Bovine Demineralized Bone Matrix Research Articles

CYTOTOXICITY TEST OF BOVINE DEMINERALIZED BONE MATRIX ON HUMAN MESENCHYMAL STEM CELLS USING THE MTT ASSAY METHOD

The use of bone grafts in Indonesia continues to increase each year. Although Autograft is considered the gold standard in bone grafting, its use is often confronted with various challenges, similar to allograft. To address this issue, Bovine Demineralized Bone Matrix (DBM) can be considered as a substitute for bone grafts with the advantages of unlimited availability and more affordable costs. Currently, the Tissue Bank of Dr. Soetomo Hospital is developing bovine DBM, although there is no research yet on its potential toxicity.This study aims to evaluate whether bovine DBM has cytotoxic effects on human mesenchymal stem cells. In this experimental study, a total of 48 samples were involved, including a control group and two treatment groups (50% and 25%), each consisting of 16 samples. Mesenchymal stem cells were cultured and then treated with the addition of 50% and 25% DBM. Subsequently, cell viability was measured using the MTT Assay method.The collected data were processed by conducting normality and homogeneity tests and then analyzed using comparative tests with an independent t-test. The criteria for declaring cell toxicity were set at a viability of not less than 60% compared to the control group.The results of the MTT assay measurements showed that the mean Optical Density (OD) in the control group was 0.656 ± 0.021 (range 0.620-0.696), while in the treatment groups, it was 0.565 ± 0.022 (range 0.529-0.614) and 0.520 ± 0.022 (range 0.461-0.552), respectively. Statistically, the differences in OD between the control group and both treatment groups (50% and 25%) were significant (p<0.05). The average cell viability in both treatment groups was found to be more than 60%, indicating that Bovine Demineralized Bone Matrix is not toxic to human mesenchymal stem cells.

CAD/CAM Titanium Meshes for GBR: A Case Series with Preliminary Histologic Analysis.

Titanium has been proposed as a mesh material for guided bone regeneration (GBR) since the 1990s. To overcome difficulties in shaping and adapting meshes to the defect, digital techniques were introduced to digitally print meshes capable of fitting the bone perfectly, reproduced through the patient's CT scan. Five patients were included in this case series, and their CBCT data were acquired and sent to the producer of the titanium meshes. 3D regenerative surgery was performed with titanium meshes and a mix of demineralized bovine bone matrix (DBBM) and autogenous bone (1:1 ratio). Radiographic measures were evaluated on paraxial sections of the CBCT through a dedicated software. When possible, regenerated bone samples were obtained at implant insertion. Four out of five regenerated areas healed without local or systemic complications. One mesh was removed after 2 months and 2 weeks due to exposure. The mean vertical bone gain was 4.3 ± 1.5 mm (range: 2.5 to 7 mm). Two histologic samples were obtained. In sample 1, bone tissue area and graft material area were 44.4% and 12.5%, respectively; in sample 2, the same parameters were 15.6% and 16.9%, respectively.

Lateral Sinus Floor Elevation in the Severely Atrophied Maxilla: Concentrated Growth Factors Versus Bone Substitutes. A Controlled Clinical Trial

The present study clinically and radiographically compares the outcome of implants inserted in maxillary sinuses augmented with concentrated growth factors (CGFs) or demineralized bovine bone matrix (DBBM) in a one-stage lateral approach. In 20 patients with a residual bone height of 1 to 4 mm, lateral sinus floor elevation was performed, using CGFs or DBBM as the sole grafting material, with simultaneous implant placement. Outcome variables were implant and prosthesis failures, complications, subjective satisfaction, and radiographic changes in marginal bone level (MBL) 12 months after surgery. The patients were consecutively recruited: 10 to the CGF group and 10 to the DBBM group. No implant failed in either group at 12 months postsurgery, and there were no complications. There was no statistically significant difference in MBL change between the CGF and DBBM groups (difference of -0.3 mm, favoring the CGF group; 95% confidence interval [CI]: -0.8 to 0.2; P = .18). There was no statistically significant difference in satisfaction (difference of 0.2, favoring the CGF group; 95% CI: -0.2 to 0.6; P = .29). Within the limitations of the present study, the lateral sinus floor elevation performed with the use of CGFs as the sole grafting material showed implant survival rates and marginal bone level changes comparable to DBBM grafting.

