Significant Difference In Bone Formation Research Articles

In vivo evaluation of bone regeneration behavior of novel β-tricalcium phosphate/layered double hydroxide nanocomposite granule as bone graft substitutes.

This study was based on in vivo assessment of bone regeneration capacity of synthesized porous β-tricalcium phosphate (β-TCP) nanocomposite granules and aimed to explore the effects of fabricated β-TCP granules reinforced with layered double hydroxides (LDH) nanoclay compared to β-TCP granules, in terms of osteoconductivity and biodegradability. Granules with diameters of 2-3 mm were implanted into cavities drilled in rabbit distal femur and were left in situ for up to 3months. The mechanical study demonstrated that the presence of LDH nanoparticles in β-TCP granules resulted in a significant increase in compressive modulus from 174.4 to 231.4MPa, while the porosity was constant at 76%-80%. The results revealed that the obtained granules showed no cytotoxicity. In this study, x-ray radiographic, micro-computed tomography, and histological staining analysis were taken to evaluate the percentage of bone ingrowth and biodegradability of the porous granules. The results exhibited that both granules support bone regeneration and also the amount of new bone formation in the bone defect filled with both granules was almost six times higher than the empty defects. Although no significant difference in bone formation for two different granules was observed, a higher biodegradability was detected in β-TCP granules in comparison to β-TCP/LDH granules. Overall, the addition of LDH nanoclay (10%) enhanced the physicochemical and mechanical properties of β-TCP granules while it is biological and osteoconductity properties have been maintained and its biodegradation rate has been decreased.

Biological Evaluation of a New Sodium-Potassium Silico-Phosphate Glass for Bone Regeneration: In Vitro and In Vivo Studies.

In vitro and in vivo studies are fundamental steps in the characterization of new implantable materials to preliminarily assess their biological response. The present study reports the in vitro and in vivo characterizations of a novel experimental silicate bioactive glass (BG) (47.5B, 47.5SiO2-10Na2O-10K2O-10MgO-20CaO-2.5P2O5 mol.%). Cytocompatibility tests were performed using human mature osteoblasts (U2OS), human mesenchymal stem cells (hMSCs) and human endothelial cells (EA.hy926). The release of the early osteogenic alkaline phosphatase (ALP) marker suggested strong pro-osteogenic properties, as the amount was comparable between hMSCs cultivated onto BG surface and cells cultivated onto polystyrene control. Similarly, real-time PCR revealed that the osteogenic collagen I gene was overexpressed in cells cultivated onto BG surface without biochemical induction. Acute toxicity tests for the determination of the median lethal dose (LD50) allowed classifying the analyzed material as a slightly toxic substance with LD50 = 4522 ± 248 mg/kg. A statistically significant difference in bone formation was observed in vivo through comparing the control (untreated) group and the experimental one, proving a clear osteogenic effect induced by the implantation at the defect site. Complete resorption of 47.5B powder was observed after only 3 months in favor of newly formed tissue, thus confirming the high osteostimulatory potential of 47.5B glass.

The sole and combined effects of gelatin-thrombin matrix and freeze-dried bone allograft on early bone healing.

Gelatin-thrombin matrix (GTM) is a hemostatic agent with applications in maxillofacial surgery consisting of human-derived thrombin and bovine-derived gelatin matrix. The aim of this study was to evaluate the efficacy of GTM alone or with freeze-dried bone allograft (FDBA) in improving early bone healing. Forty-six adult male Sprague-Dawley rats were used. All animals were randomly assigned to a control group (n = 4) and three study groups (n = 14). Each group was divided into two subgroups for histomorphometric and histological analyses at weeks 1 and 4. The new bone formation, inflammation, fibrosis, necrosis, foreign body reaction, and bone healing scores were evaluated based on the histopathological findings. Multiple comparisons were performed using the Kruskal-Wallis test. Variables that were not normally distributed were evaluated using the Mann-Whitney U test. At 1 week, the GTM + FDBA group showed less bone formation (mean ± SD: 0.08 ± 0.03 mm2), compared with the study and control groups (FDBA: 0.15 ± 0.06 mm2; GTM: 0.13 ± 0.06 mm2). At 4 weeks, the GTM group (0.48 ± 0.1 mm2) showed more bone formation than the GTM + FDBA group (0.33 ± 0.17 mm2). Foreign body reactions were observed at weeks 1 and 4 in all GTM-containing groups. Within the limitations of this study, GTM group did not show a significant difference in bone formation compared with the control group. GTM did not inhibit bone healing at 1 and 4 weeks, and no significant difference was observed compared with the control groups. GTM was more effective for bone healing when administered without FDBA. GTM is considered safe when bone hemorrhage is encountered.

