Rat Calvaria Defects Research Articles

Synthetic Bone Blocks Produced by Additive Manufacturing in the Repair of Critical Bone Defects.

This study evaluated the efficacy of synthetic bone blocks, composed of hydroxyapatite (HA) or β-tricalcium phosphate (B-TCP), which were produced by additive manufacturing and used for the repair of critical size bone defects (CSDs) in rat calvaria. Sixty rats were divided into five groups (n = 12): blood clot (CONTROL), 3D-printed HA (HA), 3D-printed β-TCP (B-TCP), 3D-printed HA + autologous micrograft (HA+RIG), and 3D-printed β-TCP + autologous micrograft (B-TCP+RIG). CSDs were surgically created in the parietal bone and treated with the respective biomaterials. The animals were euthanized at 30 and 60 days postsurgery for microcomputed tomography (micro-CT) histomorphometric, and immunohistochemical analysis to assess new bone formation. Micro-CT analysis showed that both biomaterials were incorporated into the animals' calvaria. The HA+RIG group, especially at 60 days, exhibited a significant increase in bone formation compared with the control. The use of 3D-printed bioceramics resulted in thinner trabeculae but a higher number of trabeculae compared with the control. Histomorphometric analysis showed bone islands in close contact with the B-TCP and HA blocks at 30 days. The HA blocks (HA and HA+RIG groups) showed statistically higher new bone formation values with further improvement when autologous micrografts were included. Immunohistochemical analysis showed the expression of bone repair proteins. At 30 days, the HA+RIG group had moderate Osteopontin (OPN) staining, indicating that the repair process had started, whereas other groups showed no staining. At 60 days, the HA+RIG group showed slight staining, similar to that of the control. Osteocalcin (OCN) staining, indicating osteoblastic activity, showed moderate expression in the HA and HA+RIG groups at 30 days, with slight expression in the B-TCP and B-TCP+RIG groups. The combination of HA blocks with autologous micrografts significantly enhanced bone repair, suggesting that the presence of progenitor cells and growth factors in the micrografts contributed to the improved outcomes. It was concluded that 3D-printed bone substitute blocks, associated with autologous micrografts, are highly effective in promoting bone repair in CSDs in rat calvaria.

Human versus Rat PRF on Collagen Membranes: A Pilot Study of Mineralization in Rat Calvaria Defect Model.

Platelet-rich fibrin, the coagulated plasma fraction of blood, is commonly used to support natural healing in clinical applications. The rat calvaria defect is a standardized model to study bone regeneration. It remains, however, unclear if the rat calvaria defect is appropriate to investigate the impact of human PRF (Platelet-Rich Fibrin) on bone regeneration. To this end, we soaked Bio-Gide® collagen membranes in human or rat liquid concentrated PRF before placing them onto 5 mm calvarial defects in Sprague Dawley rats. Three weeks later, histology and micro-computed tomography (μCT) were performed. We observed that the collagen membranes soaked with rat PRF show the characteristic features of new bone and areas of mineralized collagen matrix, indicated by a median mineralized volume of 1.5 mm3 (range: 0.9; 5.3 mm3). Histology revealed new bone growing underneath the membrane and hybrid bone where collagen fibers are embedded in the new bone. Moreover, areas of passive mineralization were observed. The collagen membranes soaked with human PRF, however, were devoid of histological features of new bone formation in the center of the defect; only occasionally, new bone formed at the defect margins. Human PRF (h-PRF) caused a median bone volume of 0.9 mm3 (range: 0.3-3.3 mm3), which was significantly lower than what was observed with rat PRF (r-PRF), with a BV median of 1.2 mm3 (range: 0.3-5.9 mm3). Our findings indicate that the rat calvaria defect model is suitable for assessing the effects of rat PRF on bone formation, but caution is warranted when extrapolating conclusions regarding the efficacy of human PRF.

Active and Passive Mineralization of Bio-Gide® Membranes in Rat Calvaria Defects.

