation was observed in the two test groups. The major portion of the space originally created by the capsules in the test groups was occupied by biomaterial particles embedded in connective tissue. At baseline, the mean volume of newly formed bone occupied 23% of the original capsule space in the animals grafted with Bio-Oss, 12.6% in those implanted with Biogran, and 94.1% in those that received empty control capsules. Six months after capsule removal, the corresponding values were 21.5%, 13.2%, and 91.7%, respectively. No statistically significant differences were observed between baseline, and the 3-, and 6-month observation times in terms of bone volume for any of the three treatment groups (p>0.05)., Conclusion: Bone produced by GTR with and without implantation of Bio-Oss, or Biogran, is stable on a long-term basis, but bone formation is obstructed by implantation of these biomaterials.

Aim: To examine in a discriminating capsule model whether denaturation of demineralized bone matrix (DBM) by heating may influence bone formation., Materials and Methods: DBM was produced from the long bones of rats. Half the portion of DBM was denatured by heating in distilled water for 20 min at temperatures between 70 degrees C and 90 degrees C. Prior to the study, the destruction of the osteoinductive properties of the DBM was confirmed in three rats following intramuscular implantation. Thirty, 4-month-old, male albino rats of the Wistar strain were used in the study. Following surgical exposure of the mandibular ramus, a hemispherical Teflon capsule (internal diameter = 5.0 mm) was placed, with its open part facing the lateral aspect of the ramus. On one side (test side), the capsule was loosely packed with denatured DBM, while on the contralateral side, serving as control, the capsule was loosely packed with the same amount of non-denatured DBM. After healing periods of 30, 60, and 120 days, groups of 10 animals were killed and 40-70 microm thick undecalcified sections of the capsules were produced. Three sections from each specimen, representing the mid-portion of the capsule, were subjected to histological analysis and computer-assisted planimetric measurements., Results: Increasing amounts of newly formed bone were observed in both test and control capsules during the experimental period. At 4 months, the new bone formed in the control capsules occupied 46.7% of the cross-sectional area of the capsules, while it was only 19.1% in the test capsules (P<0.05)., Conclusion: Denaturation of DBM by heating significantly reduces bone formation by guided tissue regeneration.

Aim: To evaluate whether bone formation by guided tissue regeneration (GTR) and demineralized bone matrix (DBM) can be enhanced by the use of a cell-permeable Teflon barrier allowing the penetration of undifferentiated mesenchymal cells from the surrounding soft tissues., Material and Methods: DBM was produced from the long bones of rats, and its bone-inductive properties were tested in three rats prior to the study by intramuscular implantation. Thirty, 4-month-old, male albino rats of the Wistar strain were used. Following surgical exposure of the mandibular ramus, a cell-permeable Teflon capsule, loosely packed with DBM, was placed with its opening facing the lateral surface of the ramus (test side). At the contralateral side, serving as control, a non-perforated (cell-occlusive) Teflon capsule, loosely packed with the same amount of DBM, was placed. After healing periods of 30, 60, and 120 days, groups of 10 animals were killed, and 40-70 microm thick undecalcified sections of the capsules were produced., Results: Computer-assisted planimetric measurements on the histological sections disclosed similar amounts of newly formed bone in both test and control capsules. After 4 months, the new bone in the control capsules occupied 45.0% of the cross-sectional area of the capsule, while it was 50.5% in the test capsules. This difference was not statistically significant (P<0.05)., Conclusion: Similar amounts of bone formed in cell-permeable and cell-occlusive capsules grafted with DBM, suggesting that invasion of undifferentiated mesenchymal cells from the surrounding soft tissues into the barrier-protected area is unnecessary for bone formation with GTR.