3D-printed Biomimetic Bioactive Glass Scaffolds for Bone Regeneration in Rat Calvarial Defects.

The pore geometry of scaffold intended for the use in the bone repair or replacement is one of the most important parameters in bone tissue engineering. It affects not only the mechanical properties of the scaffold but also the amount of bone regeneration after implantation. Scaffolds with five different architectures (cubic, spherical, x, gyroid, and diamond) at different porosities were fabricated with bioactive borate glass using the selective laser sintering (SLS) process. The compressive strength of scaffolds with porosities ranging from 60% to 30% varied from 1.7 to 15.5 MPa. The scaffold’s compressive strength decreased significantly (up to 90%) after 1-week immersion in simulated body fluids. Degradation of scaffolds is dependent on porosity, in which the scaffold with the largest surface area has the largest reduction in strength. Scaffolds with traditional cubic architecture and biomimetic diamond architecture were implanted in 4.6 mm diameter full-thickness rat calvarial defects for 6 weeks to evaluate the bone regeneration with or without bone morphogenetic protein 2 (BMP-2). Histological analysis indicated no significant difference in bone formation in the defects treated with the two different architectures. However, the defects treated with the diamond architecture scaffolds had more fibrous tissue formation and thus have the potential for faster bone formation. Overall, the results indicated that borate glass scaffolds fabricated using the SLS process have the potential for bone repair and the addition of BMP-2 significantly improves bone regeneration.

Evaluation of immediate cell viability and repair of osteotomies for implants using drills and piezosurgery. A randomized, prospective, and controlled rabbit study.

The objective of this study was to evaluate the immediate bone damage and bone repair after osteotomy for implants using conventional drills and drills powered by piezoelectric motor. Twelve rabbits received a tibial osteotomy with a conventional drill (control), with a diamond like carbon drill (DLC) and with a piezoelectric drill (Piezo). The animals were euthanized immediately, 30 and 60 days postoperatively. The tibias were removed and analyzed by means of histomorphometry, immunohistochemistry and microtomography. The immediate damage to the tissue at the cavity margins was similar (P > .05). At 30 and 60 days, the amount of newly formed bone tissue was similar (P > .05). Osteocalcin was intense score at 60 days in Piezo group. Microtomography revealed that bone volume at 30 days (control 3.8; DLC 4.3; and Piezo 2.4) and 60 days (control 4.9; DLC 4.82; and Piezo 3.95) were similar. There was a significant difference in bone formation between 30 (2.4) and 60 days (3.95) for the Piezo group (P = .016). The immediate effects and repair of cavities made using conventional, DLC coated, or ultrasound drills were similar.