Bio-Gide® is a collagen membrane routinely used in guided bone regeneration. Recent studies have shown that this collagen membrane has osteoconductive properties, meaning that it can support the growth of new bone. However, it has also been observed that the collagen membrane has areas of mineralized fibers which can occur spontaneously and independently of osteoblasts. To better understand how this works, we established a model using minced collagen membranes to reduce the active mineralization of intact collagen membranes in favor of passive mineralization. We thus compared the original intact membrane with a minced collagen membrane in a 5 mm calvarial defect model in Sprague Dawley rats. After three weeks of healing, histology and microcomputed tomography (μCT) were performed. Histological analysis confirmed the osteoconductive properties, with new bone growing inside the intact collagen membrane. However, in minced collagen membranes, the osteoconductive properties were restricted to the defect margins. Interestingly, histology revealed large mineralized areas indicating passive mineralization with no signs of bone formation. In the μCT analysis, the intact collagen membranes caused a higher median mineralized volume (1.5 mm3) compared with the minced group (0.4 mm3), but this lacked significance (p = 0.09). The μCT analysis needs to be interpreted carefully, particularly in defects filled with minced membranes, considering that the mineralized tissue may not necessarily be bone but also the result of passive mineralization. Taken together, the findings suggest that Bio-Gide® collagen membranes support bone formation while also exhibiting potential for passive mineralization.

Repair of a rat calvaria defect with injectable strontium (Sr)-doped polyphosphate dicalcium phosphate dehydrate (P-DCPD) ceramic bone grafts.

The trace element strontium (Sr) enhances new bone formation. However, delivering Sr, like other materials, in a sustained manner from a ceramic bone graft substitute (BGS) is difficult. We developed a novel ceramic BGS, polyphosphate dicalcium phosphate dehydrate (P-DCPD), which delivers embedded drugs in a sustained pattern. This study assessed the in vitro and in vivo performance of Sr-doped P-DCPD. In vitro P-DCPD and 10%Sr-P-DCPD were nontoxic and eluents from 10%Sr-P-DCPD significantly enhanced osteoblastic MC3T3 cell differentiation. A sustained, zero-order Sr release was observed from 10%Sr-P-DCPD for up to 70 days. When using this BGS in a rat calvaria defect model, both P-DCPD and 10% Sr-P-DCPD were found to be biocompatible and biodegradable. Histologic data from decalcified and undecalcified tissue showed that 10%Sr-P-DCPD had more extensive new bone formation compared with P-DCPD 12-weeks after surgery and the 10%Sr-P-DCPD had more organized new bone and much less fibrous tissue at the defect margins. The new bone was formed on the surface of the degraded ceramic debris within the bone defect area. P-DCPD represented a promising drug-eluting BGS for repair of critical bone defects.

Effect of local and systemic administration of atorvastatin for improving bone healing on critical defects.

This study aimed to evaluate the impact of atorvastatin, administered both locally and systemically, on critical defects in the calvaria of rats. Thirty-six adult rats were randomly assigned to three groups, with all bone defects covered by a collagen membrane. The groups received different treatments: distilled water (GAD), where membranes were soaked in distilled water; systemic application of atorvastatin (GAS) at a dosage of 3.6mg/kg/day through gavage; and local application of atorvastatin (GAL). After 14 and 28 days, all animals were euthanized, and various assessments were conducted, including histometric analysis, measurement of linear residual defect, evaluation of newly formed bone area, determination of membrane and soft tissue area, cell count, and immunohistochemical analysis. Group GAS exhibited a significant reduction in residual defect compared to the other groups (p<0.05) and a lower number of osteocytes (p<0.05) in comparison with other groups. On day 28, both GAL and GAS groups showed a higher number of inflammatory cells compared to GAD (p<0.05). Immunolabeling of CD31 was similar for both groups, but in the case of osteocalcin, there was a significant increase in labeling for groups GAS and GAL between days 14 and 28 postoperative (p<0.05). In conclusion, systemic atorvastatin demonstrated enhanced osteogenesis in critical calvaria defects in rats, suggesting its efficacy in promoting bone regeneration without exerting a notable anti-inflammatory effect.

Poly(lactic acid/caprolactone) bilayer membrane achieves bone regeneration through a prolonged barrier function.