Objectives: Grafting of deproteinized bovine bone or bioactive glass has been suggested as an adjunct to guided tissue regeneration (GTR) for the treatment of periodontal and peri-implant bone defects but the influence of these materials on bone formation is not clarified. The aim of the study was to examine the long-term influence of deproteinized bovine bone (Bio-Oss) or bioactive glass (Biogran) on bone formation produced by GTR., Material and Methods: Eighteen rats were used. Following incisions along the inferior border of the mandible, muscle-periosteal flaps were raised to expose the mandibular ramus. Rigid, hemispherical, Teflon capsules (6 mm internal diameter and 1 mm peripheral collar) loosely packed with a standardized quantity of either deproteinized bovine bone (test group 1) or bioactive glass (test group 2), or empty capsules (control group) were then placed with their open part facing the lateral surface of the ramus (one capsule per animal). After 1 year, the capsules were removed by a reentry operation, and the animals were sacrificed. Histological specimens of the augmented sites were prepared, and the volumes of (1). newly formed bone, (2). graft particles, and (3). soft connective tissue in the space originally created by the capsule were estimated by a point-counting technique in three to four histological sections, taken by uniformly random sampling., Results: Limited bone formation was observed in the two test groups. The major part of the space originally created by the capsules was occupied by graft particles embedded in connective tissue. The mean volume of newly formed bone occupied only 23% of the total space in the animals grafted with Bio-Oss and 12.6% in those grafted with Biogran. In the control animals, however, 88.2% (p<0.01) of the space was filled with newly formed bone. There were no signs of ongoing bone formation in any of the three experimental groups., Conclusion: It is concluded that grafting of Bio-Oss or Biogran as an adjunct to GTR arrests bone formation.

Objectives: To evaluate in a discriminating "capsule" model whether local application of gentamicin may have an added effect on bone formation produced by Bio-Oss and guide tissue regeneration (GTR)., Material and Methods: Thirty male 3-month-old Wistar rats were used. After elevation of muscle-periosteal flaps, a rigid hemispherical Teflon capsule, loosely packed with 0.025 g of Bio-Oss impregnated with 2 mg/ml gentamicin sulfate (Garamycin), was placed with its open part facing the lateral bone surface of the mandibular ramus (test) in one side of the jaw. A capsule filled only with Bio-Oss (control) was placed on the contralateral side of the jaw. After healing periods of 1, 2 and 4 months, groups of 10 animals were sacrificed and the specimens were processed for histological examination. The volumes of (1) the space created by the capsule, (2) newly formed bone, (3) Bio-Oss particles, (4) loose connective tissue, and (5) acellular space in the capsule were estimated by a point-counting technique in three to four histological sections of each specimen, taken by uniformly random sampling., Results: The histological evaluation showed limited but increasing bone fill in the capsules from 1 to 4 months in both the test and control sides. After 4 months, the newly formed bone occupied 11.9% (CV: 0.39) of the space created by the capsules at the test sides versus 13.2% (CV: 0.41) at the control sides. There was no statistical significant difference between test and control specimens at any observation time (p>0.05)., Conclusion: It is concluded that local application of gentamicin has no added effect on bone formation when combined with Bio-Oss and GTR.

Aim: To evaluate in a discriminating capsule model whether bone formation by guided tissue regeneration (GTR) may be influenced by concomitant implantation of demineralized bone matrix (DBM)., Materials and Methods: Thirty 4-month-old male albino rats of the Wistar strain were used in the study. Following surgical exposure of the mandibular ramus, a hemispherical, Teflon capsule (5.0 mm in diameter), loosely packed with a standardized amount of DBM, was placed with its open part facing the lateral bone surface of the ramus. At the contralateral side, an empty capsule was placed, serving as control. After healing periods of 15, 30, and 120 days, groups of 10 animals were sacrificed and 40-70 microm thick undecalcified sections of the capsules were produced. In the sections, the cross-sectional areas of (1) the space created by the capsule, (2) newly formed bone, (3) DBM particles, (4) loose connective tissue as well as the (5) height of the capsules, and (6) that of the newly formed bone were measured., Results: Increasing bone fill was observed in both test and control sites from 30 to 120 days. After 30 days of healing, the mean amount of bone was approx. 3% of the cross-sectional area of the capsules at the test sites while it was 8% in the control sites (p<0.05). However, no statistically significant differences were observed between the test (46%) and control (64%) sites after 120 days regarding any of the measured parameters (p>0.05). The newly formed bone in the DBM group at 120 days, on the other hand, appeared more dense than that in the control capsules., Conclusion: DBM used as an adjunct to GTR did not provide any added effect on bone formation but increased the density of the newly formed bone.

