Showed Bone Formation Research Articles

Isolation and expansion of osteogenic cells from bone marrow and in vivo testing at ectopic sites

BACKGROUND: Cell therapy is a strategy to enhance bone formation in regenerative dentistry. Osteogenic cells can be isolated and expanded from total bone marrow by the use of biodegradable microcarriers. It remains, however, unknown whether the cells on the surface of microcarriers can provide bone substitutes with an osteogenic potential. METHODS: We therefore isolated and expanded rat bone marrow cells on the surface of CultiSpher G microcarriers in a spinner-flask bioreactor. Isolated cells were viable as determined by MTT assay and stained positive for alkaline phosphatase activity. CultiSpher G microcarriers loaded with osteogenic cells were mixed with two bone substitutes, natural bone mineral, and hydroxyapatite/tricalcium, phosphate before subcutaneous implantation into immunodeficient mice. RESULTS: Isolated cells showed positive formazan formation indicating cell viability. Alkaline phosphatase activity being a marker gene for osteogenic differentiation was detectable. However, implants recovered after eight weeks from ectopic sites failed to show bone formation, even though blood vessels were present. CONCLUSIONS: These results suggest that the presence of osteogenic cells on the surface of microcarriers is not sufficient to provide bone substitutes with a bone forming capacity. The lack of bone formation at ectopic sites does not rule out the possibility that supplementation of bone substitutes with cell-loaded microcarrier support bone regeneration at orthotopic sites. Moreover, our data do not exclude the possibility that the current concept can be optimized to achieve ectopic bone formation.

The Role of Endothelial Progenitor Cells in Prevascularized Bone Tissue Engineering: Development of Heterogeneous Constructs

In vitro prevascularization of bone grafts with endothelial progenitor cells (EPCs) is a promising strategy to improveimplant survival. In this study we show bone formation in constructs that contain multipotent stromal cells (MSCs) and EPCs. Early and late EPCs from peripheral blood and bone marrow of adult goats were characterized for differentiation markers and functional responses. EPCs from peripheral blood are more proliferative than bone-marrow-derived EPCs, express higher numbers of endothelial markers for longer periods of time, and form more intricate networks. We demonstrate that EPCs derived from peripheral blood contribute to osteogenic differentiation by MSCs in vitro, and that MSCs support the proliferation of EPCs and stabilize the formed cellular networks. In vivo, EPCs from peripheral blood assemble into early blood vessel networks, which are more pronounced in the presence of MSCs. These results show that the EPCs isolated from peripheral blood are suitable for prevascularization strategies, and that coseeding of EPCs and MSCs is favorable for bone formation after 6 weeks.

Early bone formation in furrows at titanium implants. A study in the rabbit tibia

Aim Previous studies have shown a preference for bone formation in macroscopic furrows as compared to other parts of the implant surface. The aim of the study was to describe the early events of bone integration of furrowed oxidized titanium implants. Materials and Methods New Zealand white rabbits and 54 implants were used in the study. The test oxidized implants had a 110 μm wide furrow added to the superior flank whilst the control oxidized implants had no furrow. The implants were retrieved after 7, 14 and 28 days for light microscopy and for micro-CT evaluation. Results The bone contact and bone area values increased with time for all implants. A statistically significant preference for bone formation in the furrows was seen for test implants and for the inferior thread flank for control implants compared to the opposing superior thread flank. The 3D reconstructions from micro-CT showed bone formation as thin rims along the implant surface either continuous or as solitary rims. Conclusion It is concluded, that bone formation is observed more often at the lower flank and in macroscopic furrows at the threads than opposing upper flanks of oxidized titanium implants. This was more marked at seven days than later time points and in areas with no primary contacts with adjacent host bone. Bone integration of the implants seemed to occur by bone condensation directly at and along the implant surface and following the path of the implant threads.

