Critical-sized Calvarial Defects Research Articles

Sustained release of ancillary amounts of testosterone and alendronate from PLGA coated pericard membranes and implants to improve bone healing.

Testosterone and alendronate have been identified as two bone healing compounds which, when combined, synergistically stimulate bone regeneration. This study describes the development of a novel ultrasonic spray coating for sustained release of ancillary amounts of testosterone and alendronate encapsulated in PLGA 5004A as a carrier. Due to the low amounts of testosterone and alendronate used, sensitive in vitro assays were developed to determine in vitro release. The ultrasonic spray coating technology was optimized for coating titanium screws and pericardial collagen membranes, with the aim to improve osseo-integration and (guided) bone regeneration, respectively, without interfering with their primary mode of action. In vitro release analysis of collagen membranes and screws showed up to 21 days sustained release of the compounds without a burst release. Subsequent preclinical studies in rat and rabbit models indicated that testosterone and alendronate coated membranes and screws significantly improved bone regeneration in vivo. Coated membranes significantly improved the formation of new bone in a critical size calvarial defect model in rats (by 160% compared to controls). Coated screws implanted in rabbit femoral condyles significantly improved bone implant contact (69% vs 54% in controls), bone mineral density (121%) and bone volume (119%) up to 1.3 mm from the implant. Based on the results obtained, we suggest that implants or membranes enabled with local sustained delivery of ancillary amounts of testosterone and alendronate can be a promising system to stimulate local bone regeneration resulting in improved osseo-integration of implants and improved healing of bone defects and fractures.

Polyelectrolyte multilayer composite coating on 316 L stainless steel for controlled release of dual growth factors accelerating restoration of bone defects

A composite coating of polyelectrolyte multilayers (PEMs) consisting of collagen, a chitosan barrier, and poly-γ-glutamic acid was fabricated using a spin coating technique to investigate and overcome the limited osseointegration capacity of 316 L stainless steel (316 L SS). To further enhance the biocompatibility, bone morphogenetic protein 2 (BMP-2) and basic fibroblast growth factor-2 (FGF-2) were loaded separately as dual growth factors, allowing for progressive drug release following the natural process of bone regeneration. The first burst release of FGF-2 triggered the proliferation of surrounding cells, and the subsequent release of BMP-2 stimulated their differentiation. The microstructure, surface potential, hardness, reduced Young's modulus, and wettability were assessed using scanning electron microscopy, nanoindentation, and water contact angle. The formation of apatite layers after immersion in simulated body fluid confirmed the bioactivity of this PEM. PEMs loaded with BMP-2 and FGF-2 showed a long sustained release of growth factors for up to 48 days. The biological properties were studied in vitro with rat bone mesenchymal stem cells (rBMSCs) and in vivo using a rat critical-sized calvarial defect model. PEMs loaded with growth factors further stimulated the proliferation and osteogenic differentiation of rBMSCs and the histology results indicated that new bone tissues could directly grow onto the PEMs. These findings suggest that PEM composite coating possesses significant potential for surface modification and long-term drug release of metallic implants to assist with bone restoration.

Addition of lactoferrin and substance P in a chitin/PLGA-CaSO4 hydrogel for regeneration of calvarial bone defects

Calcium-based injectable hydrogels with various bioactive active molecules possess a great potential for bone regeneration. Herein, we have synthesized a chitin-PLGA-calcium sulfate hydrogel (CSG) containing bioactive molecules - lactoferrin (LF) and substance P (SP). SEM and XRD analysis revealed that CS crystal growth was altered with the addition of LF. Rheological measurements indicated that the injectability of the hydrogels was maintained after the addition of LF, however, there was a reduction in storage modulus after LF addition. The addition of LF increased stem cell proliferation whereas, SP enhanced the cell migration. Osteogenic gene expression revealed that LF concentration at 25 μg/mg of CSG was optimal for a favourable outcome. To this optimized LF containing CSG, SP was incorporated and 0.05 μg/mg was found to be most effective (CSG-L3S2) in vitro studies. Further, the μ-CT and histological studies confirmed that CSG-L3S2 showed enhanced bone regeneration compared to the controls in critical-sized calvarial defect of mice. Thus the results indicate that a combination of the chemotactic agent (SP), pleiotropic growth protein (LF), and CS in the chitin-PLGA hydrogel could be a promising approach for non-load bearing bone defects.