Guided bone regeneration (GBR) is a treatment strategy used to recover bone volume. Barrier membranes are a key component of GBR protocols, and their properties can impact treatment outcomes. This study investigated the efficacy of an experimental, slow-degrading, bilayer barrier membrane for application in GBR using in vivo animal models. A synthetic copolymer of poly(lactic acid/caprolactone) (PLCL) was used to prepare a slow-degrading bilayer membrane. The biodegradability of PLCL was evaluated by subcutaneous implantation in a rat model. The barrier function of the PLCL membrane was investigated in a rat calvaria defect model and compared with commercially available membranes composed of type I collagen (Col) and poly(lactic-co-glycolic acid) (PLGA). An alveolar bone defect model in beagle dogs was used to simulate GBR protocols to evaluate the bone regeneration ability of the experimental PLCL membrane. The PLCL membrane showed slow biodegradation, resulting in an efficient and prolonged barrier function compared with commercial materials. In turn, this barrier function enabled the space-making ability of PLCL membrane and facilitated bone regeneration. In the alveolar bone defect model, significantly greater regeneration was achieved by treatment with PLCL membrane compared with Col and PLGA membranes. Additionally, a continuous alveolar ridge contour was observed in PLCL-treated bone defects. In conclusion, the PLCL bilayer membrane is a promising biomaterial for use in GBR given its slow degradation and prolonged barrier function.

Anti-Fibronectin Aptamer Modifies Blood Clot Pattern and Stimulates Osteogenesis: An Ex Vivo Study.

Scaffold (SCA) functionalization with aptamers (APT) provides adsorption of specific bioactive molecules on biomaterial surfaces. The aim of this study was to observe if SCA enriched with anti-fibronectin APT can favor coagulum (PhC) and osteoblasts (OSB) differentiation. 20 μg of APT was functionalized on SCA by simple adsorption. For PhC formation, SCAs were inserted into rat calvaria defects for 17 h. Following proper transportation (buffer solution PB), OSBs (UMR-106 lineage) were seeded over PhC + SCAs with and without APT. Cells and PhC morphology, PhC cell population, protein labeling and gene expression were observed in different time points. The APT induced higher alkaline phosphatase and bone sialoprotein immunolabeling in OSB. Mesenchymal stem cells, leukocytes and lymphocytes cells were detected more in the APT group than when scaffolds were not functionalized. Additionally, an enriched and dense fibrin network and different cell types were observed, with more OSB and white blood cells in PhC formed on SCA with APT. The gene expression showed higher transforming growth factor beta 1 (TGF-b1) detection in SCA with APT. The SCA functionalization with fibronectin aptamers may alter key morphological and functional features of blood clot formation, and provides a selective expression of proteins related to osteo differentiation. Additionally, aptamers increase TGF-b1 gene expression, which is highly associated with improvements in regenerative therapies.

Chondrocytes supplemented to bone graft-containing scaffolds expedite cranial defect repair

Critical maxillofacial bone fractures do not heal spontaneously, thus, often there is a need to facilitate repair via surgical intervention. Gold standard approaches, include the use of autologous bone graft, or devices supplemented with osteogenic growth factors and bone substitutes. This research aimed to employ a critical size calvaria defect model, to determine if the addition of chondrocytes to collagen-containing bone graft substitute, may expedite bone repair. As such, using a critical size rat calvaria defect, we implanted a collagen scaffold containing bone graft substitute (i.e., Bone graft scaffold, BG) or BG supplemented with costal chondrocytes (cBG). The rats were subjected to live CT imaging at 1, 6, 9, and 12 weeks following the surgical procedure and sacrificed for microCT imaging of the defect site. Moreover, serum markers and histological evaluation were assessed to determine osseous tissue regeneration and turnover. Live CT and microCT indicated cBG implants displayed expedited bone repair vs, BG alone, already at 6 weeks post defect induction. cBG also displayed a shorter distance between the defect edges and greater mineral apposition distance compared to BG. Summerizing, the data support the addition of chondrocytes to bone substitute, accelerates the formation of new bone within a critical size defect.

Effects of degradation products from gelatin spongy scaffolds on angio-osteogenic capacity

ABSTRACT The objective of this study was to determine whether degradation products from spongy gelatin scaffolds can enhance angiogenesis and orthotopic bone regeneration. Spongy gelatin disks were prepared using gelatin solution concentrations ranging from 1% to 7% (v/w) within cylindrical tubes through programmed freezing, lyophilization, cutting, and dehydrothermal crosslinking and implanted in critical-sized defects of rat calvaria for up to 8 weeks. Analyses of disk implantation into rat calvaria defects by microfocus X-ray computed tomography and histomorphometry indicated that the bone volume was significantly larger in the 5% and 7% gelatin sponge groups than in the 1% and 3% gelatin sponge groups and tended to increase progressively from 5% to 7%. The histomorphometric analysis also showed that the largest number of new vessels was formed in the defect treated with 5% gelatin sponge compared to other gelatin sponges. Immunohistochemistry of matrix metalloproteinase (MMP) indicated that the gelatin concentration in the disks affected the appearance of MMP-2 and MMP-9 positive cells around the skeleton of the gelatin sponges. The degradation products of gelatin by MMP-9 and prolyl endopeptidase enhanced the formation of a capillary-like structure in human umbilical vein endothelial cells in vitro. These results suggested that the higher-density gelatin sponges tended to supply their own molecules via biodegradation, resulting in enhanced orthotopic osteogenesis through the expected function of gelatin molecules in angiogenesis and bone formation.