Buccal fat pad versus sandwich graft for treatment of oroantral defects: A comparison

Aim:To compare the efficacy of buccal fat pad (BFP) graft with sandwich graft (hydroxyapatite crystals embedded within collagen sheath) in closure of oroantral defects.Materials and Methods:A 2-year prospective study was conducted; 20 patients were included in the study were divided into two groups having 10 patients in each. Group I patients underwent surgical closure of oroantral fistula with sandwich graft and Group II patients with buccal pad of fat.Results:In Group I, the mean pain scores were 7.60 ± 0.84, 3.90 ± 1.10, 2.30 ± 1.16, 1.10 ± 0.99 and 0.40 ± 0.70 at immediate post-op., 1, 3, 6 and 12 week time intervals, respectively, whereas in Group II these were 7.30 ± 0.67, 3.50 ± 0.53, 1.70 ± 0.48, 1.00 ± 0.47 and 0.30 ± 0.48, respectively, at the corresponding time intervals. In Group I, swelling was seen to be present in 10 (100%), 7 (70%), 2 (20%) and nil (0%) patients at 1, 3, 6 and 12 weeks, respectively, whereas in Group II, it was seen to be present in 10 (100%), 10 (100%), 2 (20%) and nil (0%) patients at the corresponding time intervals. At 1 week, infection was seen to be present in 1 (10%) patient of Group I and 2 (20%) patients of Group II. At 3 and 6 weeks, infection was seen to be present in 1 (10%) patient of Group I and none of the patients of group II. No radiologic evidence of bone formation was seen in either group up to 1 week. At 3 week interval, there were 6 (60%) patients in Group I and nil (0%) in Group II showing bone formation, thus showing a statistically significant difference between the two groups. By 6 week time interval, radiologic evidence of bone formation was seen in 9 (90%) patients of Group I but in no patients of Group II, thereby showing a statistically very highly significant (P < 0.001) difference between the two groups. In Group I, in 1 (10%) patient, graft was rejected by first week; however, no further graft rejection took place. In Group II, no case of graft rejection was reported.Conclusions:The sandwich graft technique yielded a more promising closure of oroantral communication by provision of a more biologically apt base in terms of regeneration of lost bone structure at the floor of the maxillary sinus

Histological Findings Following the Use of a Space‐Making Device for Bone Reformation and Implant Integration in the Maxillary Sinus of Primates

Previous studies have shown that membrane elevation results in predictable bone formation in the maxillary sinus provided that implants can be placed as tent poles. In situations with an extremely thin residual crest which impairs implant placement, it is possible that a space-making device can be used under the sinus membrane to promote bone formation prior to placement of implants. The present study was conducted to test the hypothesis that the use of a space-making device for elevation of the sinus membrane will result in predictable bone formation at the maxillary sinus floor to allow placement of dental implants. Eight tufted capuchin primates underwent bilateral sinus membrane elevation surgery, and a bioresorbable space-making device, about 6 mm wide and 6 mm in height, was placed below the elevated membrane on the sinus floor. An oxidized implant (Nobel Biocare AB, Gothenburg, Sweden) was installed in the residual bone protruding into the created space at one side while the other side was left without an implant. Four animals were sacrificed after 6 months of healing. The remaining four animals received a second implant in the side with a space-making device only and followed for another 3 months before sacrifice. Implant stability was assessed through resonance frequency analysis (RFA) using the Osstell (Osstell AB, Gothenburg, Sweden) at installation, 6 months and 9 months after the first surgery. The bone-implant contact (BIC) and bone area inside the threads (BA) were histometrically evaluated in ground sections. Histologically there were only minor or no signs of bone formation in the sites with a space-making device only. Sites with simultaneous implant placement showed bone formation along the implant surface. Sites with delayed implant placement showed minor or no bone formation and/or formation of a dense fibrous tissue along the apical part of the implant surface. In the latter group the apical part of the implant was not covered with the membrane but protruded into the sinus cavity. The use of a space-making device, with the design used in the present study, does not result in bone formation at the sinus floor. However, membrane elevation and simultaneous placement of the device and an implant does result in bone formation at the implant surface while sites with implants placed 6 months after membrane elevation show only small amounts of bone formation. It is suggested that lack of stabilization of the device and/or a too extensive elevation of the membrane may explain the results.