Effect of wheat gluten on improved thermal cross-linking and osteogenesis of hydroxyapatite-gelatin composite scaffolds

Promising strategies to stabilize gelatin or collagen include glutaraldehyde-based chemical cross-linking or dehydrothermal treatment at different temperatures (120–180 °C). However, these procedures require 24–48 h for complete cross-linking to occur. The present study aims to evaluate the role of wheat gluten on enhancing thermal cross-linking of silica-nanohydroxyapatite (nanoHA)-gelatin composite scaffolds within a shorter period (2 h). Changes in properties were evaluated by varying the ratio of gelatin and gluten in silica-nanoHA matrix (60 wt% ceramic: 40 wt% polymer). The results showed that the scaffolds cross-linked at 170 °C were stable in phosphate-buffered saline for 21 days. It was crystalline and porous in nature. However, the scaffolds with high weight percentage of wheat gluten were brittle, while those with low gluten degraded fast in vitro. The mesenchymal stem cells could adhere, proliferate and differentiate into osteogenic lineage on wheat gluten-containing scaffolds for 21 days (mainly medium concentration). The scaffold also supported new bone formation in critical-sized rat calvarial defect, showing its osteoconductive and osteointegrative nature. In short, this study showed the potential of wheat gluten on improving thermal cross-linking within a shorter period and its suitability to use as a biomimetic bone graft for bone tissue engineering.

Bone Regeneration Using PEVAV/β-Tricalcium Phosphate Composite Scaffolds in Standardized Calvarial Defects: Micro-Computed Tomographic Experiment in Rats.

Bone regeneration using beta-tricalcium phosphate (β-TCP) can be practiced using a biocomposite scaffold. Poly(ethylene-co-vinylalcohol)/poly(δ-valerolactone)/β-tricalcium phosphate (PEVAV/β-TCP) composite scaffolds showed promising in vitro results. This study evaluated the bone regenerative potential of PEVAV/β-TCP biocomposite scaffolds in standardized calvarial defects in a rat model over 4 and 10 weeks. Bilateral calvarial defects (5 mm in diameter and about 1.5 mm thick, equivalent to the thickness of the calvaria) were created in 40 male Wistar albino rats. The defects were grafted with either commercially available β-TCP (positive control), PEVAV/β-TCP 70, or PEVAV/β-TCP 50, or left empty (negative control), depending on the group to which the animal was randomly assigned, to be covered before flap closure with resorbable collagen membrane (RCM). At 4 and 10 weeks post-surgery, the collected rat calvaria were evaluated using micro computed tomography (micro-CT) analysis, to assess the newly formed bone volume (NFBV), newly formed bone mineral density (NFBMD), and remaining graft volume (RGV). The results showed that calvarial defects grafted with the PEVAV/β-TCP biocomposite exhibited higher NFBV than did control defects, both at 4 and 10 weeks post-surgery. Furthermore, calvarial defects grafted with PEVAV/β-TCP 70 showed the highest NFBV among all grafting conditions, with a statistically significant difference recorded at 10 weeks post-surgery. The PEVAV/β-TCP composite scaffold showed potentiality for the regeneration of critical-sized calvarial bone defects in a rat model.