Effects of platelet-rich fibrin produced by three centrifugation protocols on bone neoformation in defects created in rat calvaria

This study evaluated the potential of Leukocyte-platelet-rich fibrin (L-PRF; fixed angle centrifugation protocol), Advanced-platelet-rich fibrin (A-PRF; low-speed fixed angle centrifugation protocol), and Horizontal-platelet-rich fibrin (H-PRF; horizontal centrifugation protocol) in bone neoformation in critical size defects (CSDs) in rat calvaria. Thirty-two rats were divided into groups: Control (C), L-PRF, A-PRF, and H-PRF. 5 mm diameter CSDs were created in the animals’ calvaria. Defects from group Control (C) were filled with blood clots, while defects from groups L-PRF, A-PRF, and H-PRF were filled with respective platelet-rich fibrin (PRF) membranes. L-PRF, A-PRF, and H-PRF were prepared from animal blood collection and specific centrifugation protocols. At 14 and 30 days, calcein (CA) and alizarin (AL) injections were performed, respectively. Animals were euthanized at 35 days. Microtomographic, laser confocal microscopy, and histomorphometric analyzes were performed. Data were statistically analyzed (ANOVA, Tukey, p < .05). L-PRF, A-PRF, and H-PRF groups showed higher values of bone volume (BV), newly formed bone area (NFBA), and precipitation of CA and AL than the C group (p < .05). The H-PRF group showed higher values of BV, number of trabeculae (Tb. N), NFBA, and higher precipitation of AL than the A-PRF and L-PRF groups (p < .05). Therefore, it can be concluded that: i) L-PRF, A-PRF, and H-PRF potentiate bone neoformation in CSDs in rat calvaria; ii) H-PRF demonstrated more biological potential for bone healing.

Lycopene enhances bone neoformation in calvaria bone defects of ovariectomized rats.

Osteoporosis can affect a significant part of the population and fractures are the most common complications associated with this disease, leading to high public health costs. Thus, the prevention of fractures is relevant to individuals with signs and symptoms as well as to the health system. Postmenopausal osteoporosis has been associated with oxidative stress, emphasizing the importance of an efficient defense system to maintain bone health. Lycopene is a carotenoid with antioxidant properties that may stimulate osteoblastogenesis and inhibit osteoclastogenesis. The purpose of this investigation was to analyze the influence of lycopene in the bone neoformation of calvaria defects in ovariectomized rats utilizing the concentration of 45 mg/kg. Wistar Hannover female rats were divided into ovariectomized and sham groups. The ovariectomized animals received 45 mg/kg lycopene (OvxL) or water (Ovx) by daily gavage the day after ovariectomy/sham surgery for 16 weeks. Twelve weeks after ovariectomy, there were performed 5-mm calvaria defects followed by euthanasia after 4 weeks. Samples of bone tissue were collected to perform morphological and morphometrical analysis of the neoformed bone area, and percentage with Software Image J. Morphological evaluation showed mature bone with more osteocytes in the group OVxL when compared to the other groups. The morphometrical analysis demonstrated a significant increase of bone neoformation in the group OvxL (p<0.05). The data obtained suggest that lycopene benefits bone repair in the absence of estrogenic hormones.

Effectiveness of Apigenin, Resveratrol, and Curcumin as Adjuvant Nutraceuticals for Calvarial Bone Defect Healing: An In Vitro and Histological Study on Rats

Bone healing is a major clinical issue, especially in bone defects of critical dimensions. Some studies have reported in vivo positive effects on bone healing by some bioactive compounds, such as the phenolic derivatives found in vegetables and plants, such as resveratrol, curcumin, and apigenin. The aim of this work was (1) to analyze in vitro in human dental pulp stem cells the effects of these three natural compounds on the gene expression of related genes downstream to RUNX2 and SMAD5, key factor transcriptions associated with osteoblast differentiation, in order to better understand the positive effects that can occur in vivo in bone healing, and (2) to evaluate in vivo the effects on bone healing of critical-size defects in the calvaria in rats of these three nutraceuticals tested in parallel and for the first time administered by the gastric route. Upregulation of the RUNX2, SMAD5, COLL1, COLL4, and COLL5 genes in the presence of apigenin, curcumin, and resveratrol was detected. In vivo, apigenin induced more consistent significant bone healing in critical-size defects in rat calvaria compared to the other study groups. The study findings encourage a possible therapeutic supplementation with nutraceuticals during the bone regeneration process.

