Inhibiting wear particles-induced osteolysis with naringin.

PURPOSE: The purpose of this study was to determine the effects of naringin on osteoclastogenesis and osteolysis both in vitro and in vivo. METHODS: In this research osteoclasts were generated from mouse bone marrow monocytes with the receptor activator of NF-КB ligand and the macrophage colony stimulating factor. Naringin, at a concentration of 1, 10, 50, and 100 µg/mL, was respectively added to the medium. Seven days later, the osteoclasts were determined through tartrate-resistant acid phosphatase (TRAP) staining. Mature osteoclasts were isolated from newborn rabbits and cultured for three days on bone slices. Naringin at a concentration of 1, 10, 50, and 100 µg/mL was respectively added to the medium. The resorption bone slices were quantified, and the area was calculated after toluidine blue and Mayer-hematoxylin staining. Polymethyl methacrylate (PMMA) particles were implanted on the calvariae of C57BL/J6 mice. Naringin, at a dose of 50 µg/kg and 100 µg/kg, was respectively given intraperitoneally for seven. Seven days later, the calvariae were removed and processed for pathological analysis. RESULTS: The result indicated that naringin treatment effectively inhibited in vitro osteoclastogenesis and inhibited mature osteoclasts. In vivo data indicated that naringin strongly inhibited PMMA-induced osteolysis. CONCLUSION: Naringin can effectively inhibit osteoclastogenesis and suppress wear particles-induced osteolysis and might be useful in the treatment or prevention of wear particles-induced osteolysis and aseptic loosening for its effect on osteoclast generation and function.

Chronic calcaneal osteomyelitis associated with soft-tissue defect could be successfully treated with platelet-rich plasma: a case report.

Chronic osteomyelitis associated with soft-tissue defect following surgical management is a severe complication for orthopaedic surgeons. Traditionally, the treatment protocol for the notorious complication involved thorough debridement, bone grafting, long-term antibiotic use and flap surgery. Alternatively, platelet-rich plasma (PRP), a high concentration of platelets collected via centrifugation, has been successfully used as an adjuvant treatment for bone and soft-tissue infection in medical practices. PRP has numerous significant advantages, including stypsis, inflammation remission and reducing the amount of infected fluid. It increases bone and soft-tissue healing and allows fewer opportunities for transplant rejection. Through many years of studies showing the advantages of PRP, it has become preferred organic product for the clinical treatment of infections, especially for chronic osteomyelitis associated with soft-tissue defect. To promote the clinical use of this simple and efficacious technique in trauma, we report the case of a patient with chronic calcaneal osteomyelitis associated with soft-tissue defect that healed uneventfully with PRP.

Combination of platelet-rich plasma with degradable bioactive borate glass for segmental bone defect repair.

Porous scaffold biomaterials may offer a clinical alternative to bone grafts; however, scaffolds alone are typically insufficient to heal large bone defects. Numerous studies have demonstrated that osteoinductive growth factor significantly improves bone repair. In this study, a strategy combining degradable bioactive borate glass (BG) scaffolds with platelet-rich plasma (PRP) was tested. The bone defect was filled with BG alone, BG combined with autologous PRP or left empty. Bone formation was analyzed at 4, 8 and 12 weeks using both histology and radiology. The PRP treated group yielded better bone formation than the pure BG scaffold as determined by both histology and microcomputer tomography after 12 weeks. In conclusion, PRP improved bone healing in a diaphyseal rabbit model on BG. The combination of PRP and BG may be an effective approach to repair critical defects.

Well-dispersed platelet lysate entrapped nanoparticles incorporate with injectable PDLLA-PEG-PDLLA triblock for preferable cartilage engineering application.

Platelet lysate (PL) as a cost-effective cocktail of growth factors is an emerging ingredient in regenerative medicine, especially in cartilage tissue engineering. However, most studies fail to pay attention to PL's intrinsic characteristics and incorporate it directly with scaffolds or hydrogels by simple mixture. Currently, the particle size distribution of PL was determined to be scattered. Directly introducing PL into a thermosensitive poly(d,l-lactide)-poly(ethylene glycol)-poly(d,l-lactide) (PLEL) hydrogel disturbed its sol-gel transition. Electrostatic self-assembly heparin (Hep) and ε-poly-l-lysine (EPL) nanoparticles (NPs) were fabricated to improve the dispersity of PL. Such PL-NPs-incorporated PLEL gels retained the initial gelling capacity and showed a long-term PL-releasing ability. Moreover, the PL-loaded composite hydrogels inhibited the inflammatory response and dedifferentiation of IL-1ß-induced chondrocytes. For in vivo applications, the PLEL@PL-NPs system ameliorated the early cartilage degeneration and promoted cartilage repair in the late stage of osteoarthritis. RNA sequencing analysis indicated that PL's protective effects might be associated with modulating hyaluronan synthase 1 (HAS-1) expression. Taken together, these results suggest that well-dispersed PL by Hep/EPL NPs is a preferable approach for its incorporation into hydrogels and the constructed PLEL@PL-NPs system is a promising cell-free and stepwise treatment option for cartilage tissue engineering.