In vivo regeneration functionalities of experimental organo-biomaterials containing water-soluble nacre extract

BackgroundNovel multifunctional biomaterials were recently designed to allow for an optimized tissue regeneration process. PurposeTo comprehensively assess (photographic, radiographic and histological) the in vivo functionality of demineralized bovine bone matrix (DBM) associated with an experimental marine organic extract (MOE) from nacre in a sheep ectopic grafting model. Materials and methodsSynthesis of MOE was based on mixing powdered nacre (0.05 g, particles average size <0.1 mm) with acetic acid (5 mL, pH 7) under constant stirring for 72 hours (25 °C). Polyethylene tubes (3/animal, n = 4, diameter: 5.0 mm × length: 10.0 mm) from the control (empty) or experimental groups (DBM or DBM + MOE) were then intramuscularly implanted into the lumbar regions of sheep (n = 8, 2-years old, ≈45 kg). Animals were euthanized at 3 and 6 months to allow for the collection of tissue samples. Tissue samples were fixed in formalin 10% (buffered, 7 days) in preparation for photographic, radiographic and histological assessments. Acquired images were then analyzed using digital image analysis software to quantify the amount of neoformed tissues, whereas radiographic and histological analyses were performed to determine radiopacity and classification of tissues deposited inside of the tubes. ResultsPhotographic and radiographic analyses have shown that both pure (unaltered) and MOE-modified DBM were capable of depositing neoformed tissues (at 3 and 6 months), where higher levels of deposition and radiopacity were observed on groups treated with experimental materials. Histological results, however, demonstrated that tissues formed from both unaltered and MOE-modified DBM were only fibrous connective in origin. ConclusionsAs an ectopic grafting in sheep, the experimental organo-biomaterial association applied did not reveal any osteoinductive property but led to a fibrous tissue repair only.

&lt;i&gt;In Vitro&lt;/i&gt; Comparative Study of Osteogenic Differentiation Ability between Adipose and Bone Marrow Mesenchymal Stem Cell Applied to Bovine Demineralized Bone Matrix

Ideal bone graft must possess the desirable trait such as osteoconductive, osteoinductive and osteogenesis. Demineralized Bone Matrix (DBM) provides both osteoconductive and osteoinductive trait. Referring to the tissue engineering principle, the addition of mesenchymal stem cell would add the osteogenic trait to this procedure. The design of this study is experimental using Bovine DBM. Bone Marrow Mesenchymal Stem Cell (BMSCs) and Adipose Mesenchymal Stem Cells (ASCs) were taken from New Zealand white rabbit. There are two groups of treatment, divided into DBM implanted with BMSCs and DBM implanted with ASCs. Each BMSCs and ASCs groups is incubated in the normal and osteogenic culture plate. Evaluation is performed by counting the osteoblast and immunohistochemistry stain using Alkaline Phosphate and Osteocalcin. After 4 weeks of incubation, we found that the osteoblast count in BMSCs groups is higher compared to the ASCs groups in both culture condition (p&lt;0.01) along with Alkaline Phosphate staining (p&lt;0.05), while the Osteocalcin staining showed insignificant differences (p&gt;0.05). This study revealed that xenogenic bovine DBM can act as the potential osteoinductive scaffold for the MSCs to differentiate. The tissue engineering application by combining MSCs and Bovine DBM can be considered as an alternative in managing bone defect cases.

In vitro and in vivo evaluation of xenogeneic bone putty with the carrier of hydrogel derived from demineralized bone matrix.