Collagen Membranes Functionalized with 150 Cycles of Atomic Layer Deposited Titania Improve Osteopromotive Property in Critical-Size Defects Created on Rat Calvaria.

The membranes used in bone reconstructions have been the object of investigation in the field of tissue engineering, seeking to improve their mechanical strength and add other properties, mainly the osteopromotive. This study aimed to evaluate the functionalization of collagen membranes, with atomic layer deposition of TiO2 on the bone repair of critical defects in rat calvaria and subcutaneous biocompatibility. A total of 39 male rats were randomized into four groups: blood clot (BC), collagen membrane (COL), COL 150-150 cycles of titania, and COL 600-600 cycles of titania. The defects were created in each calvaria (5 mm in diameter) and covered according to each group; the animals were euthanized at 7, 14, and 28 days. The collected samples were assessed by histometric (newly bone formed, soft tissue area, membrane area, and residual linear defect) and histologic (inflammatory cells and blood cells count) analysis. All data were subjected to statistical analysis (p < 0.05). The COL150 group showed statistically significant differences compared to the other groups, mainly in the analysis of residual linear defects (1.5 ± 0.5 × 106 pixels/µm2 for COL 150, and around 1 ± 0.5 × 106 pixels/µm2 for the other groups) and newly formed bone (1500 ± 1200 pixels/µm for COL 150, and around 4000 pixels/µm for the others) (p < 0.05), demonstrating a better biological behavior in the chronology of defects repair. It is concluded that the collagen membrane functionalized by TiO2 over 150 cycles showed better bioactive potential in treating critical size defects in the rats' calvaria.

Polydioxanone Membrane Compared with Collagen Membrane for Bone Regeneration

Guided bone regeneration (GBR) is an approach that induces osteopromotion through the regenerative membranes. These barriers exhibit bioactive behavior and mechanical function. Polydioxanone is a synthetic option, already used in medicine and dentistry, with good results in bone regeneration. This study aimed to evaluate bone repair in critical defects in rat calvaria using a polydioxanone membrane (Plenum® Guide) compared with a commercially available collagen-based membrane (Bio-Gide®). The bone defects were filled with Plenum® Osshp, a synthetic bone graft, hydroxyapatite:β-tricalcium phosphate, 70:30%, Group PG (Plenum® Guide + Plenum® Osshp), and Group BG (Geistlich Bio-Gide® + Plenum® Osshp). The specimens were submitted to immunohistochemical (RUNX2 and OPN), gene expression (RUNX2, IBSP, and VEGF), histometric, and microtomography analyses after 07, 15, 30, and 60 days postoperative. PG group showed greater immunolabeling area for RUNX2 and OPN, higher gene expression of VEGF (3.15 ± 0.85), and IBSP (24.9 ± 0.59). However, there was no statistical difference between groups in the histometric analysis regarding the percentage of connective tissue PG (0.83 ± 0.45), BG (0.70 ± 0.34), neoformed bone PG (0.60 ± 0.4), BG (0.65 ± 0.51), and remaining biomaterial PG (0.84 ± 0.31), BG (0.91 ± 0.33). In addition, there was no statistical difference between groups by micro-CT analysis. The absorbable-synthetic membrane, Plenum® Guide, is an effective membrane for guided bone regeneration.