Incorporation of cerium oxide in hollow mesoporous bioglass scaffolds for enhanced bone regeneration by activating the ERK signaling pathway.

Hierarchically porous structures and bioactive compositions of artificial biomaterials play a positive role in bone defect healing and new bone regeneration. Herein, cerium oxide nanoparticles-modified bioglass (Ce-BG) scaffolds were firstly constructed by the incorporation of hollow mesoporous Ce-BG microspheres in CTS via a freeze-drying technology. The interconnected macropores in Ce-BG scaffolds facilitated the in-growth of bone cells/tissues from material surfaces into the interiors, while the hollow cores and mesopore shells in Ce-BG microspheres provides more active sites for bone mineralization. The cerium oxide nanoparticles in the scaffolds rapidly promoted the proliferation and osteogenic differentiation of human bone marrow-derived mesenchymal stem cells (hBMSCs), as confirmed by the up-regulation of osteogenesis-related markers such as OCN, ALP and COL-1. The enhanced osteoinductivity of Ce-BG scaffolds was mainly related to the activated ERK pathway, and it was blocked by adding a selective ERK1/2 inhibitor (SCH772984). In vivo rat cranial defect models revealed that Ce-BG scaffolds accelerated collagen deposition, osteoblast formation and bone regeneration as compared to BG scaffolds. The exciting results demonstrated that the synergistic effects between hierarchically porous structures and cerium oxide nanoparticles contributed to osteogenic ability, and hollow mesoporous Ce-BG scaffolds would be a novel platform for bone regeneration.

Study of biocompatibility of small intestinal submucosa (SIS) with Schwann cells in vitro.

No satisfactory method currently exists for repairing long peripheral nerve defects. Efforts have been made to fabricate bioactive artificial nerve conduits, comprised of a biomaterial pre-seeded with Schwann cells (SCs), which creating a favorable micro-environment for axonal regeneration, to be an alternative to autografting by means of tissue engineering. Small intestinal submucosa (SIS) possesses special biological characteristics and is comprehensively researched for tissue repairing at varied tissues and organs. This study investigated the biocompatibility of SIS with SCs in vitro. Cultured rat SCs were seeded on SIS. Cell morphology was observed by light microscopy, scanning electron microscopy and transmission electron microscope. The viability of SCs was measured by MTT assay. Secretion of NGF-beta and BDNF was quantitatively assessed by ELISA, and NGF-beta mRNA and BDNF mRNA were semi-quantitatively assessed by RT-PCR. The results indicated that SCs could adhere, migrate and proliferate on the surface of SIS in good condition with productive function of secreting growth factors. SIS has a good biocompatibility with SCs and SIS pre-seeded with SCs has potential to be an alternate candidate of autografting for repairing long peripheral nerve defects.

Comparative study of porous hydroxyapatite/chitosan and whitlockite/chitosan scaffolds for bone regeneration in calvarial defects.

Hydroxyapatite (HAP; Ca10(PO4)6(OH)2) and whitlockite (WH; Ca18Mg2(HPO4)2(PO4)12) are widely utilized in bone repair because they are the main components of hard tissues such as bones and teeth. In this paper, we synthesized HAP and WH hollow microspheres by using creatine phosphate disodium salt as an organic phosphorus source in aqueous solution through microwave-assisted hydrothermal method. Then, we prepared HAP/chitosan and WH/chitosan composite membranes to evaluate their biocompatibility in vitro and prepared porous HAP/chitosan and WH/chitosan scaffolds by freeze drying to compare their effects on bone regeneration in calvarial defects in a rat model. The experimental results indicated that the WH/chitosan composite membrane had a better biocompatibility, enhancing proliferation and osteogenic differentiation ability of human mesenchymal stem cells than HAP/chitosan. Moreover, the porous WH/chitosan scaffold can significantly promote bone regeneration in calvarial defects, and thus it is more promising for applications in tissue engineering such as calvarial repair compared to porous HAP/chitosan scaffold.

Sonochemical synthesis of fructose 1,6-bisphosphate dicalcium porous microspheres and their application in promotion of osteogenic differentiation.