Guided bone regeneration with polypropylene barrier in rabbit's calvaria: A preliminary experimental study

ObjectivesThis study aimed to evaluate the bone formation process in experimental defects created on rabbit calvarial, in which one of the bone defects was covered by the impermeable membrane before suturing the skin flap, while the other was closed only by the cutaneous flap. The experimental holes were filled only by the blood clot. Material and methodsSixteen New Zealand female rabbits weighing between 3.5 and 4 kg were used. Two experimental bone defects were made in the rabbit calvarial. The holes were filled only with the blood clot and one of them was covered with an impermeable polypropylene membrane. A histological analysis was made at 21 and 42 days following the surgery. Histological evaluation consisted of the following: 1. inflammatory process; 2. Bone repair; 3. Bone remodeling; 4. Presence of osteoid matrix and mineralization, and 5. Formation of hematopoietic tissue. Each characteristic was analyzed semi quantitatively. ResultsThere was a statistical difference between the test and the control group at 21 days of healing in the following items: presence of cementation line (p = 0.012), presence of osteoid tissue (p = 0.012), and trabecular bone tissue development and mineralization (p = 0.012). A greater amount of lamellar bone tissue (mature) was also observed in the test group compared to the control group. ConclusionThe semiquantitative analysis showed that at 21 days there was a superiority of the repair process in the test group; at 42 days there was no significant difference in bone formation between the two groups; and that the polypropylene membrane is feasible to be used in GBR. Clinical significanceThe impermeable polypropylene barrier is feasible for use in the guided bone regeneration technique. It can be used only on the blood clot, without the need for grafting, and can be easily removed a few days after surgery. These results are unprecedented.

The Use of Tooth Particles as a Biomaterial in Post-Extraction Sockets. Experimental Study in Dogs.

Objectives: The objective of this study was to evaluate new bone formation derived from freshly crushed extracted teeth, grafted immediately in post-extraction sites in an animal model, compared with sites without graft filling, evaluated at 30 and 90 days. Material and Methods: The bilateral premolars P2, P3, P4 and the first mandibular molar were extracted atraumatically from six Beagle dogs. The clean, dry teeth were ground immediately using the Smart Dentin Grinder. The tooth particles obtained were subsequently sieved through a special sorting filter into two compartments; the upper container isolating particles over 1200 μm, the lower container isolated particles over 300 μm. The crushed teeth were grafted into the post-extraction sockets at P3, P4 and M1 (test group) (larger and smaller post-extraction alveoli), while P2 sites were left unfilled and acted as a control group. Tissue healing and bone formation were evaluated by histological and histomorphometric analysis after 30 and 90 days. Results: At 30 days, test site bone formation was greater in the test group than the control group (p < 0.05); less immature bone was observed in the test group (25.71%) than the control group (55.98%). At 90 days, significant differences in bone formation were found with more in the test group than the control group. No significant differences were found in new bone formation when comparing the small and large alveoli post-extraction sites. Conclusions: Tooth particles extracted from dog’s teeth, grafted immediately after extractions can be considered a suitable biomaterial for socket preservation.

Effects of low-level laser therapy and platelet concentrate on bone repair: Histological, histomorphometric, immunohistochemical, and radiographic study

PurposeLow-level laser therapy (LLLT) can increase bone metabolism, cell proliferation, and maturation, and reduce inflammation, while platelet concentrate (PC) assists bone healing process by releasing proteins and growth factors. Here, we evaluated the efficacy of combined LLLT and PC therapy in the healing of critical-size bone defects. Materials and methodsCalvarial critical-size defects 5 mm in diameter were made in 48 Wistar rats. Bones were removed, milled, and used as autogenous bone grafts. Animals were randomized into four groups: LP (LLLT + PC), PC, L (LLLT), and C (control, autogenous graft only). Animals were sacrificed at day 30 and 60 post-surgery. Specimens were submitted to radiographic (digital and conventional), histological, histomorphometric, and immunohistochemical analyses. ResultsDigital radiography was shown to be a better image analysis method compared with conventional radiography. Histological analysis demonstrated a significant difference in bone formation between animals in group L (p = 0.049) at day 60 than in other groups. Higher rates of inflammatory infiltrates and fibrosis were observed in the LP and PC groups at days 30 and 60, whereas the groups not receiving PC showed a higher rate of bone maturity. The inflammatory processes were reduced in the animals in the L group, together with new bone formation and maturation. Groups L and C had higher scores of positive osteocalcin immunostaining in bone and extracellular matrix. ConclusionsLLLT reduces inflammation and contributes to increased bone formation. PC treatment was shown to maintain connective tissue and to induce fibrosis during bone repair. Combined LLLT and PC treatment did not improve bone repair.