The demineralized bone matrix (DBM) putty is a traditional bone graft utilized to facilitate the repair and reconstruction of bone. Recent studies indicated the DBM putties with the various carriers were different in bone repairing ability. In order to prepare a kind of DBM putty with a good biocompatibility and bioactivity, the DBM gel was processed from the DBM and the feasibility as a carrier for the DBM putty was evaluated. After the bovine DBM gel was prepared, the BMPs content as well as the ability to promote osteogenic differentiation of MC3T3-E1 cells in vitro were investigated. Then the DBM putty was prepared and filled into the rat calvarial defect model to evaluate the bone repairing ability by micro-CT and histology. The result showed there was 2.953 ± 0.054ng BMP contained in per gram of the DBM gel. And the ALP production of MC3T3-E1 cells in the DBM gels group increased with prolonged culturing, the mineralized nodules formed in MC3T3-E1 cells on 14th day after co-culture. The putty prepared by DBM gel was easy to handle without loss of DBM particles at room temperature. In the rat calvarial bone defect experiment, histological observation showed more mature bone formed in the DBM putty group than that in the type I collagen group at 12weeks, which indicated the bone putty prepared by DBM gel exhibited a better bone repair capability.

Application of bovine bone versus bovine DBM graft on bone healing of radial defect in rat

Reconstruction of the fractured bone and bone healing biology is compound processes, and the effectiveness of osteosynthesis is determined by their success. Currently, autografts or allografts and mineral bone substitutes are used commonly for skeletal restoration. The aim of this study is to evaluate the effects of bovine demineralized bone matrix (DBM) and bovine bone (bovine xenografts) on the bone healing process in rat. Sixteen female Sprague-Dawley rats were used in separate four groups: autograft (positive control, n = 4), untreated defect (negative control, n = 4), bovine DBM (n = 4), and bovine bone graft (n = 4). Radiological, histopathological, and biomechanical evaluations were done during the study, and results were recorded and analyzed statistically. At the 56th postoperative day, there was no significant differences in different groups in the biomechanical and radiographical criteria (P > 0.05), but there were significant differences in the mode of union and cancellous bone formation between the groups in histopathological evaluation at the 56th postoperative day (P < 0.05); the present study showed that there was no difference in the radiological and biomechanical performances of the healing bones between the bovine DBM and bovine bone graft.

Maxillary Sinus Floor Elevation Using Platelet-Rich Plasma Combined With Either Biphasic Calcium Phosphate or Deproteinized Bovine Bone.

Maxillary sinus floor elevation procedure has the objective of augmenting available bone in atrophic posterior maxilla to allow dental implants placement. The main aim of this prospective study was to evaluate clinically and histomorphometrically the performance of biphasic calcium phosphate (BCP) used in conjunction with platelet-rich plasma (PRP) compared with demineralized bovine bone matrix (DBBM) and PRP in sinus floor elevation surgery. Patients candidate to sinus floor elevation were treated using either BCP and PRP or DBBM and PRP. Biopsies were retrieved using trephine bur during implant placement surgery 6 months after grafting. Clinical success of implants was evaluated 1 year after prosthesis delivery. Histomorphometric analysis was performed assessing the relative volume of newly formed bone. A total of 20 patients were recruited, and 20 sinus augmentation procedures were performed, 10 for each group. A total of 42 implants were placed. One year after prosthetic loading a 100% implant survival rate was reported in both groups. Histomorphometric analysis revealed that the mean amount of new bone formation was 18.6 ± 3.3% in BCP group and it was 21.9 ± 4.9% in DBBM group, without statistically significant difference. In BCP group a greater amount of collagen type I was found with respect to DBBM group. Both grafting materials led to an excellent performance regarding implant survival rate after 1 year follow-up, without any significant adverse sequelae. A similar capability of inducing new bone formation was observed in both groups, even though the higher quantity of collagen type I in BCP group may suggest a greater potential for bone formation over time as compared with DBBM.