Histological Outcomes on the Development of New Space-making Devices for Maxillary Sinus Floor Augmentation

Previous studies have pointed out that the mere elevation of the maxillary sinus membrane promotes bone formation without the use of augmentation materials. This experimental study aimed at evaluating if the two-stage procedure for sinus floor augmentation could benefit from the use of a space-making device in order to increase the bone volume to enable later implant installation with good primary stability. Six male tufted capuchin primates (Cebus apella) were subjected to extraction of the three premolars and the first molar on both sides of the maxilla to create an edentulous area. The sinuses were opened using the lateral bone-wall window technique, and the membrane was elevated. One resorbable space-making device was inserted in each maxillary sinus, and the bone window was returned in place. The animals were euthanatized after 6 months, and biopsy blocks containing the whole maxillary sinus and surrounding soft tissues were prepared for ground sections. The histological examination of the specimens showed bone formation in contact with both the schneiderian membrane and the device in most cases even when the device was displaced. The process of bone formation indicates that this technique is potentially useful for two-stage sinus floor augmentation. The lack of stabilization of the device within the sinus demands further improvement of space-makers for predictable bone augmentation. It is concluded that (1) the device used in this study did not trigger any important inflammatory reaction; (2) when the sinus membrane was elevated, bone formation was a constant finding; and (3) an ideal space-making device should be stable and elevate the membrane to ensure a maintained connection between the membrane and the secluded space.

SEM-EDS and biomechanical evaluation of implants with different surface treatments: an initial study

Aim Alterations in implant surfaces can affect periimplant bone formation and shorten the healing time. The goal of the present study was a comparative scanning electron microscopy (SEM)/energy dispersive spectrometry (EDS) and biomechanical evaluation of implants subjected to different surface treatments. Materials and Methods Four implant surfaces were analyzed in the present study: machined commercial implants (TU); porous-surfaced commercial implants blasted with Al2O3 microspheres and acid-etched (TJA); laser beam-irradiated experimental implants (Laser) and laser beam-irradiated experimental implants with hydroxyapatite coating (HA). One sample for each surface underwent pre-surgery SEM / EDS analysis. Thirty- two implants (8 for each surface treatment) were then inserted into the tibia of 4 rabbits. After 8 weeks, the animals were euthanized and the implants retrieved by reverse torque and processed for post-surgery SEM / EDS analysis. Results HA implants presented higher removal torque values when compared to Laser, TJA and TU groups. Post-surgery SEM micrographs clearly showed bone formation on all the examined surfaces; however, in the TU group bone covered only some areas of the implant surface, while in TJA, Laser and HA groups the entire implant surfaces were overlaid by newly formed bone. EDS analysis supported the results obtained by SEM and removal torque, showing that concentration of Ca and P increased from TU to TJA, Laser and HA implants. Conclusions Implants with surfaces modified by laser beam with or without apatite coating showed more promising results.

Effect of Autologous Bone Marrow Stromal Cell Seeding and Bone Morphogenetic Protein-2 Delivery on Ectopic Bone Formation in a Microsphere/Poly(Propylene Fumarate) Composite

A biodegradable microsphere/scaffold composite based on the synthetic polymer poly(propylene fumarate) (PPF) holds promise as a scaffold for cell growth and sustained delivery vehicle for growth factors for bone regeneration. The objective of the current work was to investigate the in vitro release and in vivo bone forming capacity of this microsphere/scaffold composite containing bone morphogenetic protein-2 (BMP-2) in combination with autologous bone marrow stromal cells (BMSCs) in a goat ectopic implantation model. Three composites consisting of 0, 0.08, or 8 microg BMP-2 per mg of poly(lactic-co-glycolic acid) microspheres, embedded in a porous PPF scaffold, were combined with either plasma (no cells) or culture-expanded BMSCs. PPF scaffolds impregnated with a BMP-2 solution and combined with BMSCs as well as empty PPF scaffolds were also tested. The eight different composites were implanted subcutaneously in the dorsal thoracolumbar area of goats. Incorporation of BMP-2-loaded microspheres in the PPF scaffold resulted in a more sustained in vitro release with a lower burst phase, as compared to BMP-2-impregnated scaffolds. Histological analysis after 9 weeks of implantation showed bone formation in the pores of 11/16 composites containing 8 microg/mg BMP-2-loaded microspheres with no significant difference between composites with or without BMSCs (6/8 and 5/8, respectively). Bone formation was also observed in 1/8 of the BMP-2-impregnated scaffolds. No bone formation was observed in the other conditions. Overall, this study shows the feasibility of bone induction by BMP-2 release from microspheres/scaffold composites.