Copper peptide-incorporated 3D-printed silk-based scaffolds promote vascularized bone regeneration

Bioactive ions are promising materials for vascularized bone regeneration. However, trace elements, e.g., copper, exert their effects in a narrow concentration range and cause cytotoxicity when over-accumulated, which limits their potential application in tissue engineering. In this regard, naturally occurring nanosized metallopeptides provide new insight into the novel usage of metal ions. Here, 3D-printed silk-based scaffolds with sustained release of copper peptide, a copper ion-specific binding tripeptide, were fabricated and exhibited similar therapeutic effects to free copper ions, with lower toxicity. We found that copper peptide promoted the polarization of M2 macrophages, which exhibited an anti-inflammatory phenotype and assisted in bone tissue repair during early stage. Moreover, the released copper peptide provided a microenvironment to stimulate transplanted bone marrow-derived stem cell (BMSC) proliferation and cytokine secretion. The secreted cytokines further promoted angiogenesis in vitro and in vivo. In addition, in vivo critical-sized calvarial defect regeneration experiments further demonstrated that fabricated scaffolds seeded with BMSC, resulting in better vascularized bone repair. Altogether, the results indicated that copper peptide incorporated silk-based scaffolds are promising materials for vascularized bone tissue regeneration, as well as providing new approaches for bioactive ion-based biomaterial design.

In situ bone regeneration with sequential delivery of aptamer and BMP2 from an ECM-based scaffold fabricated by cryogenic free-form extrusion

In situ tissue engineering is a powerful strategy for the treatment of bone defects. It could overcome the limitations of traditional bone tissue engineering, which typically involves extensive cell expansion steps, low cell survival rates upon transplantation, and a risk of immuno-rejection. Here, a porous scaffold polycaprolactone (PCL)/decellularized small intestine submucosa (SIS) was fabricated via cryogenic free-form extrusion, followed by surface modification with aptamer and PlGF-2123-144*-fused BMP2 (pBMP2). The two bioactive molecules were delivered sequentially. The aptamer Apt19s, which exhibited binding affinity to bone marrow-derived mesenchymal stem cells (BMSCs), was quickly released, facilitating the mobilization and recruitment of host BMSCs. BMP2 fused with a PlGF-2123-144 peptide, which showed “super-affinity” to the ECM matrix, was released in a slow and sustained manner, inducing BMSC osteogenic differentiation. In vitro results showed that the sequential release of PCL/SIS-pBMP2-Apt19s promoted cell migration, proliferation, alkaline phosphatase activity, and mRNA expression of osteogenesis-related genes. The in vivo results demonstrated that the sequential release system of PCL/SIS-pBMP2-Apt19s evidently increased bone formation in rat calvarial critical-sized defects compared to the sequential release system of PCL/SIS-BMP2-Apt19s. Thus, the novel delivery system shows potential as an ideal alternative for achieving cell-free scaffold-based bone regeneration in situ.

Influence of Age on Calvarial Critical Size Defect Dimensions: A Radiographic and Histological Study.

Calvarial critical-size defect has been used to assess techniques and materials in the bone regeneration field. Previous studies utilized young adult rats with 3 months of age, which might not reflect the geriatric conditions. This study aimed to assess the dimensions of the calvarial critical-size defect in aged rats.Seventy-two rats in a randomized block design were allocated into a control young adult (11–12 weeks), and a test old group (22–24 months). Both groups were divided according to bone defect's size: 3 mm, 5 mm, and 7 mm defects, which were surgically created and followed for 4 and 8 weeks. Radiographic and histologic analyses were performed. Based on the results, additional groups with 4 mm defect size were added following the same protocols. Young groups yielded higher bone volumes, defect closure percentages, and density of newly formed bone. Closure of cranial defects was only observed in 3 mm defects in both age groups after 8 weeks; however, the 4 mm defect group demonstrated bony bridging after 8 weeks in young but not old rats. Results confirmed that 5-mm defect is considered a critical size for calvarial bone defects in young adult rats; however, 4 mm defect might be considered critical size for the aged rats after 8 weeks.

Spongostan™ Leads to Increased Regeneration of a Rat Calvarial Critical Size Defect Compared to NanoBone® and Actifuse.