It has been reported that local application of bone grafts or synthetic bone substitutes (filler materials) may favour bone formation when used in combination with guided tissue regeneration (GTR). Therefore, the aim of the present investigation was to evaluate the effect of application of chitin beads (a bioabsorbable natural polymer) as a bone substitute in bone formation by GTR. The experiment was carried out in 25 rats. The mandibular ramus was exposed on one side after elevation of a muscle-periosteal flap, and a teflon capsule filled with chitin beads (2.0 mm in diameter) was placed with its opening facing the lateral aspect of the ramus. On the contralateral side of the jaw, serving as control, an empty teflon capsule was placed in the same manner. Groups of 5 animals were sacrificed at 7, 15, 30, 60 and 120 days following capsule placement. Histological analysis demonstrated that the amount of newly formed bone was similar in both experimental and control specimens, amounting to approximately 3% of the central/largest, cross-sectional area created by the capsule at 15 days, and to approximately 9% of this area at 30 days following capsule placement. At 60 and 120 days, however, the amount of newly formed bone observed in the control specimens was twice as large as that observed in the test specimens, amounting to approximately 31% of the cross-sectional area created by the capsule at 60 days, and to approximately 45% at 120 days. It is concluded that, although chitin beads (2.0 mm in diameter) are biocompatible, their presence retards bone formation in the model system used.

Background: Promising results have been reported following treatment of periodontal and peri-implant bone defects with deproteinized bovine bone grafts, but their influence on bone formation has not been clarified., Purpose: The goal of this study was to examine whether implantation of deproteinized bovine bone (Bio-Oss, Geistlich AG, Wolhusen, Switzerland) influences bone formation when used as an adjunct to guided bone augmentation (GBA)., Materials and Methods: A rigid, hemispherical, Teflon capsule was loosely packed with a standardized quantity of Bio-Oss and placed with its open part facing the lateral surface of the mandibular ramus (test) in 30 rats. At the contralateral side of the jaw, an empty capsule was placed (control). Groups of 10 animals were sacrificed after 1, 2, and 4 months. The volumes of the space created by the capsule and of the (1) newly formed bone, (2) remaining Bio-Oss particles, (3) soft connective tissue, and (4) acellular space in the capsule were estimated by a point-counting technique in three or four histologic sections, taken by uniformly random sampling., Results: Bone formation at 1 month was limited in both tests and controls. After 2 months, the mean volume of the newly formed bone occupied 9.0% of the space created by the capsule in the test specimens compared with 23.8% in the control specimens (p < .01). After 4 months, the respective figures were 11.6% (tests) versus 38.7% (controls) (p < .01)., Conclusion: It can be concluded that Bio-Oss, used as an adjunct to GBA, interferes with bone formation.

The aim of this study was to investigate the effect of bone grafting, suture transplantation, and guided tissue regeneration (GTR) treatment on healing of craniectomy defects involving the sagittal cranial suture and on the growth of the cranial vault. Fifty 4-week-old rats were included in the study. A 5.0 mm wide trephine defect was created with its midline corresponding to the sagittal cranial suture between the coronal and occipital cranial vault sutures. The animals were randomly allocated to five groups of 10 animals. Group A: The cranial defect was left untreated. Group B: An occipital bone graft was placed into the cranial defect. Group C: A cranial bone graft including a portion of the frontal suture was placed in the cranial defect. Group D: The cerebral and galeal aspect of the defect were covered with an e-PTFE membrane. Group E: The animals were sham-operated, no defect was created. In all animals, two gutta-percha points were placed demarcating the lateral borders of the parietal bones. Histological analysis at 4 months following surgery showed that the untreated cranial defects healed with fibrous connective tissue. The bone-grafted defects healed partially with bone and connective tissue in the periphery of the bone graft. The healing of suture-grafted defects resembled that of the bone-grafted defects, since the transplanted suture became completely obliterated with bone. The membrane-treated defects healed with bone and a suture-like tissue resembling the sagittal suture of the sham-operated controls. Cephalometric measurements demonstrated that membrane-treated and sham-operated control animals exhibited significantly more (P < 0.05) coronal growth (approximately 1.2 mm) than that of the remaining three groups of animals (approximately 0.7 mm). These findings were supported by the craniometry measurements demonstrating that sham-operated control and membrane-treated specimens presented significantly more cranial width than that of the remaining groups of animals (P < 0.05). It can be concluded that complete osseous healing, creation of a new sagittal suture, and increased cranial growth can be achieved by the treatment of craniectomy defects with the GTR technique.