In vivo evaluation of a collagen membrane in bone neoformation: A morphological and histomorphometric study

ObjectiveGuided bone regeneration (GBR) is a technique that involves the placement of mechanical barriers to protect the blood clot, and create an isolated space to prevent competition from epithelial and connective tissues in bone augmentation treatments. Collagen membranes stand out from other materials available for performing regenerative surgeries, and are widely used because of their ability to promote cell adhesion and homeostasis, and their biocompatibility, ease of handling, and low immunogenicity. In this context, researchers have investigated xenogenic membranes/barriers that cost less and have slower resorption rates. The current study aimed to assess the osteogenic potential induced by a crosslinked, synthesized xenogenic membrane 100 µm thick when applied in vivo to critical defects in rat calvaria. Material and methodsCritical size defects were created in the calvaria of thirty male Wistar rats, and randomly divided into the following two groups: G1 – clot covered with a commercial xenogenic membrane (Lumina-Coat®, Criteria, Brazil), and G2 – clot covered with a synthesized xenogenic membrane. The animals were euthanized after 7, 15 and 30 days, and samples of calvaria were processed to perform morphometric evaluations to measure bone neoformation in the defect region. In addition, ultrastructural characterization of the collagen membranes was performed by scanning electron microscope. The quantitative analyses were carried out by adopting a significance level of 5%. ResultsThe ultrastructural characterization revealed that the synthesized membrane had thicker collagen fibers and a more cohesive surface, compared with the Lumina-Coat® collagen membrane (G1). There was no significant difference in bone neoformation between the membranes (p>0.05), at any of the time periods analyzed. The bone quantification area increased significantly over time for both membranes (p<0.05). ConclusionThe synthesized membrane exhibited morphological characteristics similar to those of the commercial membrane evaluated, allowed potentially active participation in the bone neoformation process, and served as a low-cost alternative for GBR procedures.

Comparative study of the effect of different temperatures on bovine bone used for bone repair of critical calvaria defects in rats.

To evaluate the osteoconductive potential of inorganic biomaterials of bovine origin submitted to different temperatures in the bone repair of critical defects in rat calvaria. Forty-eight rats were divided into four groups according to the material used to fill the defect: control group (GC), the defect was filled only with blood clot (n=12); GBO, defect filled with Bio-Oss®, deproteinzed at 300°C (n=12); GOX, defect filled with Inorganic GenOx®, deproteinzed from 850 to 1200°C (n=12) and G700, defect filled with Inorganic GenOx 700, deproteinzed at 700°C (n=12). In each animal's calvaria, a trephine bur with 5 mm internal diameter was used to produce a 6 mm-diameter central defect. Gen Derm® resorbable bovine membrane was superimposed over all defects. Subsequently, animals were euthanized at 30 and 60 days after surgery. The pieces were sent for histological and histometric analysis to evaluate the following variables: bone neoformation, presence of biomaterial, mononuclear and polymorphonuclear leukocytes, presence of other tissues (granulation and medullary) and maturation of collagen fibers. The most representative group for bone neoformation was GC. At 30 days, there was a higher mean of mature bone tissue (75.8). At 60 days, there was no statistical difference between the GC (64.9), GBO (32.9), GOX (45.3), and G700 (26.6) groups. GBO presented the highest amount of biomaterial after 30 days (115.9) and 60 days (118.5). All bovine biomaterials were biocompatible and osteoconductive. GOX promoted the best bone repair of the studied materials.

Screening of Hydroxyapatite Biomaterials for Alveolar Augmentation Using a Rat Calvaria Critical-Size Defect Model: Bone Formation/Maturation and Biomaterials Resolution.

Natural (bovine-/equine-/porcine-derived) or synthetic hydroxyapatite (HA) biomaterials appear to be the preferred technologies among clinicians for bone augmentation procedures in preparation for implant dentistry. The aim of this study was to screen candidate HA biomaterials intended for alveolar ridge augmentation relative to their potential to support local bone formation/maturation and to assess biomaterial resorption using a routine critical-size rat calvaria defect model. Eighty adult male Sprague Dawley outbred rats obtained from a approved-breeder, randomized into groups of ten, were used. The calvaria defects (ø8 mm) either received sham surgery (empty control), Bio-Oss (bovine HA/reference control), or candidate biomaterials including bovine HA (Cerabone, DirectOss, 403Z013), and bovine (403Z014) or synthetic HA/ß-TCP (Reprobone, Ceraball) constructs. An 8 wk healing interval was used to capture the biomaterials' resolution. All biomaterials displayed biocompatibility. Strict HA biomaterials showed limited, if any, signs of biodegradation/resorption, with the biomaterial area fraction ranging from 22% to 42%. Synthetic HA/ß-TCP constructs showed limited evidence of biodegradation/erosion (biomaterial area fraction ≈30%). Mean linear defect closure in the sham-surgery control approximated 40%. Mean linear defect closure for the Bio-Oss reference control approximated 18% compared with 15-35% for the candidate biomaterials without significant differences between the controls and candidate biomaterials. None of the candidate HA biomaterials supported local bone formation/maturation beyond the native regenerative potential of this rodent model, pointing to their limitations for regenerative procedures. Biocompatibility and biomaterial dimensional stability could suggest their potential utility as long-term defect fillers.