Bone substitute materials are becoming increasingly important in oral and maxillofacial surgery. Reconstruction of critical size bone defects is still challenging for surgeons. Here, we compared the clinically applied organic bone substitute materials NanoBone® (nanocrystalline hydroxyapatite and nanostructured silica gel; n = 5) and Actifuse (calcium phosphate with silicate substitution; n = 5) with natural collagen-based Spongostan™ (hardened pork gelatin containing formalin and lauryl alcohol; n = 5) in bilateral rat critical-size defects (5 mm diameter). On topological level, NanoBone is known to harbour nanopores of about 20 nm diameter, while Actifuse comprises micropores of 200–500 µm. Spongostan™, which is clinically applied as a haemostatic agent, combines in its wet form both nano- and microporous topological features by comprising 60.66 ± 24.48 μm micropores accompanied by nanopores of 32.97 ± 1.41 nm diameter. Micro-computed tomography (µCT) used for evaluation 30 days after surgery revealed a significant increase in bone volume by all three bone substitute materials in comparison to the untreated controls. Clearly visual was the closure of trepanation in all treated groups, but granular appearance of NanoBone® and Actifuse with less closure at the margins of the burr holes. In contrast, transplantion of Spongostan™ lead to complete filling of the burr hole with the highest bone volume of 7.98 ccm and the highest bone mineral density compared to all other groups. In summary, transplantation of Spongostan™ resulted in increased regeneration of a rat calvarial critical size defect compared to NanoBone and Actifuse, suggesting the distinct nano- and microtopography of wet Spongostan™ to account for this superior regenerative capacity. Since Spongostan™ is a clinically approved product used primarily for haemostasis, it may represent an interesting alternative in the reconstruction of defects in the maxillary region.

Sinusoidal electromagnetic fields accelerate bone regeneration by boosting the multifunctionality of bone marrow mesenchymal stem cells

BackgroundThe repair of critical-sized bone defects is always a challenging problem. Electromagnetic fields (EMFs), used as a physiotherapy for bone defects, have been suspected to cause potential hazards to human health due to the long-term exposure. To optimize the application of EMF while avoiding its adverse effects, a combination of EMF and tissue engineering techniques is critical. Furthermore, a deeper understanding of the mechanism of action of EMF will lead to better applications in the future.MethodsIn this research, bone marrow mesenchymal stem cells (BMSCs) seeded on 3D-printed scaffolds were treated with sinusoidal EMFs in vitro. Then, 5.5 mm critical-sized calvarial defects were created in rats, and the cell scaffolds were implanted into the defects. In addition, the molecular and cellular mechanisms by which EMFs regulate BMSCs were explored with various approaches to gain deeper insight into the effects of EMFs.ResultsThe cell scaffolds treated with EMF successfully accelerated the repair of critical-sized calvarial defects. Further studies revealed that EMF could not directly induce the differentiation of BMSCs but improved the sensitivity of BMSCs to BMP signals by upregulating the quantity of specific BMP (bone morphogenetic protein) receptors. Once these receptors receive BMP signals from the surrounding milieu, a cascade of reactions is initiated to promote osteogenic differentiation via the BMP/Smad signalling pathway. Moreover, the cytokines secreted by BMSCs treated with EMF can better facilitate angiogenesis and osteoimmunomodulation which play fundamental roles in bone regeneration.ConclusionIn summary, EMF can promote the osteogenic potential of BMSCs and enhance the paracrine function of BMSCs to facilitate bone regeneration. These findings highlight the profound impact of EMF on tissue engineering and provide a new strategy for the clinical treatment of bone defects.

Bovine-Derived Xenografts Immobilized With Cryopreserved Stem Cells From Human Adipose and Dental Pulp Tissues Promote Bone Regeneration: A Radiographic and Histological Study.

