The Exosome-Mediated Bone Regeneration: An Advanced Horizon Toward the Isolation, Engineering, Carrying Modalities, and Mechanisms.

Exosomes, nanoparticles secreted by various cells, composed of a bilayer lipid membrane, and containing bioactive substances such as proteins, nucleic acids, metabolites, etc., have been intensively investigated in tissue engineering owing to their high biocompatibility and versatile biofunction. However, there is still a lack of a high-quality review on bone defect regeneration potentiated by exosomes. In this review, the biogenesis and isolation methods of exosomes are first introduced. More importantly, the engineered exosomes of the current state of knowledge are discussed intensively in this review. Afterward, the biomaterial carriers of exosomes and the mechanisms of bone repair elucidated by compelling evidence are presented. Thus, future perspectives and concerns are revealed to help devise advanced modalities based on exosomes to overcome the challenges of bone regeneration. It is totally believed this review will attract special attention from clinicians and provide promising ideas for their future works.

Extracellular vesicles secreted by human gingival mesenchymal stem cells promote bone regeneration in rat femoral bone defects.

Extracellular vesicles (EVs), important components of paracrine secretion, are involved in various pathological and physiological processes of the body. In this study, we researched the benefits of EVs secreted by human gingival mesenchymal stem cells (hGMSC-derived EVs) in promoting bone regeneration, thereby providing new ideas for EVs-based bone regeneration therapy. Here, we successfully demonstrated that hGMSC-derived EVs could enhance the osteogenic ability of rat bone marrow mesenchymal stem cells and the angiogenic capability of human umbilical vein endothelial cells. Then, femoral defect rat models were created and treated with phosphate-buffered saline, nanohydroxyapatite/collagen (nHAC), a grouping of nHAC/hGMSCs, and a grouping of nHAC/EVs. The results of our study indicated that the combination of hGMSC-derived EVs and nHAC materials could significantly promote new bone formation and neovascularization with a similar effect to that of the nHAC/hGMSCs group. Our outcomes provide new messages on the role of hGMSC-derived EVs in tissue engineering, which exhibit great potential in bone regeneration treatment.

Case Report: Application of Mixed Reality Combined With A Surgical Template for Precise Periapical Surgery.

Objective: The etiology of apical diseases is diverse, and most are due to incomplete root canal therapy. The common clinical manifestations include gingival abscess, fistula and bone destruction. The currently existing limitation of procedures is that surgeons cannot visually evaluate the surgical areas. We sought to combine mixed reality (MR) technology with a 3-dimensional (3D) printed surgical template to achieve visualization in apical surgery. Notably, no reports have described this application. Methods: We created visual 3D (V3D) files and transferred them into the HoloLens system. We explained the surgical therapy plan to the patient using a mixed reality head-mounted display (MR-HMD). Then, the 3D information was preliminarily matched with the operative area, and the optimal surgical approach was determined by combining this information with 3D surgical guide plate technology. Results: We successfully developed a suitable surgical workflow and confirmed the optimal surgical approach from the buccal side. We completely exposed the apical lesion and removed the inflammatory granulation tissue. Conclusion: We are the first group to use the MR technique in apical surgery. We integrated the MR technique with a 3D surgical template to successfully accomplish the surgery. Desirable outcomes using minimally invasive therapy could be achieved with the MR technique.

D-RADA16-RGD-Reinforced Nano-Hydroxyapatite/Polyamide 66 Ternary Biomaterial for Bone Formation.

BACKGROUND: Nano-hydroxyapatite/polyamide 66 (nHA/PA66) is a composite used widely in the repair of bone defects. However, this material is insufficient bioactivity. In contrast, D-RADA16-RGD self-assembling peptide (D-RADA16-RGD sequence containing all D-amino acids is Ac-RADARADARADARADARGDS-CONH2) shows admirable bioactivity for both cell culture and bone regeneration. Here, we describe the fabrication of a favorable biomaterial material (nHA/PA66/D-RADA16-RGD). METHODS: Proteinase K and circular dichroism spectroscopy were employed to test the stability and secondary structural properties of peptide D-RADA16-RGD respectively. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used to characterize the surface of these materials. Confocal laser scanning (CLS), cell counting kit-8 tests (CCK-8), alizarin red S staining, cell immunofluorescence analysis and Western blotting were involved in vitro. Also biosafety and bioactivity of them have been evaluated in vivo. RESULTS: Proteinase K and circular dichroism spectroscopy demonstrated that D-RADA16-RGD in nHA/PA66 was able to form stable-sheet secondary structure. SEM and TEM showed that the D-RADA16-RGD material was 7-33 nm in width and 130-600 nm in length, and the interwoven pore size ranged from 40 to 200 nm. CLS suggests that cells in nHA/PA66/D-RADA16-RGD group were linked to adjacent cells with more actin filaments. CCK-8 analysis showed that nHA/PA66/D-RADA16-RGD revealed good biocompatibility. The results of Alizarin-red S staining and Western blotting as well as vivo osteogenesis suggest nHA/PA66/D-RADA16-RGD exhibits better bioactivity. CONCLUSION: This study demonstrates that our nHA/PA66/D-RADA16-RGD composite exhibits reasonable mechanical properties, biocompatibility and bioactivity with promotion of bone formation.