Human tissue-engineered bone produced in clinically relevant amounts using a semi-automated perfusion bioreactor system: a preliminary study

The aim of this study was to evaluate a semi-automated perfusion bioreactor system for the production of clinically relevant amounts of human tissue-engineered bone. Human bone marrow stromal cells (hBMSCs) of eight donors were dynamically seeded and proliferated in a perfusion bioreactor system in clinically relevant volumes (10 cm(3)) of macroporous biphasic calcium phosphate scaffolds (BCP particles, 2-6 mm). Cell load and distribution were shown using methylene blue staining. MTT staining was used to demonstrate viability of the present cells. After 20 days of cultivation, the particles were covered with a homogeneous layer of viable cells. Online oxygen measurements confirmed the proliferation of hBMSCs in the bioreactor. After 20 days of cultivation, the hybrid constructs became interconnected and a dense layer of extracellular matrix was present, as visualized by scanning electron microscopy (SEM). Furthermore, the hBMSCs showed differentiation towards the osteogenic lineage as was indicated by collagen type I production and alkaline phosphatase (ALP) expression. We observed no significant differences in osteogenic gene expression profiles between static and dynamic conditions like ALP, BMP2, Id1, Id2, Smad6, collagen type I, osteocalcin, osteonectin and S100A4. For the donors that showed bone formation, dynamically cultured hybrid constructs showed the same amount of bone as the statically cultured hybrid constructs. Based on these results, we conclude that a semi-automated perfusion bioreactor system is capable of producing clinically relevant and viable amounts of human tissue-engineered bone that exhibit bone-forming potential after implantation in nude mice.

Comparative study of biphasic calcium phosphates with different HA/TCP ratios in mandibular bone defects. A long‐term histomorphometric study in minipigs

Three biphasic calcium phosphate (BCP) bone substitute materials with hydroxyapatite (HA)/tricalcium phosphate (TCP) ratios of 20/80, 60/40, and 80/20 were compared to coagulum, particulated autogenous bone, and deproteinized bovine bone mineral (DBBM) in membrane-protected bone defects. The defects were prepared in the mandibles of 24 minipigs that were divided into four groups of six with healing times of 4, 13, 26, and 52 weeks, respectively. The histologic and histomorphometric evaluation focused on differences in amount and pattern of bone formation, filler degradation, and the interface between bone and filler. Collapse of the expanded polytetrafluoroethylene barrier membrane into the coagulum defects underlined the necessity of a filler material to maintain the augmented volume. Quantitatively, BCP 20/80 showed bone formation and degradation of the filler material similar to autografts, whereas BCP 60/40 and BCP 80/20 rather equaled DBBM. Among the three BCP's, the amount of bone formation and degradation of filler material seemed to be inversely proportional to the HA/TCP ratio. The fraction of filler surface covered with bone was highest for autografts at all time points and was higher for DBBM than BCP 80/20 and 60/40 at the early healing phase. TRAP-positive multinucleated cells were identified on BCP and DBBM surfaces without showing typical signs of resorption lacunae.