Adipose tissue-derived stem cells (ADSCs) and dental pulp stem cells (DPSCs) have become promising sources for bone tissue engineering. Our study aimed at evaluating bone regeneration potential of cryopreserved ADSCs and DPSCs combined with bovine-derived xenografts with 10% porcine collagen. In vitro studies revealed that although DPSCs had higher proliferative abilities, ADSCs exhibited greater mineral depositions and higher osteogenic-related gene expression, indicating better osteogenic differentiation potential of ADSCs. After applying cryopreserved ADSCs and DPSCs in a critical-sized calvarial defect model, both cryopreserved mesenchymal stem cells significantly improved bone volume density and new bone area at 2, 4, and 8 weeks. Furthermore, the combined treatment with ADSCs and xenografts was more efficient in enhancing bone repair processes compared to combined treatment with DPCSs at all-time points. We also evaluated the sequential early bone healing process both histologically and radiographically, confirming a high agreement between these two methods. Based on these results, we propose grafting of the tissue-engineered construct seeded with cryopreserved ADSCs as a useful strategy in accelerating bone healing processes.

Mesenchymal stem cells and three-dimensional-osteoconductive scaffold regenerate calvarial bone in critical size defects in swine.

Craniofacial bones protect vital organs, perform important physiological functions, and shape facial identity. Critical‐size defects (CSDs) in calvarial bones, which will not heal spontaneously, are caused by trauma, congenital defects, or tumor resections. They pose a great challenge for patients and physicians, and significantly compromise quality of life. Currently, calvarial CSDs are treated either by allogenic or autologous grafts, metal or other synthetic plates that are associated with considerable complications. While previous studies have explored tissue regeneration for calvarial defects, most have been done in small animal models with limited translational value. Here we define a swine calvarial CSD model and show a novel approach to regenerate high‐quality bone in these defects by combining mesenchymal stem cells (MSCs) with a three‐dimensional (3D)‐printed osteoconductive HA/TCP scaffold. Specifically, we have compared the performance of dental pulp neural crest MSCs (DPNCCs) to bone marrow aspirate (BMA) combined with a 3D‐printed HA/TCP scaffold to regenerate bone in a calvarial CSD (>7.0 cm2). Both DPNCCs and BMA loaded onto the 3D‐printed osteoconductive scaffold support the regeneration of calvarial bone with density, compression strength, and trabecular structures similar to native bone. Our study demonstrates a novel application of an original scaffold design combined with DPNCCs or BMA to support regeneration of high‐quality bone in a newly defined and clinically relevant swine calvarial CSD model. This discovery may have important impact on bone regeneration beyond the craniofacial region and will ultimately benefit patients who suffer from debilitating CSDs.

Xenogeneic bone mineral is efficient for the repair of critical-sized rat calvarial defects

Aim. To evaluate the efficiency of bone repair on a critical-sized rat calvarial defect model using our original xenogeneic bone mineral, widely established Geistlich Bio-Oss®, and autologous bone graft.Materials and Methods. We created a critical-sized calvarial defect in Sprague-Dawley rats (n = 48) and then divided them into 4 groups (unfilled defect, autologous bone graft, Geistlich BioOss® and our original xenogeneic bone mineral, 12 rats per group). Rats were sacrificed upon 4 and 12 months (6 rats per time point) with the following excision of the implant and adjacent tissues. 3D structure, extent of mineralisation, and bone volume were measured by means of microcomputed tomography. Microanatomy of the explants and adjacent tissue was investigated by haematoxylin and eosin staining.Results. The highest and the lowest bone volume was expectedly detected when the defect was filled with the autologous bone graft or remained unfilled, respectively. Replacement of the defect by the original bone mineral entailed better regeneration as compared to Geistlich Bio-Oss. Bone mineral density, bone thickness and the extent of mineralisation did not differ significantly between the experimental groups and were close to the positive control values, indicating efficient bone repair.Conclusions. Original xenogeneic bone mineral promotes induction of bone regeneration as compared to Geistlich Bio-Oss®, a commercially available bone mineral widely used in the clinical practice.