A Biomechanical Analysis of Treatment Options for Enchondromas of the Hand

Enchondromas are the most common benign bone tumor affecting the hand. Standard treatment has consisted of curettage with bone grafting. It has become increasingly common for surgeons to use biologic cement in these cases. The purpose of this study was to evaluate different treatment options used to fill voids after curettage of hand enchondromas to determine if any provide more rigid fixation. A cadaveric model of hand enchondromas was designed by making a standardized size corticotomy (0.6 × 1.0 cm) in 60 cadaver metacarpals. Resultant voids were then filled with either no material, bovine demineralized bone matrix (Synthes Paoli, PA, USA), or one of two different calcium phosphate bone cements: a carbonated apatite (Synthes Norian SRS Skeletal Repair System) and a hydroxyapatite (Stryker HydroSet) calcium phosphate. An apex dorsal three-point bend was applied to the metacarpals through an MTS machine, and load to failure and stiffness were recorded. Biomechanically, load to failure for intact metacarpals was significantly superior to those in which a corticotomy was created (p = 0.04). There was a significant increase in load to failure between the metacarpals treated with the calcium phosphate bone cement and the negative controls (p = 0). In a biomechanical analysis of a cadaveric model of hand enchondromas, calcium phosphate bone cement provided significantly increased strength as compared to curettage alone and approximated the strength of intact metacarpals. It is unknown whether the use of biologic cements in this clinical setting leads to less postoperative immobilization, earlier digital motion, a quicker return to work, or increased patient satisfaction.

English

Stimulating bone production may be applied to the management of fractures, non-unions, and osteomyelitis. Hydroxyapatite is compatible and osteoconductive for bone regeneration. This study was conducted to evaluate the effect of osteoinductive activity of xenogenic demineralized bone matrix (DBM) in combination with osteoconductive hydroxyapatite (HA) in osseous location. Twenty four adult New Zealand white rabbits (male, weight: 2.5 to 3.0 kg) were randomly divided into four groups. All rabbits were anesthetized with intramuscular administration of ketamin and xylazine. After preparing the operation site, the right radius was exposed, and a full thickness 5 mm defect was created in the mid-diaphyseal region. The defects were filled with DBM and hydroxyapatite in Group I, DBM in Group II, hydroxyapatite in Group III and Group IV was considered as a control group. The formation and healing of bone were determined by histological and mechanical analysis during 8 weeks. Implantation of combination of DBM and hydroxyapatite yielded significant (p &lt; 0.05) bone formation resulting in histological, and mechanical evidence of union compared to other groups. The second group which received DBM showed better bone healing compared to the third group. No instances of union were observed in the hydroxyapatite and control groups. Our results suggest that bovine DBM may be more useful in combination with hydroxyapatite as a therapeutic adjuvant in clinical situations when local formation of bone is needed. &nbsp; Key words:&nbsp;Demineralized bone matrix, coralline hydroxyapatite, bone healing, rabbit.

Influence of the association between simvastatin and demineralized bovine bone matrix on bone repair in rats

Simvastatin, a drug used to lower blood cholesterol, has been reported to have an anabolic effect on bone. The purpose of this study was to evaluate the influence of simvastatin and demineralized bovine bone matrix (DBBM) on the repair of rat calvarial defects. Defects of 5 mm were created in 64 rats, divided into four groups: no local treatment (control); treatment with DBBM (DBBM); treatment with a combination of simvastatin solution (2.2 mg/50 μl) and DBBM (DBBMSIM-1); and treatment with simvastatin solution (0.5 mg/50 μl) and DBBM (DBBMSIM-2). Animals were sacrificed on postoperative day 30 or 60, after which the calvariae were X-rayed and prepared for histomorphometric evaluation. The data were submitted to statistical analysis (p < 0.05). X-rays revealed that, on postoperative day 30, animals treated with a lower dose of simvastatin presented the lowest bone density, whereas on postoperative day 60 the use of simvastatin, regardless of the dose, resulted in lower density than that observed in control and DBBM group samples. Histomorphometric analysis revealed that, on postoperative day 30, both DBBM and DBBMSIM-1 had a negative impact on bone formation. On postoperative day 60, none of the combinations tested impaired bone repair. These results showed that the association between DBBM and simvastatin had a negative impact on bone repair.