Regeneration of the Osteochondral Defect by a Wollastonite and Macroporous Fibrin Biphasic Scaffold.

Osteochondral defect refers to the damage of cartilage as well as subchondral bone. Cartilage tissue engineering focusing on the regeneration of cartilage and disregarding the subchondral bone always leads to partial regeneration of the damage, resulting in poor mechanical and physiological properties. A scaffold suitable for in situ inductive regeneration of both types of tissues is urgently needed. In this study, a biphasic scaffold integrated by macro-porous fibrin and 3D-printed wollastonite (containing 8% MgSiO3 (CS-Mg8)) scaffolds, either preloaded with rabbit bone marrow mesenchymal stem cells (BMSCs) or not, were fabricated and used to repair osteochondral defects in vivo (full thickness osteochondral defects in rabbits, 4 mm in diameter and 4 mm in depth with bone marrow blood effusion). The fibrin scaffold had a pore size of 100-200 μm, and was degraded gradually and reached weight loss over 80% at 28 days. The presence of BMSCs could accelerate the degradation rate. BMSCs could well proliferate in the fibrin scaffold along with time prolongation. The CS-Mg8 scaffold possessed a regular structure of cross stacked CS-Mg8 rods, and was degraded rather slowly with a mass loss of 8.5% at 28 days. BMSCs adhered and showed well spreading on the CS-Mg8 scaffold, without apparent proliferation in vitro. In vivo transplantation of the biphasic scaffolds, either preloaded with BMSCs or not, could induce the regeneration of both cartilage and subchondral bone to a great extent. Loading of BMSCs enabled better regeneration of cartilage layer, leading to smoother macroscopic appearance, good integrity with surrounding tissue and tide mark formation. However, no significant difference in bone formation and in gene expression was found with and without BMSCs loading.

Local Administration of Bisphosphonate-soaked Hydroxyapatite for the Treatment of Osteonecrosis of the Femoral Head in Rabbit.

Background:Systemic administration of bisphosphonates has shown promising results in the treatment of osteonecrosis of the femoral head (ONFH). However, few studies have evaluated the efficacy of local zoledronate (ZOL) administration in the treatment of ONFH. The purpose of this study was to investigate whether local administration of bisphosphonate-soaked hydroxyapatite (HA) could improve bone healing in an experimental rabbit model of ONFH.Methods:This experimental study was conducted between October 2014 and June 2015. Forty-five rabbits underwent simulated ONFH surgery. Immediately following surgery, they were divided into three groups: model (untreated, n = 15), HA (treated with HA alone, n = 15), and HA + ZOL (treated with HA soaked in a low-dose ZOL solution, n = 15). Histological, immunohistochemical, and quantitative analyses were performed to evaluate bone formation and resorption 2, 4, and 8 weeks after surgery.Results:Gross bone matrix and hematopoietic tissue formation were observed in the HA + ZOL group 4 weeks after surgery. The immunohistochemical staining intensities for 5-bromodeoxyuridine, runt-related transcription factor 2, osteocalcin, osteopontin, and osteoprotegerin were significantly higher in the HA + ZOL group than that in the model (P < 0.001, P < 0.001, P < 0.001, P < 0.001, and P = 0.018, respectively) and HA groups (P = 0.003, P = 0.049, P < 0.001, P = 0.020, and P = 0.019, respectively), whereas receptor activator of the nuclear factor-κB ligand staining intensity was significantly lower in the HA + ZOL group than that in the model and HA groups (P = 0.029 and P = 0.015, respectively) 4 weeks after surgery. No significant differences in bone formation or bone resorption marker expression were found between the three groups 2 or 8 weeks after surgery (P > 0.05).Conclusions:Local administration of HA soaked in a low-dose ZOL solution increased new bone formation while inhibiting bone resorption in an animal model of ONFH, which might provide new evidence for joint-preserving surgery in the treatment of ONFH.