Bone formation with functionalized 3D printed poly-ε-caprolactone scaffold with plasma-rich-fibrin implanted in critical-sized calvaria defect of rat

Background/purposeSpace-making is one of the essential factors for bone regeneration in severe bony defect. To test the hypothesis that an appropriately designed scaffold may be beneficial for the bone formation in defect, the new bone formed in the critical-size calvarial defect of rat was examined after implanted with a 3D-printed poly-ɛ-caprolactone (PCL) scaffold, retaining with and without plasma rich fibrin (PRF). Materials and methodsThirty-two rats were divided into four groups (control, PCL, PRF, and PCL-plus-PRF). A custom-made 3D-printed PCL scaffold, 900 μm in pore size, retaining with and without PRF, was implanted into a critical-sized calvarial defect, 6 mm in diameter. Animals were sacrificed at week-4 or 8 after implantation for assessing the new bone formation by dental radiography, micro-computed tomography (μ-CT), and histology. ResultsBy radiography and μ-CT, significantly greater mineralization areas/volumes were observed in defects with 3D-printed scaffold groups compared to that without the scaffold in both two-time points. However, no advantage was found by adding PRF. Histology showed that bone tissues grew into the central zone of the critical defect when 3D-printed PCL scaffold was present. In contrast, for the groups without the scaffolds, new bones were formed mostly along defect borders, and the central zones of the defects were collapsed and healed with thin connective tissue. ConclusionOur results suggest that the use of a 900 μm pore size 3D-printed PCL scaffold may have the potential in facilitating the new bone formation.

Effect of extracellular matrix and dental pulp stem cells on bone regeneration with 3D printed PLA/HA composite scaffolds.

The demand for bone grafting procedures in various fields of medicine is increasing. Existing substitutes in clinical practice do not meet all the criteria required for an ideal bone scaffold, so new materials are being sought. This study evaluated bone regeneration using a critical-size Wistar rat's calvarial defect model. 12 male and 12 female rats were evenly divided into 3 groups: 1. Negative and positive (Geistlich Bio-Oss®) controls; 2. polylactic acid (PLA) and PLA/hydroxyapatite (HA); 3. PLA/HA cellularised with dental pulp stem cells (DPSC) and PLA/HA extracellular matrix (ECM) scaffolds. PLA/HA filament was created using hot-melt extrusion equipment. All scaffolds were fabricated using a 3D printer. DPSC were isolated from the incisors of adult Wistar rats. The defects were evaluated by micro-computed tomography (µCT) and histology, 8 weeks after surgery. µCT revealed that the Bio-Oss group generated 1.49 mm3 and PLA/HA ECM 1.495 mm3 more bone volume than the negative control. Histology showed a statistically significant difference between negative control and both (Bio-Oss and PLA/HA ECM) groups in rats of both genders. Moreover, histology showed gender-specific differences in all experimental groups and a statistically significant difference between cellularised PLA/HA and PLA/HA ECM groups in female rats. Qualitative histology showed the pronounced inflammation reaction during biodegradation in the PLA group. In conclusion, the bone-forming ability was comparable between the Bio-Oss and PLA/HA ECM scaffolds. Further research is needed to analyse the effects of ECM and PLA/HA ratio on osteoregeneration.

Prevascularized hydrogels with mature vascular networks promote the regeneration of critical-size calvarial bone defects in vivo.