Effects of demineralized bone matrix and a ‘Ricinus communis’ polymer on bone regeneration: a histological study in rabbit calvaria

The aim of the present study was to histologically analyze the effects of bovine and human demineralized bone matrix and a Ricinus communis polymer on the bone regeneration process. Two surgical bone defects were created in rabbit calvaria, one on the right and the other on the left side of the parietal suture. Eighteen rabbits were divided into three groups. In Group I, the experimental defect was treated with bovine demineralized bone matrix, Group II with human demineralized bone matrix, and in Group III, the experimental cavity was treated with polyurethane resin derived from Ricinus communis oil. The control defects were filled with the animals' own blood. The animals were sacrificed after 7 and 15 weeks. Histological analysis revealed that in all groups (control and experimental), bone regeneration increased with time. The least time required for bone regeneration was noted in the control group, with a substantial decrease in the thickness of the defect. All materials proved to be biologically compatible, but polyurethane resorbed more slowly and demonstrated considerably better results than the demineralized bone matrices.

Experimental bone defect healing with xenogenic demineralized bone matrix and bovine fetal growth plate as a new xenograft: radiological, histopathological and biomechanical evaluation

The following study was designed to evaluate xenogenic bovine demineralized bone matrix (DBM) and new xenograft (Bovine fetal growth plate) effects on bone healing process. Twenty male White New Zealand rabbits were used in this study. In group I (n = 10) the defect was filled by xenogenic DBM and in group II (n = 10) the defect was filled by a segment of bovine fetal growth plate and was fixed by cercelage wire. Radiological, histopathological, and biomechanical evaluations were performed blindly and results scored and analyzed statistically. Statistical tests did not support significant differences between two groups radiographically (P > 0.05). There was a significant difference for union at the 28th postoperative radiologically (P < 0.05). Xenograft was superior to DBM group at the 28th postoperative day for radiological union (P < 0.03). Histopathological and biomechanical evaluation revealed no significant differences between two groups. In conclusion, the results of this study indicate that satisfactory healing occurred in rabbit radius defect filled with xenogenic bovine DBM and xenogenic bovine fetal growth plate. Complications were not identified and healing was faster in two grafting groups.

Xenogenic demineralized bone matrix and fresh autogenous cortical bone effects on experimental bone healing: radiological, histopathological and biomechanical evaluation.

BackgroundBone grafting is used to enhance healing in osteotomies, arthrodesis, and multifragmentary fractures and to replace bony loss resulting from neoplasia or cysts. They are source of osteoprogenitor cells and induce bone formation and provide mechanical support for vascular and bone ingrowth. Autografts are used commonly but quantity of harvested bone is limited. The aim of this study is to evaluate autograft and new xenogenic bovine demineralized bone matrix (DBM) effects on bone healing process.Materials and methodsTwenty male White New Zealand rabbits were used in this study. In group I (n = 10) the defect was filled by xenogenic DBM and in autograft group the defect was filled by fresh autogenous cortical graft and fixed by cercelage wire. Radiological, histopathological and biomechanical evaluations were performed blindly and results scored and analyzed statistically.ResultsStatistical tests did not reveal any significant differences between two groups on the 14th postoperative day radiographically (P > 0.05). There was a significant difference for union on 28th and 42nd postoperative days and for remodeling at on the 56th postoperative day radiologically (P < 0.05). Statistical tests did not support any significant differences between two groups for radiological bone formation (P > 0.05). Histopathological and biomechanical evaluation revealed no significant differences between two groups.ConclusionsThe results of this study indicate that satisfactory healing occurred in rabbit radius defect filled with xenogenic bovine DBM. Complications were not identified and healing was faster, same as in cortical autogenous grafting.

Assessment of bovine biomaterials containing bone morphogenetic proteins bound to absorbable hydroxyapatite in rabbit segmental bone defects

To evaluate the osteo-regenerative capacity of two proprietary bone grafting materials, using a segmental defect model in both radial diaphyses of rabbits. The right defect was filled with pooled bone morphogenetic proteins (pBMPs) bound to absorbable ultrathin powdered hydroxyapatite (HA) mixed with inorganic and demineralized bone matrix and bone-derived collagen, derived from bovine bone (Group A). The left defect was filled with bovine demineralized bone matrix and pBMPs bound to absorbable ultrathin powdered HA (Group B). In both groups, an absorbable membrane of demineralized bovine cortical was used to retain the biomaterials in the bone defects, and to guide the tissue regeneration. The rabbits were euthanized 30, 90 and 150 days after surgery. Radiographic, tomographic and histologic evaluations were carried out on all specimens. At 30 days, the demineralized cortical bone cover was totally resorbed in both groups. HA was totally resorbed from Group A defects, whereas HA persisted in Group B defects. A prominent foreign body reaction was evident with both products, more pronounced in sections from Group B. At 90 days, the defects in Group B exhibited more new bone than Group A. However, at 150 days after surgery, neither treatment had stimulated complete repair of the defect. The partial bone healing of the segmental defect occurred with low or none performance of the biomaterials tested.