Obesity does not affect the healing of femur fractures in mice

Obesity is reported to be both protective and deleterious to bone. Lipotoxicity and inflammation might be responsible for bone loss through inhibition of osteoblasts and activation of osteoclasts. However, little is known whether obesity affects the process of fracture healing. Therefore, we studied the effect of high fat diet-induced (HFD) obesity on callus formation and bone remodelling in a closed femur fracture model in mice. Thirty-one mice were fed a diet containing 60kJ% fat (HFD) for a total of 20 weeks before fracture and during the entire postoperative observation period. Control mice (n=31) received a standard diet containing 10kJ% fat. Healing was analyzed using micro-CT, biomechanical, histomorphometrical, immunohistochemical, serum and protein biochemical analysis at 2 and 4 weeks after fracture. HFD-fed mice showed a higher body weight and increased serum concentrations of leptin and interleukin-6 compared to controls. Within the callus tissue Western blot analyses revealed a higher expression of transcription factor peroxisome proliferator-activated receptor y (PPARy) and a reduced expression of runt-related transcription factor 2 (RUNX2) and bone morphogenetic protein (BMP)-4. However, obesity did not affect the expression of BMP-2 and did not influence the receptor activator of nuclear factor κB (RANK)/RANK ligand/osteoprotegerin (OPG) pathway during fracture healing. Although the bones of HFD-fed animals showed an increased number of adipocytes within the bone marrow, HFD did not increase callus adiposity. In addition, radiological and histomorphometric analysis could also not detect significant differences in bone formation between HFD-fed animals and controls. Accordingly, HFD did not affect bending stiffness after 2 and 4 weeks of healing. These findings indicate that obesity does not affect femur fracture healing in mice.

A comparative study of zinc, magnesium, strontium-incorporated hydroxyapatite-coated titanium implants for osseointegration of osteopenic rats

Surface modification techniques have been applied to generate titanium implant surfaces that promote osseointegration for the implants in cementless arthroplasty. However, its effect is not sufficient for osteoporotic bone. Zinc (Zn), magnesium (Mg), and strontium (Sr) present a beneficial effect on bone growth, and positively affect bone regeneration. The aim of this study was to confirm the different effects of the fixation strength of Zn, Mg, Sr-substituted hydroxyapatite-coated (Zn-HA-coated, Mg-HA-coated, Sr-HA-coated) titanium implants via electrochemical deposition in the osteoporotic condition. Female Sprague–Dawley rats were used for this study. Twelve weeks after bilateral ovariectomy, all animals were randomly divided into four groups: group HA; group Zn-HA; group Mg-HA and group Sr-HA. Afterwards, all rats from groups HA, Zn-HA, Mg-HA and Sr-HA received implants with hydroxyapatite containing 0%, 10% Zn ions, 10% Mg ions, and 10% Sr ions. Implants were inserted bilaterally in all animals until death at 12weeks. The bilateral femurs of rats were harvested for evaluation. All treatment groups increased new bone formation around the surface of titanium rods and push-out force; group Sr-HA showed the strongest effects on new bone formation and biomechanical strength. Additionally, there are significant differences in bone formation and push-out force was observed between groups Zn-HA and Mg-HA. This finding suggests that Zn, Mg, Sr-substituted hydroxyapatite coatings can improve implant osseointegration, and the 10% Sr coating exhibited the best properties for implant osseointegration among the tested coatings in osteoporosis rats.

Novel bone substitute material in alveolar bone healing following tooth extraction: an experimental study in sheep.