Human bone mesenchymal stem cells (hBMSCs) have the ability to differentiate into bone and cartilage for clinical bone regeneration. Biomaterials with an innate ability to stimulate osteogenic differentiation of hBMSCs into bone and cartilage are considered attractive candidates for the applications in bone tissue engineering and regeneration. In this paper, we synthesized fructose 1,6-bisphosphate dicalcium (Ca2FBP) porous microspheres by the sonochemical method, and investigated the ability of Ca2FBP for the promotion of the osteogenic differentiation of hBMSCs. After the hBMSCs were co-cultured with the sterilized powder of Ca2FBP porous microspheres for different times, the cell proliferation assay, alkaline phosphatase activity assay, quantitative real-time polymerase chain reaction and western blotting were performed to investigate the bioactivity and osteogenic differentiation performance of the as-prepared product. Compared with hydroxyapatite nanorods, Ca2FBP porous microspheres show a superior bioactivity and osteoinductive potential, and can promote the cell differentiation of hBMSCs in vitro, thus, they are promising for applications in the tissue engineering field such as dental and bone defect repair.

[Periosteum construction in vitro by small intestinal submucosa combined with bone marrow mesenchymal stem cell].

OBJECTIVE: To investigate the practicability of periosteum construction in vitro by small intestinal submucosa (SIS) as a tissue scaffold with bone marrow mesenchymal stem cells (BMSC). METHODS: The bone marrow mesenchymal stem cells were cultivated by the conventional method in vitro, and then co-cultivated with SIS. The morphological feature of the complex material, the extracellular matrices, the adhesion and proliferation of BMSC were observed by optical microscope, electronic microscope, scanning electronic microscope and histologic evaluation respectively. RESULTS: BMSCs adhered and proliferated on SIS, secreted a great deal of extracellular matrices with active cellular function. BMSCs grew well there in layers, and the thickness became thicker as time passed, acting like a biological periosteum. CONCLUSION: Combined culture of SIS and BMSC in vitro can construct a tissue engineering periosteum similar to the natural one, thus providing a base for further study of its osteogenic regeneration in vivo.

Hydroxyapatite coatings with oriented nanoplate arrays: synthesis, formation mechanism and cytocompatibility.

Hydroxyapatite (HA) is the main inorganic constituent of natural bones and teeth with c-axis orientation and a(b)-axis orientation, respectively. Designing HA coatings (HACs) with specific orientation and morphology is an important strategy to improve their biological properties. Herein, we report, for the first time, the hydrothermal synthesis of HACs with oriented nanoplate arrays according to the following steps: (i) deposition of brushite/chitosan coatings (BCCs) on Ti6Al4V substrates; and (ii) transformation of HACs with oriented nanoplate arrays from BCCs after hydrothermal treatment with alkaline solutions. After soaking the BCCs in a NaOH solution under hydrothermal conditions, the Ca2+ and PO4 3- ions are released from the coatings because of the dissolution reaction of brushite, and they react with OH- ions to form HA nanoplates. Interestingly, these HA nanoplates with a preferential c-plane orientation are perpendicular to the coating surfaces. Hydrothermal reaction time and Ca/P ratio of BCCs have great effects on the morphologies of HA nanoplates. On increasing the reaction time from 3 h to 3 days or decreasing the Ca/P ratio from 2.0 to 1.0, the widths (or lengths) of HA nanoplates increase gradually. Simulated body fluid immersion (SBF) tests reveal that the HACs with oriented nanoplate arrays can promote the formation of apatite on the surfaces, suggesting their good in vitro bioactivity. Moreover, human bone marrow stromal cells (hBMSCs) have been used as cell models to investigate cytocompatibility of the HACs. The hBMSCs on the HACs have better cell adhesion, spreading, proliferation and osteogenic differentiation than those on Ti6Al4V substrates because the HACs are similar to the minerals of human hard tissues in chemical composition, morphology and crystallographic orientation. Therefore, HACs with oriented nanoplate arrays have great potential for use as implants of human hard tissues.

A novel injectable borate bioactive glass cement as an antibiotic delivery vehicle for treating osteomyelitis.