Adequate vascularization of scaffolds is a prerequisite for successful repair and regeneration of lost and damaged tissues. It has been suggested that the maturity of engineered vascular capillaries, which is largely determined by the presence of functional perivascular mural cells (or pericytes), plays a vital role in maintaining vessel integrity during tissue repair and regeneration. Here, we investigated the role of pericyte-supported-engineered capillaries in regenerating bone in a critical-size rat calvarial defect model. Prior to implantation, human umbilical vein endothelial cells and human bone marrow stromal cells (hBMSCs) were cocultured in a collagen hydrogel to induce endothelial cell morphogenesis into microcapillaries and hBMSC differentiation into pericytes. Upon implantation into the calvarial bone defects (8mm), the prevascularized hydrogels showed better bone formation than either untreated controls or defects treated with autologous bone grafts (positive control). Bone formation parameters such as bone volume, coverage area, and vascularity were significantly better in the prevascularized hydrogel group than in the autologous bone group. Our results demonstrate that tissue constructs engineered with pericyte-supported vascular capillaries may approximate the regenerative capacity of autologous bone, despite the absence of osteoinductive or vasculogenic growth factors.

Bone regeneration in critical-size calvarial defect using functional biocompatible osteoinductive herbal scaffolds and human umbilical cord Wharton’s Jelly-derived mesenchymal stem cells

Critical size calvarial bone defects and their repair is the major challenge in orthopaedic surgery. Bone tissue engineering using varied combinations of scaffolds, stem cells, and growth factors is an emerging choice of treatment for variety of bone defects. Present study aimed to compare and evaluate the potential of poly ε-caprolactone (PCL), PCL- graphene oxide (GO), PCL-GO-Cissus quadrangularis (CQ), and human umbilical cord-derived mesenchymal stem cells (hUCMSCs) seeded PCL (PCL- hUCMSCs), PCL-GO-CQ-hUCMSCs scaffolds to heal critical size calvarial bone defect in the rat models. Healthy adult Wister female rats (N = 30) with average body weight of 350 ± 30 g were divided to 6 groups with five animals in each group. Single critical size calvarial defects of 8 mm were created in the skull of each rat and same were treated with respective scaffolds. The outcome of the treatments was evaluated at 6 weeks and 12 weeks in terms of weight, haematological parameters and biochemical parameters for biocompatibility. New bone regeneration/healing was analysed using digital radiography and micro-computed tomography. Quality of bone formed was analysed by bone mineral density. Histological analysis of the bone tissues was performed using an optical microscope. All the scaffolds were biocompatible and there was no adverse effect of these scaffolds on animals. Higher bone regeneration was observed after 12 weeks of transplantation than 6 weeks. However, PCL-GO-CQ-hUCMSCs scaffolds exhibited highest bone regeneration 12 weeks post-transplantation. The unique combination of PCL-GO-CQ-hUCMSCs scaffold proved to be beneficial in bone regeneration by aiding nearly complete healing of defect site. Further studies with PCL-GO-CQ-hUCMSCs could pave the way for human clinical trials.

Influence of the Degree of Deacetylation of Chitosan and BMP-2 Concentration on Biocompatibility and Osteogenic Properties of BMP-2/PLA Granule-Loaded Chitosan/β-Glycerophosphate Hydrogels.

Compositions based on chitosan/β-glycerophosphate hydrogels with highly porous polylactide granules can be used to obtain moldable bone graft materials that have osteoinductive and osteoconductive properties. To eliminate the influence of such characteristics as chain length, degree of purification, and molecular weight on a designed material, the one-stock chitosan sample was reacetylated to degrees of deacetylation (DD%) of 19.5, 39, 49, 55, and 56. A study of the chitosan/β-glycerophosphate hydrogel with chitosan of a reduced DD% showed that a low degree of deacetylation increased the MSCs (multipotent stromal cells) viability rate in vitro and reduced the leukocyte infiltration in subcutaneous implantation to Wistar rats in vivo. The addition of 12 wt% polylactide granules resulted in optimal composite mechanical and moldable properties, and increased the modulus of elasticity of the hydrogel-based material by approximately 100 times. Excessive filling of the material with PLA (polylactide) granules (more than 20%) led to material destruction at a ~10% strain. Osteoinductive and osteoconductive properties of the chitosan hydrogel-based material with reacetylated chitosan (39 DD%) and highly porous polylactide granules impregnated with BMP-2 (bone morphogenetic protein-2) have been demonstrated in models of orthotopic and ectopic bone formation. When implanted into a critical-size calvarial defect in rats, the optimal concentration of BMP-2 was 10 μg/mL: bone tissue areas filled the entire material’s thickness. Implantation of the material with 50 μg/mL BMP-2 was accompanied with excessive growth of bone tissue and material displacement beyond the defect. Significant osteoinductive and osteoconductive properties of the material with 10 μg/mL of BMP-2 were also shown in subcutaneous implantation.