Electrospun cotton wool-like nanocomposite (ECWN) is a novel synthetic bone substitute that incorporates amorphous calcium phosphate nanoparticles into a biodegradable synthetic copolymer poly(lactide-co-glycolide). The objectives of this study were to develop a tooth extraction socket model in sheep for bone graft research and to compare ECWN and bovine-derived xenograft (BX) in this model. Sixteen cross-bred female sheep were used. Bilateral mandibular premolars were extracted atraumatically. Second and third premolar sockets were filled (Latin-square allocation) with BX, ECWN or left unfilled. Resorbable collagen membranes were placed over BX and selected ECWN grafted sockets. Eight sheep per time period were sacrificed after 8 and 16weeks. Resin-embedded undemineralised sections were analysed for descriptive histology and histomorphometric analyses. At 8weeks, there were with no distinct differences in healing among the different sites. At 16weeks, osseous healing followed a fine trabecular pattern in ECWN sites. Non-grafted sites showed thick trabeculae separated by large areas of fibrovascular connective tissue. In BX grafted sites, xenograft particles were surrounded by newly formed bone or fibrovascular connective tissue. There were no statistically significant differences in bone formation across the four groups. However, ECWN sites had significantly less residual graft material than BX sites at 16weeks (P=0.048). This first description of a tooth extraction socket model in sheep supports the utility of this model for bone graft research. The results of this study suggested that the novel material ECWN did not impede bone ingrowth into sockets and showed evidence of material resorption.

The effects of combined low level laser therapy and mesenchymal stem cells on bone regeneration in rabbit calvarial defects

ObjectiveThis study evaluated the effect of Low Level Laser Therapy (LLLT) and Mesenchymal Stem Cells (MSCs) on bone regeneration. Background dataAlthough several studies evaluated the effects of MSCs and LLLT, there is little information available regarding in vivo application of LLLT in conjunction with MSCs. MethodsForty-eight circular bone defects (6mm in diameter) were prepared in the calvaria of 12 New-Zealand white rabbits. The defects of each animal were randomly assigned to 4 groups: (C) no treatment; (L) applying LLLT; (SC) filled with MSCs; (SCL) application of both MSCs and LLLT. LLL was applied on alternate days at wavelength of 810nm, power density of 0.2W/cm2 and a fluency of 4J/cm2 using a Gallium–Aluminum–Arsenide (GaAlAs) diode laser. The animals were sacrificed after 3weeks and then histological samples were evaluated to determine the amount of new bone formation and the remaining scaffold and inflammation. ResultsThe histological evaluation showed a statistically significant increase in new bone formation of LLLT group relative to the control and the other two experimental groups (p<0.05). There was no significant difference in bone formation of the control group compared to experimental groups filled with MSCs. Laser irradiation had no significant effect on resorption of the scaffold material. In addition, inflammation was significantly reduced in LLLT group compared to the control defects and the other two experimental groups. ConclusionLow level laser therapy could be effective in bone regeneration but there is no evidence of a synergistic effect when applied in conjunction with MSCs.

Systemic vitamin D supplementation and local bone formation after maxillary sinus augmentation - a randomized, double-blind, placebo-controlledclinical investigation.

Maxillary sinus augmentation procedures with bone replacement grafts aimed to increase bone height in the posterior maxilla. During healing, bone particles are partially resorbed and replaced by the patient's own bone. Vitamin D plays an essential role in calcium homeostasis and is critical for bone formation and remodeling. This randomized, double-blind, placebo-controlled clinical investigation studied whether oral supplementation with vitamin D3 (5000 IU) combined with calcium (600mg) impacts bone formation and remodeling after maxillary sinus augmentation compared to a placebo medication containing calcium alone (n=10/group). Bone cores were harvested at the time of implant placement (6-8months) for histological analysis. Serum 25-hydroxyvitamin D (25-OHD) levels were comparable between both groups at the baseline (P=nonsignificant [n.s.]). Vitamin D3+ calcium supplementation improved significantly serum 25-OHD levels (placebo vs. vitamin D3 group: 25-OHD ng/ml: 31.13±7.06 vs. 61.11±20.42, P≤0.01); however, no statistically significant difference in bone formation or graft resorption was detected between groups. However, in the vitamin D3 group, a significant association was found between increased vitamin D levels and number of bone-resorbing osteoclasts around graft particles suggesting that local bone remodeling might be more pronounced when serum vitamin D levels were improved (r=0.92, P≤0.05). Vitamin D3+ calcium supplementation improves serum vitamin D levels and potentially impacts local bone remodeling on a cellular level. However, no statistically significant difference in bone formation or graft resorption was detected between groups.