The Bone Tissue Responses to Prehydrated and Collagenated Cortico‐Cancellous Porcine Bone Grafts: A Study in Rabbit Maxillary Defects

Bone substitutes should have osteoconductive properties and be completely replaced with new bone with time. Adding collagen gel to prehydrated and collagenated porcine bone (PCPB) particles results in a sticky and moldable material which facilitates clinical handling. However, the possible influence of the gel on the bone tissue response is not known. The objective of the study was to evaluate the bone tissue responses to PCPB graft with or without collagen gel and to evaluate the resorption/degradation properties of the biomaterials. Fourteen rabbits were used in the study. Bilateral bone defects, 5 x 8 x 3 mm, were created in the maxilla and filled with PCPB + collagen gel (test) or with PCPB only (control) and covered with a collagen membrane. Animals were killed after 2 (n = 3), 4 (n = 3), and 8 weeks (n = 8) for histological and morphometrical evaluations. There were no differences between test and control defects. Both materials showed bone formation directly on the particles by typical osteoblastic seams. The bone area increased with time (2-8 weeks) for both sides, from 16.2% (control) and 19.2% (test) to 42.7 and 43.8%, respectively. The PCPB, whether mixed with collagen gel or not, was resorbed by osteoclasts as well as part of remodeling with the formation of osteons within the particles. Morphometry showed a decrease of PCPB area from 19.4% (control) and 23.8% (test) after 2 weeks to 3.7 and 9.3% after 8 weeks, respectively. Mixing collagen gel and PCPB to facilitate the clinical handling does not influence the bone tissue responses to the material, which exhibited osteoconductive properties and was resorbed with time.

A Case of Psoas Ossification From the Use of BMP-2 for Posterolateral Fusion at L4–L5

Case report. Describe a complication from the use of bone morphogenetic protein-2 (BMP-2) for posterolateral lumbar fusion. N/A, case report. The patient is a 69-year-old white male with a 1 year history of back and right leg pain with a foot drop due to spinal stenosis and a degenerative spondylolisthesis at L4-L5. Surgery for decompression and instrumented fusion was performed using BMP. Within 3 months after surgery the patient developed pain along his iliac wing, groin, and greater trochanter on the right side. Work up over the next several months included a bone scan, radiographs, Dexa scan, computed tomography scan, and magnetic resonance imaging with gadolinium and fat suppression. These tests showed extensive bone formation in the psoas and iliacus on the right side. The bone has persisted at least 2 years after surgery and shows no sign of resorption. BMP-2 can cause heterotopic bone formation when used for posterolateral lumbar fusion.

Effect of seeding technique and scaffold material on bone formation in tissue‐engineered constructs

The aim of the present study was to test the hypothesis that both scaffold material and the type of cell culturing contribute to the results of in vivo osteogenesis in tissue-engineered constructs in an interactive manner. CaCO3 scaffolds and mineralized collagen scaffolds were seeded with human trabecular bone cells at a density of 5 x 10(6) cells/cm(3) and were left to attach under standard conditions for 24 h. Subsequently, they were submitted to static and dynamic culturing for 14 days (groups III and IV, respectively). Dynamic culturing was carried out in a continuous flow perfusion bioreactor. Empty scaffolds and scaffolds that were seeded with cells and kept under standard conditions for 24 h served as controls (groups I and II, respectively). Five scaffolds of each biomaterial and from each group were implanted into the gluteal muscles of rnu rats for 6 weeks. Osteogenesis was assessed quantitatively by histomorphometry and expression of osteocalcin (OC) and vascular endothelial growth factor (VEGF) was determined by immunohistochemistry. CaCO3 scaffolds exhibited 15.8% (SD 3.1) of newly formed bone after static culture and 22.4% (SD 8.2) after dynamic culture. Empty control scaffolds did not show bone formation, and scaffolds after 24 h of standard conditions produced 8.2% of newly formed bone (SD 4.0). Differences between the controls and the scaffolds cultured for 14 days were significant, but there was no significant difference between static and dynamic culturing. Mineralized collagen scaffolds did not show bone formation in any group. There was a significant difference in the expression of OC within the scaffolds submitted to static versus dynamic culturing in the CaCO3 scaffolds. VEGF expression did not show significant differences between static and dynamic culturing in the two biomaterials tested. It is concluded that within the limitations of the study the type of biomaterial had the dominant effect on in vivo bone formation in small tissue-engineered scaffolds. The culture period additionally affected the amount of bone formed, whereas the type of culturing may have had a positive effect on the expression of osteogenic markers but not on the quantity of bone formation.