Polydopamine-induced hydroxyapatite coating facilitates hydroxyapatite/polyamide 66 implant osteogenesis: an in vitro and in vivo evaluation.

BACKGROUND: Hydroxyapatite/polyamide 66 (HA/P66) has been clinically used for several years owing to its good biocompatibility and bioactivity. However, it has been found that the osseointegration process of the HA/P66 implant takes a large amount of time because of the small amount of HA on its surface. METHODS: To increase the amount of HA and aid faster osseointegration, we prepared a HA coating using a biomimetic process assisted by polydopamine (PDA) on the HA/P66 substrate. The surface properties of the substrate modified by PDA and HA were characterized, and the capacity of biomaterials for osteogenic induction was investigated both in vitro and in vivo. RESULTS: The HA coating was successfully prepared on the HA/P66 substrate and verified by X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), and X-ray diffraction (XRD). The HA coating remained firmly attached to the underlying PDA-HA/P66 substrate even after strong ultrasound treatment for 1 h, and the calcium and phosphorus of the HA coating was continuously released in vitro in a slow manner. The formation of the HA coating on the PDA film greatly increased the hydrophilicity and surface roughness of HA/P66. In cell-based experiments, as compared with the HA/P66 substrate, the HA coating formation on the PDA film could facilitate the functions of C3H10T1/2 cells, including cell adhesion, proliferation, spreading, alkaline phosphatase activity, calcium nodule formation, and expression of osteogenic differentiation-related proteins. In addition, the HA/P66 scaffolds modified with PDA and HA coatings were implanted in rabbit femoral condyles. At 8 weeks after surgery, micro-computed tomography scanning (micro-CT) and hematoxylin-eosin (HE) staining revealed that more new bones were formed around the HA/P66 scaffold that was modified with a PDA-assisted HA coating. CONCLUSION: These results indicate that the preparation of a PDA-assisted HA coating by using a biomimetic process significantly improves the capacity of biomaterials for osteogenic induction.

In vitro and in vivo biocompatibility and osteogenesis of graphene-reinforced nanohydroxyapatite polyamide66 ternary biocomposite as orthopedic implant material.

Graphene and its derivatives have been receiving increasing attention regarding their application in bone tissue engineering because of their excellent characteristics, such as a vast specific surface area and excellent mechanical properties. In this study, graphene-reinforced nanohydroxyapatite/polyamide66 (nHA/PA66) bone screws were prepared. The results of scanning electron microscopy observation and X-ray diffraction data showed that both graphene and nHA had good dispersion in the PA66 matrix. In addition, the tensile strength and elastic modulus of the composites were significantly improved by 49.14% and 21.2%, respectively. The murine bone marrow mesenchymal stem cell line C3H10T1/2 exhibited better adhesion and proliferation in graphene reinforced nHA/PA66 composite material compared to the nHA/PA66 composites. The cells developed more pseudopods, with greater cell density and a more distinguishable cytoskeletal structure. These results were confirmed by fluorescent staining and cell viability assays. After C3H10T1/2 cells were cultured in osteogenic differentiation medium for 7 and 14 days, the bone differentiation-related gene expression, alkaline phosphatase, and osteocalcin were significantly increased in the cells cocultured with graphene reinforced nHA/PA66. This result demonstrated the bone-inducing characteristics of this composite material, a finding that was further supported by alizarin red staining results. In addition, graphene reinforced nHA/PA66 bone screws were implanted in canine femoral condyles, and postoperative histology revealed no obvious damage to the liver, spleen, kidneys, brain, or other major organs. The bone tissue around the implant grew well and was directly connected to the implant. The soft tissues showed no obvious inflammatory reaction, which demonstrated the good biocompatibility of the screws. These observations indicate that graphene-reinforced nHA/PA66 composites have great potential for application in bone tissue engineering.

Development and in vitro characterization of drug delivery system of rifapentine for osteoarticular tuberculosis.

The study was to develop and evaluate the rifapentine-loaded poly(lactic acid-co-glycolic acid) (PLGA) microspheres (RPMs) for the treatment of osteoarticular tuberculosis to avoid critical side effects caused by oral regimens of antibiotics or intravenous antibiotics. The RPMs were spherical with rough surfaces, and elevated amounts of rifapentine in the formulation markedly increased the particle size and drug loading, while decreased the size distribution and entrapment efficiency. The highest drug loading and encapsulation efficiency of RPMs were 23.93%±3.93% and 88.49%±8.49%, respectively. After the initial rapid drug release, the release rate gradually decreased, and approximately 80% of the encapsulated rifapentine was released after 30 days of incubation. Moreover, RPMs could effectively inhibit the growth of Staphylococcus aureus. With increasing rifapentine content, the inhibition zones were continuously enlarged while the minimal inhibitory concentration values decreased. These results suggested that RPMs were bioactive and controlled release delivery systems for the treatment of osteoarticular tuberculosis.