Biologic Response to Titanium Implants with Laser-Treated Surfaces

To examine the biologic response to titanium implant surfaces treated with a neodymium:yttrium-aluminum-garnet laser. Sixty mini-implants made of grade 2 titanium were placed in the femora of 30 Wistar rats. Thirty implants had a machined surface and the other 30 had surfaces that were roughened by laser treatment. The animals were subdivided into three groups according to bone repair periods of 15, 30, and 60 days. The samples were observed under light and electron scanning microscopes and analyzed with the Student t test. Formation of new bone trabeculae toward the surface was apparent for the laser-treated implants at 15 days. Thin layers of bone matrix in intimate contact with the surface in the area of the central screw threads were observed, indicating high biocompatibility. Similar results were seen with machined implants after 30 days. A significant difference in bone formation was observed between the implant types at 15 days. Bone-to-implant contact was better on the surfaces subjected to laser treatment than on the machined titanium implants. The development of new laser treatments, which promote alterations in the surface energy as well as in the macro- and microstructures of titanium, may lead to improved bone-to-implant contact and thus better outcomes.

Effect of cell‐seeded hydroxyapatite scaffolds on rabbit radius bone regeneration

Highly porous hydroxyapatite (HA) scaffolds were developed as bone graft substitutes using a template coating process, characterized, and seeded with bone marrow-derived mesenchymal stem cells (BMSCs). To test the hypothesis that cell-seeded HA scaffolds improve bone regeneration, HA scaffolds without cell seeding (HA-empty), HA scaffolds with 1.5 × 10(4) BMSCs (HA-low), and HA scaffolds with 1.5 × 10(6) BMSCs (HA-high) were implanted in a 10-mm rabbit radius segmental defect model for 4 and 8 weeks. Three different fluorochromes were administered at 2, 4, and 6 weeks after implantation to identify differences in temporal bone growth patterns. It was observed from fluorescence histomorphometry analyses that an increased rate of bone infiltration occurred from 0 to 2 weeks (p < 0.05) of implantation for the HA-high group (2.9 ± 0.5 mm) as compared with HA-empty (1.8 ± 0.8 mm) and HA-low (1.3 ± 0.2 mm) groups. No significant differences in bone formation within the scaffold or callus formation was observed between all groups after 4 weeks, with a significant increase in bone regenerated for all groups from 4 to 8 weeks (28.4% across groups). Although there was no difference in bone formation within scaffolds, callus formation was significantly higher in HA-empty scaffolds (100.9 ± 14.1 mm(3) ) when compared with HA-low (57.8 ± 7.3 mm(3) ; p ≤ 0.003) and HA-high (69.2 ± 10.4 mm(3) ; p ≤ 0.02) after 8 weeks. These data highlight the need for a better understanding of the parameters critical to the success of cell-seeded HA scaffolds for bone regeneration.