Cell based bone tissue engineering in jaw defects

In 6 patients the potency of bone tissue engineering to reconstruct jaw defects was tested. After a bone marrow aspirate was taken, stem cells were cultured, expanded and grown for 7 days on a bone substitute in an osteogenic culture medium to allow formation of a layer of extracellular bone matrix. At the end of the procedure, this viable bone substitute was not only re-implanted in the patient, but also simultaneously subcutaneously implanted in mice to prove its osteogenic potency. In all patients, a viable bone substitute was successfully constructed, which was proven by bone formation after subcutaneous implantation in mice (ectopic bone formation). However, the same construct was reluctant to form bone in patients with intra-oral osseous defects (orthotopic bone formation). Although biopsies, taken 4 months after reconstructing the intra-oral bone defect, showed bone formation in 3 patients, only in 1 patient bone formation was induced by the tissue-engineered construct. Although bone tissue engineering has proven its value in animal studies, extra effort is needed to make it a predictable method for reconstruction jaw defects in humans. To judge its benefit, it is important to differentiate between bone formation induced by cells from the border of the osseous defect (osteoconduction) in relation to bone matrix produced by the implanted cells (osteogenesis).

Acceleration of de novo bone formation with a novel bioabsorbable film: a histomorphometric study in vivo

Different types of barrier membranes have been used in periodontal applications for the technology of guided tissue regeneration (GTR). The aim of this study was to characterize the biological effect of novel calcium alginate film (CAF) on bone tissue regeneration by using rabbit mandible defects model. A critical size defect (5 mm in diameter) was created in the bilateral corner of mandible of 45 adult rabbits. The defects were covered with CAF served as the experimental group, or conventional collagen membrane (CCM) or left empty as the controls. Animals were killed after 1, 2, 4, 6 and 8 weeks. Morphological and histomorphometric studies were performed to evaluate their bone regeneration pattern and biological effects. Histological sections showed that bone regeneration pattern was centripetal in growth from defect rim. The quantitative histometry analysis revealed a significantly greater percentage of newly generated bone in CAF defects than that in CCM defects and empty defects from 2 to 6 weeks post-operation (P < 0.01). After 6 and 8 weeks, significantly more mature lamella bone had formed with CAF than with CCM. Empty control defects showed bone formation starting from the defect margins and incomplete healing even after 8 weeks. The CAF guided early bone growth and appeared more effective as a bioabsorbable GTR membrane than CCM. This study with mandible defect model suggests that bone defects augmented with CAF may offer most promising results from a histological and histomorphometric perspective.

Combination of anorganic bovine‐derived hydroxyapatite with binding peptide does not enhance bone healing in a critical‐size defect in a rabbit model

Anorganic bovine-derived hydroxyapatite (ABM) in combination with binding peptid (P-15) has demonstrated the capacity to improve the healing of periodontal defects. This study evaluated the benefit of ABM/P-15 to promote healing of cortical long bone defects in a rabbit model. A 5-mm segmental bone defect was created in the femur and fixed with a plate. There were two treatment groups: no implant (n = 12) and ABM/P-15 (n = 12). At 4, 8, and 12 weeks, healing of the defect was evaluated with radiographs and histomorphometric examination of the treated femora. After 4 weeks, radiographs showed bone formation without signs of complete consolidation in three of four animals in the control group and two of four ABM/P-15 treated animals. At the later course of the treatment, no radiologic difference was evident between the treatment groups. Histomorphometric evaluation revealed an area of 1.29 +/- 0.11 mm(2) and 0.97 +/- 0.21 mm(2) of newly produced bone in animals of the control group and ABM/P-15 group after 4 weeks. After 8 and 12 weeks, animals in the control group had an area of 2.44 +/- 0.62 mm(2) and 2.5 +/- 0.2 mm(2) of newly produced bone within the osteotomy gap compared to 1.6 +/- 0.65 mm(2) and 1.56 +/- 0.27 mm(2) in the ABM/P-15 group (p = 0.0004). An enhanced or accelerated ingrowth of bone, as reported in previous studies, was not observed. Our results imply that the ABM/P-15 is not a suitable graft for the treatment of critical-sized segmental defects in long bones.