Pool of bovine morphogenetic proteins and guided tissue regeneration in the treatment of intrabony periodontal defects: I- Clinical measurements

The purpose of this study was to evaluate the effect of the pool of bovine BMPs on the treatment of intrabony defects. The sample comprised 15 patients aged 26 to 57 years old presenting with 10 pairs of lesions of 2 or 3 walls or 2-3 walls ³5mm, located in the same type teeth (premolar or molar) and same jaw. The test defects were treated with combination of a pool of bovine bone morphogenetic and resorbable hydroxyapatite carrier (BMPs- HA), bovine demineralized bone matrix (MB) and coverage by a bovine collagen barrier membrane. The control defects were treated with MB-HA and covered by a bovine collagen membrane. The clinical measurements at six months after therapy in the test group revealed a reduction in the mean probing pocket depth (PPD) of 1.63 ± 1.41mm (B) and 1.93 ± 0.96mm (L) and a mean change in the clinical attachment level (CAL) of 1.60 ± 1.16mm (B) and 1.46 ± 0.97mm (L). The control group showed a mean reduction of PPD of 1.93 ± 1.34mm (B) and 2.0 ± 1.51mm (L) and a mean change of CAL of 1.03 ± 1.24mm (B) and 1.30 ± 1.14 mm (L). The analysis of variance (ANOVA) demonstrated that the changes in the clinical parameters were statistically significant (p<0.05). There were no significant differences between the test and controls subjects (Student's test, p<0.005). These findings suggest that the use of a pool of bovine BMPs do not provide added effects to GTR in the treatment of intrabony defects.

A new in vivo model for testing cartilage grafts and biomaterials: the ‘rabbit pinna punch-hole’ model

In this study an animal model was developed for evaluation of the feasibility of cartilage grafts. In the cartilage of the external ear of the rabbit multiple holes, 6 mm in diameter, were punched, leaving the adherent skin intact. Different experimental groups were evaluated. First, the punch-hole model was validated under various conditions to study spontaneous or perichondrial initiated regeneration of the cartilage defect. When both cartilage and perichondrium was excised no spontaneous repair of the cartilage defect was observed. When perichondrium is present, variable patch-like closure of the punch hole was found. As ‘golden standard’ a punched out piece of cartilage was reimplanted directly. This condition showed adequate closure of the punch hole, however, no perfect integration of graft and surrounding cartilage was observed. Secondly, to evaluate the ‘punch-hole model’ a biomaterial ,trabecular demineralized bovine bone matrix (DBM), was implanted and tested as a scaffold for tissue engineering techniques in vivo and in vitro. Direct implantation of DBM did not lead to any cartilage formation to close the defect. In vivo engineered cartilage, generated by enveloping DBM in perichondrium for 3 weeks, could adequately close the punch hole. When DBM was seeded with isolated chondrocytes in vitro before implantation in the defect, a highly fragmented graft, with some islets of viable cells was seen. To promote an efficient and reliable evaluation of cartilage grafts a semi-quantitative grading system was developed. Items such as quality, quantity and integrity of the cartilage graft were included in a histomorphological grading system to provide information about the properties of a specific cartilage graft. To validate the grading system, all conditions were scored by two independent observers. An excellent reliability ( R=0.96) was seen between the observers. In summary, the rabbit pinna punch-hole model is a reliable and efficient method for first evaluation of cartilage grafts. The results can be easily analyzed using a semi-quantitative grading system.