The aim of the present study was to evaluate whether bone tuberosities produced by GTR on the lateral surface of the mandibular ramus in rats are stable on a long-term basis. Thirty male 6-month-old albino rats of the Wistar strain were used in the study. Tissue flaps were elevated on the lateral aspect of the mandibular ramus. The periosteum was preserved (P+) on one side of the jaw while the bone was denuded (P-) on the other. A rigid, non-porous oval-shaped teflon capsule was placed on both sides with its opening facing the ramus. Six months following surgery, 10 rats were sacrificed and prepared for histology while the remaining 20 rats were subjected to a second operation during which the capsules were removed. Standardized radiographs, taken immediately before and after removal of the capsule and after 3, 6, 9 and 12 months, were subjected to planimetric measurements and subtraction radiography. Ten animals were sacrificed and prepared for histological analysis after 6 months following removal of the capsules and the remaining 10 animals after 12 months. Histology revealed that at 6 months after the placement of the capsules, 17 were completely filled with new bone. The remaining 3 capsules which were displaced exhibited only partial bone fill. The radiographic analysis revealed that after 6 months 98.6 +/- 7.6% (mean +/- SD) in average of the cross-sectional area created by the capsules was filled with new bone. Within 3 months after removal of the capsules a slight resorption of the new bone had occurred, thereby reducing the area of the bone tuberosities by 4 to 8%. No further resorption of the bone tuberosities took place from 3 to 12 months. These observations indicating that new bone produced by GTR is stable on a long-term basis, may question the general belief that non-functional bone will resorb over time.

The aim of this study was to develop an experimental model in rats of an osseous submucous cleft palate and to examine if bone regeneration can be achieved in such defects by the placement of membranes according to the principle of guided tissue regeneration (GTR). An osseous submucous cleft palate defect (5 mm x 2 mm) was created surgically in Wistar rats. Membranes of expanded polytetrafluoroethylene were placed so that they covered both the nasal and the palatal aspect of the defect before the palatal mucoperiosteal flap was repositioned and sutured. Histological analysis after seven weeks of healing showed that significantly more test animals (5/9) healed with newly formed bone and a suture-like tissue in the middle of the defects than controls treated without membranes (0/17). In specimens evaluated macroscopically after 12 weeks, the corresponding rates for the test group were 12/12 compared with 0/13. These observations indicate that an experimental model of an osseous submucous cleft palate can be established in rats, and that complete osseous healing and creation of a midpalatal suture-like tissue in the bony defect is favoured by the placement of membranes according to the principle of GTR.

The aim of this study was to explore the possibility of creating bone tuberosities on the mandibular ramus in rats, using a rigid, occlusive, Teflon capsule. The experiment was carried out in 30 rats. The mandibular ramus was exposed bilaterally. On the test side, the periosteum was left covering the lateral surface of the ramus. On the contralateral side (control), the periosteum was elevated from the lateral surface together with the flap. A hemispherical, Teflon capsule was then placed to face the periosteum or the bone surface with its open part before closure of the wound. The healing periods comprised 7-120 days. Histological analysis demonstrated increasing bone fill in the test specimens from 7 to 60 days, and a limited further increase was observed from 60 to 120 days. In the control specimens, limited bone fill was seen within the first month after surgery, but substantial amounts of new bone were produced from 30-120 days. At 120 days, the mean amount of bone obtained in the test specimens was 56% (range 39%-71%) of the total space created by the capsules, and it was 52% (range 32%-85%) in the controls. The amount of newly formed bone was equivalent to a 5-6 times increase of the original width of the mandibular ramus. These findings indicate that a secluded space created by an occlusive barrier adjacent to existing bone or periosteum may be filled out with bone tissue. This may have a great clinical impact in cranial and maxillofacial surgery.