Autologous vs. allogenic mesenchymal progenitor cells for the reconstruction of critical sized segmental tibial bone defects in aged sheep

Mesenchymal progenitor cells (MPCs) represent an attractive cell population for bone tissue engineering. Their special immunological characteristics suggest that MPCs may be used in allogenic applications. The objective of this study was to compare the regenerative potential of autologous vs. allogenic MPCs in an ovine critical size segmental defect model. Ovine MPCs were isolated from bone marrow aspirates, expanded and cultured with osteogenic medium for 2weeks before implantation. Autologous and allogenic transplantation was performed using the cell-seeded scaffolds and unloaded scaffolds, while the application of autologous bone grafts served as a control group (n=6). Bone healing was assessed 12weeks after surgery by radiology, microcomputed tomography, biomechanical testing and histology. Radiology, biomechanical testing and histology revealed no significant differences in bone formation between the autologous and allogenic groups. Both cell groups showed more bone formation than the scaffold alone, whereas the biomechanical data showed no significant differences between the cell groups and the unloaded scaffolds. The results of the study suggest that scaffold-based bone tissue engineering using allogenic cells offers the potential for an off-the-shelf product. Thus the results of this study serve as an important baseline for translation of the assessed concepts into clinical applications.

The Effect of Heparan Sulfate Application on Bone Formation during Distraction Osteogenesis

Bone morphogenetic proteins (BMPs) are recognized for their ability to induce bone formation in vivo and in vitro. Their osteogenic and osteoinductive properties are tightly regulated by the secretion of specific BMP antagonists, which have been shown to physically bind and sometimes be blocked by the extracellular proteoglycan heparan sulphate side chains (from hereon referred to as HS). The purpose of this study was to investigate if local application of 5 µg of HS proteoglycan to a bone regenerate site in a mouse model of distraction osteogenesis (DO) can accelerate bone healing and affect the expression of key members of the BMP signaling pathway. DO was performed on the right tibia of 115 adult male wild-type mice. At mid-distraction (day 11), half the group was injected locally with 5 µg of HS, while the other half was injected with saline. The mice were sacrificed at 2 time-points: mid-consolidation (34 days) and full consolidation (51 days). The distracted tibial zone was then collected for analysis by μCT, radiology, biomechanical testing, immunohistochemistry, and histology. While μCT data showed no statistically significant difference in bone formation, the results of biomechanical testing in stiffness and ultimate force were significantly lower in the HS-injected bones at 51 days, compared to controls. Immunohistochemistry results also suggested a decrease in expression of several key members of the BMP signaling pathway at 34 days. Furthermore, wound dehiscence and infection rates were significantly elevated in the HS group compared to the controls, which resulted in a higher rate of euthanasia in the treatment group. Our findings demonstrate that exogenous application of 5 µg of HS in the distracted gap of a murine model had a negative impact on bone and wound healing.

Mesenchymal Stem Cells and Platelet Gel Improve Bone Deposition within CAD-CAM Custom-Made Ceramic HA Scaffolds for Condyle Substitution

Purpose. This study evaluated the efficacy of a regenerative approach using mesenchymal stem cells (MSCs) and CAD-CAM customized pure and porous hydroxyapatite (HA) scaffolds to replace the temporomandibular joint (TMJ) condyle. Methods. Pure HA scaffolds with a 70% total porosity volume were prototyped using CAD-CAM technology to replace the two temporomandibular condyles (left and right) of the same animal. MSCs were derived from the aspirated iliac crest bone marrow, and platelets were obtained from the venous blood of the sheep. Custom-made surgical guides were created by direct metal laser sintering and were used to export the virtual planning of the bone cut lines into the surgical environment. Sheep were sacrificed 4 months postoperatively. The HA scaffolds were explanted, histological specimens were prepared, and histomorphometric analysis was performed. Results. Analysis of the porosity reduction for apposition of newly formed bone showed a statistically significant difference in bone formation between condyles loaded with MSC and condyles without (P < 0.05). The bone ingrowth (BI) relative values of split-mouth comparison (right versus left side) showed a significant difference between condyles with and without MSCs (P < 0.05). Analysis of the test and control sides in the same animal using a split-mouth study design was performed; the condyle with MSCs showed greater bone formation. Conclusion. The split-mouth design confirmed an increment of bone regeneration into the HA scaffold of up to 797% upon application of MSCs.