Bone plate composed of a ternary nano-hydroxyapatite/polyamide 66/glass fiber composite: biomechanical properties and biocompatibility.

An ideal bone plate for internal fixation of bone fractures should have good biomechanical properties and biocompatibility. In this study, we prepared a new nondegradable bone plate composed of a ternary nano-hydroxyapatite/polyamide 66/glass fiber (n-HA/PA66/GF) composite. A breakage area on the n-HA/PA66/GF plate surface was characterized by scanning electron microscopy. Its mechanical properties were investigated using bone-plate constructs and biocompatibility was evaluated in vitro using bone marrow-derived mesenchymal stem cells. The results confirmed that adhesion between the n-HA/PA66 matrix and the glass fibers was strong, with only a few fibers pulled out at the site of breakage. Fractures fixed by the n-HA/PA66/GF plate showed lower stiffness and had satisfactory strength compared with rigid fixation using a titanium plate. Moreover, the results with regard to mesenchymal stem cell morphology, MTT assay, Alizarin Red S staining, enzyme-linked immunosorbent assay, and reverse transcription polymerase chain reaction for alkaline phosphatase and osteocalcin showed that the n-HA/PA66/GF composite was suitable for attachment and proliferation of mesenchymal stem cells, and did not have a negative influence on matrix mineralization or osteogenic differentiation of mesenchymal stem cells. These observations indicate that the n-HA/PA66/GF plate has good biomechanical properties and biocompatibility, and may be considered a new option for internal fixation in orthopedic surgery.

In vitro and in vivo evaluations of nano-hydroxyapatite/polyamide 66/glass fibre (n-HA/PA66/GF) as a novel bioactive bone screw.

In this study, we prepared nano-hydroxyapatite/polyamide 66/glass fibre (n-HA/PA66/GF) bioactive bone screws. The microstructure, morphology and coating of the screws were characterised, and the adhesion, proliferation and viability of MC3T3-E1 cells on n-HA/PA66/GF scaffolds were determined using scanning electron microscope, CCK-8 assays and cellular immunofluorescence analysis. The results confirmed that n-HA/PA66/GF scaffolds were biocompatible and had no negative effect on MC3T3-E1 cells in vitro. To investigate the in vivo biocompatibility, internal fixation properties and osteogenesis of the bioactive screws, both n-HA/PA66/GF screws and metallic screws were used to repair intercondylar femur fractures in dogs. General photography, CT examination, micro-CT examination, histological staining and biomechanical assays were performed at 4, 8, 12 and 24 weeks after operation. The n-HA/PA66/GF screws exhibited good biocompatibility, high mechanical strength and extensive osteogenesis in the host bone. Moreover, 24 weeks after implantation, the maximum push-out load of the bioactive screws was greater than that of the metallic screws. As shown by their good cytocompatibility, excellent biomechanical strength and fast formation and ingrowth of new bone, n-HA/PA66/GF screws are thus suitable for orthopaedic clinical applications.

Development and characterization of an injectable cement of nano calcium-deficient hydroxyapatite/multi(amino acid) copolymer/calcium sulfate hemihydrate for bone repair.

A novel injectable bone cement was developed by integration of nano calcium-deficient hydroxyapatite/multi(amino acid) copolymer (n-CDHA/MAC) and calcium sulfate hemihydrate (CSH; CaSO4 · 1/2H2O). The structure, setting time, and compressive strength of the cement were investigated. The results showed that the cement with a liquid to powder ratio of 0.8 mL/g exhibited good injectability and appropriate setting time and mechanical properties. In vitro cell studies indicated that MC3T3-E1 cells cultured on the n-CDHA/MAC/CSH composite spread well and showed a good proliferation state. The alkaline phosphatase activity of the MC3T3-E1 cells cultured on the n-CDHA/MAC/CSH composite was significantly higher than that of the cells on pure CSH at 4 and 7 days of culture. The n-CDHA/MAC/CSH cement was implanted into critical size defects of the femoral condyle in rabbits to evaluate its biocompatibility and osteogenesis in vivo. Radiological and histological results indicated that introduction of the n-CDHA/MAC into CSH enhanced new bone formation, and the n-CDHA/MAC/CSH cement exhibited good biocompatibility and degradability. In conclusion, the injectable n-CDHA/MAC/CSH composite cement has a significant clinical advantage over pure CSH cement, and may be a promising bone graft substitute for the treatment of bone defects.