Ectopic Osteogenesis by Ex Vivo Gene Therapy Using Beta Tricalcium Phosphate as a Carrier

Injuries and other damage to large bone can result in defects that do not heal spontaneously and lead to severe functional impairment. Better therapies are greatly needed to address this worldwide problem. The objective of the present study was to determine whether adenoviral delivery of modified human BMP2 gene (AdBMP2) using beta tricalcium phosphate (ß-TCP) as a carrier could promote osteoblastic differentiation of bone marrow mesenchymal stem cells (BMSCs) and ectopic bone formation. Rabbit BMSCs were separated from tibia aspirates and expanded in vitro. The BMSCs were then infected with AdBMP-2. Expression of BMP2, alkaline phosphatase, type I collagen, osteonectin, osteopontin, and mineralization of the cells confirmed secretion of active BMP2. Cells were observed to differentiate and maintain the osteoblast phenotype. For additional in vivo experiments, subcutaneous pockets were created on the backs of nude mice, which were then implanted with AdBMP2-BMSCs/ß-TCP, Adβgal-BMSCs/ß-TCP, BMSCs/ß-TCP, or ß-TCP alone. The nude mice were sacrificed after 4 weeks for histological evaluation. Adβgal-BMSCs/ß-TCP, BMSCs/ß-TCP, and ß-TCP did not show bone formation, although extensive fibrous tissue formed in the subcutaneous space in the rats implanted with ß-TCP. However, new bone tissue formation was observed on the inner walls of the pores of the ß-TCP-treated animals, and ectopic bone formation (mainly ‘‘cartilage-bone inducing’’) was observed in the AdBMP2-BMSCs/ß-TCP composite. These results confirmed the osteogenic potential of BMSCs after AdBMP2 transduction and revealed that AdBMP2-BMSC/ß-TCP composites could provide the capacity for bone formation and maturation during the more advanced stages of healing.

Effect of low‐level laser therapy on bone repair: Histological study in rats

Bone remodeling is characterized as a cyclic and lengthy process. It is currently accepted that not only this dynamics is triggered by a biological process, but also biochemical, electrical, and mechanical stimuli are key factors for the maintenance of bone tissue. The hypothesis that low-level laser therapy (LLLT) may favor bone repair has been suggested. The purpose of this study was to evaluate the bone repair in defects created in rat lower jaws after stimulation with infrared LLLT directly on the injured tissue. Bone defects were prepared on the mandibles of 30 Holtzman rats allocated in two groups (n = 15), which were divided in three evaluation period (15, 45, and 60 days), with five animals each. control group-no treatment of the defect; laser group-single laser irradiation with a GaAlAs semiconductor diode laser device (lambda = 780 nm; P = 35 mW; t = 40 s; Theta = 1.0 mm; D = 178 J/cm(2); E = 1.4 J) directly on the defect area. The rats were sacrificed at the pre-established periods and the mandibles were removed and processed for staining with hematoxylin and eosin, Masson's Trichrome and picrosirius techniques. The histological results showed bone formation in both groups. However, the laser group exhibited an advanced tissue response compared to the control group, abbreviating the initial inflammatory reaction and promoting rapid new bone matrix formation at 15 and 45 days (P<0.05). On the other hand, there were no significant differences between the groups at 60 days. The use of infrared LLLT directly to the injured tissue showed a biostimulating effect on bone remodeling by stimulating the modulation of the initial inflammatory response and anticipating the resolution to normal conditions at the earlier periods. However, there were no differences between the groups at 60 days.