Tissue-engineered cartilage using serially passaged articular chondrocytes. Chondrocytes in alginate, combined in vivo with a synthetic (E210) or biologic biodegradable carrier (DBM)

In vitro multiplication of isolated autologous chondrocytes is required to obtain an adequate number of cells to generate neo-cartilage, but is known to induce cell-dedifferentiation. The aim of this study was to investigate whether multiplied chondrocytes can be used to generate neo-cartilage in vivo. Adult bovine articular chondrocytes, of various differentiation stages, were suspended in alginate at densities of 10 or 50 million/ml, either directly after isolation (P0) or after multiplication in monolayer for one (P1) or three passages (P3). Alginate with cells was seeded in demineralized bovine bone matrix (DBM) or a fleece of polylactic/polyglycolic acid (E210) and implanted in nude mice for 8 weeks. The newly formed tissue was evaluated by Alcian Blue and immunohistochemical staining for collagen type-II and type-I. Structural homogeneity of the tissue, composed of freshly isolated as well as serially passaged cells, was found to be enhanced by high-density seeding (50 million/ml) and the use of E210 as a carrier. The percentage of collagen type-II positive staining P3-cells was generally higher when E210 was used as a carrier. Furthermore, seeding P3-chondrocytes at the highest density (50 million/ml) enhanced collagen type-II expression. This study shows promising possibilities to generate structurally regular neo-cartilage using multiplied chondrocytes in alginate in combination with a fleece of polylactic/polyglycolic acid.

Characterization of matrix-induced osteogenesis in rat calvarial bone defects: I. Differences in the cellular response to demineralized bone matrix implanted in calvarial defects and in subcutaneous sites.

The cellular and biochemical sequences of osteogenesis induced by implanting demineralized bone matrix (DBM) in rat cranial defects and in subcutaneous sites have been studied by histological, histochemical, and biochemical techniques from days 2 to 28 after implantation. In subcutaneous sites, allogenic DBM induced cartilage cells and matrix for approximately the first 10 days which were subsequently resorbed and replaced by bone with little evidence for the classical endochondral sequence of ossification. In sharp contrast, the first cells that differentiated from the mesenchymal stem cells in the cranial defects were alkaline phosphatase (ALP) positively stained osteoblasts that appeared 3 days after implantation followed by synthesis of bone matrix which calcified shortly thereafter. A few clusters of cartilage cells were observed beginning at days 6-7 which were spatially distinct from the new bone and later resorbed. By day 28 the tissue induced in both the subcutaneous and cranial sites consisted almost solely of bone; however, the total amount of new bone in the subcutaneous implants was significantly less than the mass of bone formed in the calvarial defects. Bovine DBM induced bone formation in rat cranial defects to a very much lesser extent than allogenic DBM. A few cartilage cells were induced by bovine DBM in subcutaneous sites and rapidly resorbed and not replaced with bone. These results clearly indicate that the cellular sequence induced by allogenic and xenogenic DBM and the repair tissues synthesized are distinctly different in the cranial defects from those induced in the subcutaneous sites.

Cartilage induction for reconstruction

Objective. By ‘controlled engineering’ autologous cartilage could be produced in vivo. This cartilage was used as graft for septal reconstruction in children. The producer of cells was a pedicled perichondrium flap of the pinna, the inductor was demineralized bovine bone matrix (DBM). Two aspects were investigated (1) if cartilage production could be stimulated in a free perichondrial flap and (2) how the demineralized bone is resorbed. Methods. DBM was implanted subcutaneously in the ear, subcutaneously in the abdomen or intra-muscularly in the quadriceps both with and without a perichondrium envelope. Degradation of DBM and formation of cartilage were studied in a time series using (immuno)histochemistry. Results. Resorption of DBM was mediated by macrophages. Formation of new cartilage occurred only in the presence of perichondrium. The generated cartilage contained hyaline cartilage specific collagen and proteoglycans. In a less vascular environment this process of cartilage formation was slower and less predictable. Conclusions. (1) A free perichondrium flap is an excellent producer of cells for cartilage production. (2) The cartilage inductor (DMB) is completely resorbed by macrophages. (3) Even larger intramuscularly grafts can be generated. (4) The new formed autologous cartilage is less differentiated and seems to demonstrate better wound healing.