Effect of ozone therapy on the modulation of inflammation and on new bone formation in critical defects of rat calvaria filled with autogenous graft

ObjectiveThe aim of this study was to evaluate the effect of ozone therapy on new bone formation and inflammation modulation in defects of rat calvaria filled with autogenous bone. Material and methodsCritical size defects were created in the calvaria of 24 male Wistar rats. The animals were randomly divided into four groups according to the treatment: G1: clot; G2: clot and covered with xenogenic membrane; G3: particulate autogenous bone graft; G4: autogenous bone graft and application of 3 mL O2/O3 gas mixture (10 µg/ml). The defects were filled immediately after surgery with a bilateral retroauricular application, in the region immediately above the incision. After 21 days, the animals were euthanized, and the samples were processed for morphometric evaluations designed to measure both the intensity of the inflammatory infiltrate, and the presence of new bone formation in the defect. ResultsThe results showed a lower inflammation score and higher mean of newly formed bone in the region of the defect for the group associated with ozone therapy (G4). The bone formed in the region of the defect could be observed as being more lamellar and mineralized in the case of associated ozone therapy. ConclusionOzone therapy represents a promising adjuvant therapy to accelerate tissue regeneration.

Evaluation of a New Porcine Bone Graft on the Repair of Surgically Created Critical Bone Defects in Rat Calvaria: Histomorphometric and Microtomographic Study.

The aim of this study was to evaluate the use of a new porcine bone graft in rat calvaria bone defects. Critical defects were surgically created in 24 rats that were divided into four experimental groups according to defect filling (n = 6): Control Group (CG)—blood clot; Porcine Bone Group (PG)—porcine-derived bone substitute; (BG): Bio-Oss Group (BG)–chemically and heat-treated bovine graft; Bonefill Group (BFG)—chemically treated bovine bone substitute. Euthanasia of the animals occurred 30 days after the surgery, and the area of the original surgical defect and the surrounding tissues were removed for micro-CT and histomorphometric analysis. In the micro-CT evaluation, the PG presented statistically significant differences (p < 0.05) in comparison to the CG, BG and BFG, for the parameters percentage of Bone Volume (BV/TV), Surface Bone Density (BS/TV), Number of Trabeculae (Tb.N) and Bone Connectivity (Conn), but not for Total Porosity (Po.tot) and Trabecular Thickness (Tb.Th). The histomorphometric analysis showed that the PG presented similar results to the BG regarding newly formed bone extension and to the BG and BFG regarding newly formed bone area. The porcine-derived graft presented superior microtomographic and histomorphometric results when compared to the two bovine bone substitutes.

Diabetic bone regeneration with nanoceria-tailored scaffolds by recapitulating cellular microenvironment: Activating integrin/TGF-β co-signaling of MSCs while relieving oxidative stress

Regenerating defective bone in patients with diabetes mellitus remains a significant challenge due to high blood glucose level and oxidative stress. Here we aim to tackle this issue by means of a drug- and cell-free scaffolding approach. We found the nanoceria decorated on various types of scaffolds (fibrous or 3D-printed one; named nCe-scaffold) could render a therapeutic surface that can recapitulate the microenvironment: modulating oxidative stress while offering a nanotopological cue to regenerating cells. Mesenchymal stem cells (MSCs) recognized the nanoscale (tens of nm) topology of nCe-scaffolds, presenting highly upregulated curvature-sensing membrane protein, integrin set, and adhesion-related molecules. Osteogenic differentiation and mineralization were further significantly enhanced by the nCe-scaffolds. Of note, the stimulated osteogenic potential was identified to be through integrin-mediated TGF-β co-signaling activation. Such MSC-regulatory effects were proven in vivo by the accelerated bone formation in rat calvarium defect model. The nCe-scaffolds further exhibited profound enzymatic and catalytic potential, leading to effectively scavenging reactive oxygen species in vivo. When implanted in diabetic calvarium defect, nCe-scaffolds significantly enhanced early bone regeneration. We consider the currently-exploited nCe-scaffolds can be a promising drug- and cell-free therapeutic means to treat defective tissues like bone in diabetic conditions.