A New Technique for Reconstruction of the Atrophied Narrow Alveolar Crest in the Maxilla Using Morselized Impacted Bone Allograft and Later Placement of Dental Implants

In cases when the alveolar crest is too narrow to host an implant, lateral augmentation is required. The use of autogenous bone blocks harvested from the iliac crest is often demanded. One disadvantage is the associated patient morbidity. The purpose of this study was to clinically and histologically evaluate the use of morselized impacted bone allograft, a novel technique for reconstruction of the narrow alveolar crest. Two patients with completely edentulous maxillae and one partially edentulous, with a mean age of 77 years (range 76-79 years) were included in the study. The alveolar crest width was <3 mm without possibility to place any implant. Bone grafts were taken from a bone bank in Gävle Hospital. Bone from the neck of femur heads was milled to produce bone chips. The milled bone was partially defatted by rinsing in 37 degrees C saline solution. After compression of the graft pieces with a size of 15 mm (height), 30 mm (length), and 6 mm (width), they were then fit to adapt to the buccal surface of the atrophied alveolar crest. One piece was placed to the right and one to the left side of the midline. On both sides fibrin glue was used (Tisseel, Baxter AG, Vienna, Austria) to stabilize the graft. After 6 months of graft healing, dental implants were placed, simultaneously biopsies were harvested and in one patient two oxidized microimplants were placed. At the time of abutment connection, microimplants were retrieved with surrounding bone for histology. Fixed screw-retained bridges were fabricated in mean of 7 months after implant surgery. Radiographs were taken before and after implant surgery and after 1 year of loading. Sixteen implants with an oxidized surface were placed (TiUnite, Nobel Biocare AB, Göteborg, Sweden). After 1 year of functional loading, all implants were clinically stable. The marginal bone loss was 1.4 mm (SD 0.3) after 1 year of loading. The histological examination showed resorption and subsequent bone formation on the allograft particles. There were no signs of inflammatory cell infiltration in conjunction with the allograft. The two microimplants showed bone formation directly on the implant surface. This study shows that morselized impacted bone allograft can be used to increase the width of the atrophied narrow alveolar crest as a good alternative to autogenous bone grafts in elderly patients. The histological examination of biopsies revealed a normal incorporation process and no signs of an immunological reaction. Further studies with larger samples are of important to be able to conclude if equal results can be obtained using morselized impacted bone allograft as for autogenous bone graft.

Imaging of Mesenchymal Stem Cell Transplant by Bioluminescence and PET

Dynamic measurements of infused stem cells generally require animal euthanasia for single-time-point determinations of engraftment. In this study, we used a triple-fusion reporter system for multimodal imaging to monitor human mesenchymal stem cell (hMSC) transplants. hMSCs were transduced with a triple-fusion reporter, fluc-mrfp-ttk (encoding firefly luciferase, monomeric red fluorescent protein, and truncated herpes simplex virus type 1 sr39 thymidine kinase) by use of a lentiviral vector. Transduced cells were assayed in vitro for the expression of each functional component of the triple-fusion reporter. Transduced and control hMSCs were compared for their potential to differentiate into bone, cartilage, and fat. hMSCs expressing the reporter were then loaded into porous, fibronectin-coated ceramic cubes and subcutaneously implanted into NOD-SCID mice along with cubes that were loaded with wild-type hMSCs and empty cubes. Mice were imaged repeatedly over 3 mo by bioluminescence imaging (BLI), and selected animals underwent CT and PET imaging. Osteogenic, adipogenic, and chondrogenic potential assays revealed retained differentiation potentials between transduced and wild-type hMSCs. Signals from the cubes loaded with reporter-transduced hMSCs were visible by BLI over 3 mo. There was no signal from the empty or wild-type hMSC-loaded control cubes. PET data provided confirmation of the quantitative estimation of the number of cells at one spot (cube). Cubes were removed from some animals, and histologic evaluations showed bone formation in cubes loaded with either reporter-transduced or wild-type hMSCs, whereas empty controls were negative for bone formation. The triple-fusion reporter approach resulted in a reliable method of labeling stem cells for investigation in small-animal models by use of both BLI and small-animal PET imaging. It has the potential for translation into future human studies with clinical PET.