Can the Combination of Simvastatin and Melatonin Create a Synergistic Effect on Bone Regeneration?

The present study evaluated the potential bone regeneration capacity of combining melatonin and simvastatin, with a goal of producing more osteogenic bone substitutes. A total of 48 male Wistar rats were randomly divided into 4 groups. The following were administered into critical-sized calvarial defects of the rats: Group I-human allograft; Group II-human allograft+ 10mg melatonin; Group III-human allograft+ 0.1mg simvastatin; and Group IV-human allograft+ 10mg melatonin+ 0.1mg simvastatin. Histopathologic, histomorphometric, and microcomputed tomographic evaluations were performed postprocedurally at 4 and 8weeks. A P value < .05 was considered significant for all evaluations. Groups II and III had significantly superior regeneration compared to Group I at weeks 4 and 8. Group III had significantly superior regeneration compared to Group II, particularly in week 4. Group IV had significantly superior regeneration compared to all groups at week8. The local administration of melatonin and simvastatin resulted in increased new bone mass and quality of bone microstructure than was seen in the control group. Simvastatin shortened the defect regeneration time more effectively than melatonin did. The combined use of melatonin and simvastatin provided a synergic effect on bone regeneration, particularly in the late phase of healing.

Bi-directional regulation functions of lanthanum-substituted layered double hydroxide nanohybrid scaffolds via activating osteogenesis and inhibiting osteoclastogenesis for osteoporotic bone regeneration.

Rationale: Osteoporotic patients suffer symptoms of excessive osteoclastogenesis and impaired osteogenesis, resulting in a great challenge to treat osteoporosis-related bone defects. Based on the positive effect of rare earth elements on bone metabolism and bone regeneration, we try to prove the hypothesis that the La3+ dopants in lanthanum-substituted MgAl layered double hydroxide (La-LDH) nanohybrid scaffolds simultaneously activate osteogenesis and inhibit osteoclastogenesis.Methods: A freeze-drying technology was employed to construct La-LDH nanohybrid scaffolds. The in vitro osteogenic and anti-osteoclastogenic activities of La-LDH nanohybrid scaffolds were evaluated by using ovariectomized rat bone marrow stromal cells (rBMSCs-OVX) and bone marrow-derived macrophages (BMMs) as cell models. The in vivo bone regeneration ability of the scaffolds was investigated by using critical-size calvarial bone defect model of OVX rats.Results: La-LDH nanohybrid scaffolds exhibited three-dimensional macroporous structure, and La-LDH nanoplates arranged perpendicularly on chitosan organic matrix. The La3+ dopants in the scaffolds promote proliferation and osteogenic differentiation of rBMSCs-OVX by activating Wnt/β-catenin pathway, leading to high expression of ALP, Runx-2, COL-1 and OCN genes. Moreover, La-LDH scaffolds significantly suppressed RANKL-induced osteoclastogenesis by inhibiting NF-κB signaling pathway. As compared with the scaffolds without La3+ dopants, La-LDH scaffolds provided more favourable microenvironment to induce new bone in-growth along macroporous channels.Conclusion: La-LDH nanohybrid scaffolds possessed the bi-directional regulation functions on osteogenesis and osteoclastogenesis for osteoporotic bone regeneration. The modification of La3+ dopants in bone scaffolds provides a novel strategy for osteoporosis-related bone defect healing.