BACKGROUND: A novel injectable cement composed of chitosan-bonded borate bioactive glass (BG) particles was evaluated as a carrier for local delivery of vancomycin in the treatment of osteomyelitis in a rabbit tibial model. MATERIALS AND METHODS: The setting time, injectability, and compressive strength of the borate BG cement, and the release profile of vancomycin from the cement were measured in vitro. The capacity of the vancomycin-loaded BG cement to eradicate methicillin-resistant Staphylococcus aureus (MRSA)-induced osteomyelitis in rabbit tibiae in vivo was evaluated and compared with that for a vancomycin-loaded calcium sulfate (CS) cement and for intravenous injection of vancomycin. RESULTS: The BG cement had an injectability of >90% during the first 3 minutes after mixing, hardened within 30 minutes and, after hardening, had a compressive strength of 18 ± 2 MPa. Vancomycin was released from the BG cement into phosphate-buffered saline for up to 36 days, and the cumulative amount of vancomycin released was 86% of the amount initially loaded into the cement. In comparison, vancomycin was released from the CS cement for up 28 days and the cumulative amount released was 89%. Two months post-surgery, radiography and microbiological tests showed that the BG and CS cements had a better ability to eradicate osteomyelitis when compared to intravenous injection of vancomycin, but there was no significant difference between the BG and CS cements in eradicating the infection. Histological examination showed that the BG cement was biocompatible and had a good capacity for regenerating bone in the tibial defects. CONCLUSIONS: These results indicate that borate BG cement is a promising material both as an injectable carrier for vancomycin in the eradication of osteomyelitis and as an osteoconductive matrix to regenerate bone after the infection is cured.

Identification of one novel mutation in the EVC2 gene in a Chinese family with Ellis-van Creveld syndrome.

Ellis-van Creveld syndrome (EvC) is a rare autosomal recessive skeletal dysplasia characterized by short limbs, short ribs, postaxial polydactyly, and dysplastic nails and teeth. It is caused by biallelic mutations in the EVC or EVC2 gene. Here, we identified a novel nonsense mutation p.W828X (c.2484G>A) in exon 14 and a recurrent nonsense mutation p. R399X (c.1195C>T) in exon 10 of EVC2 gene in a Chinese boy with EvC. Identification of a novel genotype in EvC will provide clues to the phenotype-genotype relations and may assist not only in the clinical diagnosis of EvC but also in the interpretation of genetic information used for prenatal diagnosis and genetic counseling.

Novel borate glass/chitosan composite as a delivery vehicle for teicoplanin in the treatment of chronic osteomyelitis.

Composite materials composed of borate bioactive glass and chitosan (designated BGC) were investigated in vitro and in vivo as a new delivery system for teicoplanin in the treatment of chronic osteomyelitis induced by methicillin-resistant Staphylococcus aureus (MRSA). In vitro, the release of teicoplanin from BGC pellets into phosphate-buffered saline (PBS), as well as its antibacterial activity, were determined. The compressive strength of the pellets was measured after specific immersion times, and the structure of the pellets was characterized using scanning electron microscopy and X-ray diffraction. In vivo, the tibial cavity of New Zealand White rabbits was injected with MRSA strain to induce chronic osteomyelitis, treated by debridement after 4weeks, implanted with teicoplanin-loaded BGC pellets (designated TBGC) or BGC pellets, or injected intravenously with teicoplanin. After 12weeks' implantation, the efficacy of the TBGC pellets for treating osteomyelitis was evaluated using hematological, radiological, microbiological and histological techniques. When immersed in PBS, the TBGC pellets provided a sustained release of teicoplanin, while the surface of the pellets was converted to hydroxyapatite (HA). In vivo, the best therapeutic effect was observed in animals implanted with TBGC pellets, resulting in significantly lower radiological and histological scores, a lower positive rate of MRSA culture, and an excellent bone defect repair, without local or systemic side effects. The results indicate that TBGC pellets are effective in treating chronic osteomyelitis by providing a sustained release of teicoplanin, in addition to participating in bone regeneration.

[Experimental study on effect of platelet-rich plasma in repair of bone defect].

OBJECTIVE: To study the effect of platelet-rich plasma in the repair of bone defect. METHODS: Segmental bone defects of 1 cm were created in the mid-upper part of bilateral radius of 24 New Zealand white rabbits. One side was randomly chosen as the experimental side, which was filled with artificial bone with platelet-rich plasma (PRP). The other side filled with artificial bone without PRP as the control. After 2, 4, 8 and 12 weeks of implantation, the gross, radiological, histological observations, and computer graphic analysis were performed to investigate the bone healing of the defect in both sides. RESULTS: Two weeks after operation, new bone and fibrous tissue formation in both the experimental and the control sides were observed only in the areas adjacent to the cut ends of the host bone, but the amount of new tissue in the experimental side was much more than that in the control side. In the 4th and 8th weeks, the surface of the artificial bone was covered with a large amount of new bones, the artificial bone was bridged tightly with the host bone by callus in the experimental side, while new bone was limited mainly in the cut ends and was less mature in the control side. In the 12th weeks, bone defects were entirely healed in the experimental side, which were covered completely with cortical bone, while new bone formation was only observed in the ends of artificial bone and there were not continuous bone callus on the surface in the control side. CONCLUSION: Artificial bone with PRP is effective in the repair of segmental bone defects, and PRP could improve the healing of bone defect.