Abstract Objectives: To evaluate whether Bio-Oss® used as an adjunct to guided tissue regeneration (GTR) improves the healing of 1- or 2-wall intrabony defects as compared with GTR alone, and to examine whether impregnation of Bio-Oss® with gentamicin may have an added effect. Material and methods: Sixty patients, with at least one interproximal intrabony defect with probing pocket depth (PPD) >=7 mm and radiographic evidence of an intrabony component (IC) >=4 mm, were treated at random with either a resorbable membrane (GTR), a resorbable membrane in combination with Bio-Oss® impregnated with saline (DBB-), a resorbable membrane in combination with Bio-Oss® impregnated with gentamicin (DBB+), or with flap surgery (RBF). Results: All treatment modalities resulted in statistically significant clinical improvements after 1 year. Defects treated with GTR alone presented a probing attachment level (PAL) gain of 2.9 mm, a residual PPD (PPD12) of 4.9 mm, a radiographic bone level (RBL) gain of 3.1 mm, and a residual IC (IC12) of 2.7 mm. GTR combined with Bio-Oss® did not improve the healing outcome (PAL gain: 2.5 mm; PPD12: 4.9 mm; RBL gain: 2.8 mm; IC12: 3.3 mm). Impregnation of the Bio-Oss® with gentamicin 2% mg/ml resulted in clinical improvements (PAL gain: 3.8 mm; PPD12: 4.2 mm; RBL gain: 4.7 mm; IC12: 2.1 mm), superior to those of the other treatment modalities, but the difference was not statistically significant. Defects treated with only flap surgery showed the most inferior clinical response (PAL gain: 1.5 mm; PPD12: 5.1 mm; RBL gain: 1.2 mm; IC12: 4.2 mm) of all groups. Conclusion: The results failed to demonstrate an added effect of Bio-Oss® implantation in combination with GTR on the healing of deep interproximal 1- or 2-wall, or combined 1- and 2-wall intrabony defects compared with GTR alone. Local application of gentamicin, on the other hand, improved the treatment outcome but not to an extent that it was... [ABSTRACT FROM AUTHOR]

The aim of the present investigation was to evaluate the bone-forming capacity of the outer (fibrous) and inner (cambium) layer of the mandibular periosteum in skeletally mature rats. The experiment was carried out in 25 rats. The mandibular ramus was exposed on one side (experimental side) after elevation of a muscle-periosteal flap. A teflon capsule was placed with its opening facing the periosteum at the subsurface of the raised muscle-periosteal flap after suturing. In the contralateral side serving as control, the periosteum of the lateral aspect of the mandibular ramus was left intact. This time the teflon capsule was placed with its opening facing the periosteum left behind at the ramus. The histological analysis demonstrated that in all experimental and control specimens, some bone was produced at 7 days after operation. In the experimental specimens, however, both the incidence and the amount of newly-formed bone in the teflon capsules gradually decreased from 7 to 120 days, while increasing in the control capsules placed over the periosteum. At 120 days, the mean amount of new bone produced in the experimental capsules was 3% (range 0-15%) of the total space created by the capsule, while it was 68% (range 41-85%) for the control capsules. The results demonstrated that substantial amounts of bone can be produced predictably by the placement of an occlusive teflon capsule facing mandibular covered with periosteum. Bone produced from the periosteum and without continuity with existing bone becomes resorbed with time. [ABSTRACT FROM AUTHOR]