Black phosphorus-enhanced injectable hydrogel for infected soft tissue healing.

The misuse of antibiotics makes clinical treatment of soft tissue infection a huge challenge in prosthesis replacement. In this study, a black phosphorus (BP)-enhanced antibacterial injectable hydrogel (HAABP) was developed by the dynamic coordinative cross-linking among thiolated hyaluronic acid, silver ion (Ag+), and BP. HAABP has been proven to possess typical porous structures, excellent injectability, and rapid self-healing properties. In addition, the shear modulus was positive correlative to the concentration of BP. In vitro, HAABP maintained good cytocompatibility and showed a highly efficient synergistic inhibitory effect on Staphylococcus aureus through the irradiation of near infrared light and the release of Ag+. In vivo, HAABP not only inhibited the persistent infection but also accelerated the deposition of collagen fibers and angiogenesis by down-regulating the inflammatory factor TNF-α in the infectious wound defect, thereby repairing the natural barrier of tissue. This study developed a BP-enhanced injectable hydrogel that provided a simple and efficient synergistic antibacterial strategy to treat soft tissue infections around prostheses.

Epigenetic Regulation in Knee Osteoarthritis.

Osteoarthritis (OA) is a complicated disease with both hereditary and environmental causes. Despite an increase in reports of possible OA risk loci, it has become clear that genetics is not the sole cause of osteoarthritis. Epigenetics, which can be triggered by environmental influences and result in transcriptional alterations, may have a role in OA pathogenesis. The majority of recent research on the epigenetics of OA has been focused on DNA methylation, histone modification, and non-coding RNAs. However, this study will explore epigenetic regulation in OA at the present stage. How genetics, environmental variables, and epigenetics interact will be researched, shedding light for future studies. Their possible interaction and control processes open up new avenues for the development of innovative osteoarthritis treatment and diagnostic techniques.

Preliminary Study on Immediate Postoperative CT Images and Values of the Modular Polyetheretherketone Based Total Knee Arthroplasty: An Observational First-in-Human Trial.

BACKGROUND: Total knee arthroplasty (TKA) is now frequently performed and is highly successful. However, patient satisfaction after TKA is often difficult to achieve. Because of the presence of metallic prosthetic knee joints, there is a lack of imaging tools that can accurately assess the patient's postoperative prosthetic position, soft tissue impingement, and periprosthetic bone density after TKA. We conducted a clinical trial of the world's first totally modular polyetheretherketone (PEEK) TKA and determined the bone density values in the stress concentration area around the prosthesis based on postoperative computed tomography data to reconstruct a three-dimensional model of the PEEK prosthetic knee joint after implantation. Based on the model, the overhang of the prosthesis was measured at various locations on the prosthesis. METHODS: All patients who underwent PEEK-based TKA were postoperatively assessed with radiography and computed tomography (CT). Hounsfield units (HUs) for the different components of the quantitative CT assessment were measured separately. RESULTS: Ten patients (nine female and one male) aged 59-74 (mean 66.9, median 67) years were included. The HU values were as follows: PEEK prosthesis mean 182.95, standard deviation (SD) 4.90, coefficient of variation (CV) 2.68; polyethylene mean -89.41, SD 4.14, CV -4.63; lateral femoral osteochondral mean 192.19, SD 55.05, CV 28.64; lateral tibial osteochondral mean 122.94, SD 62.14, CV 42.86; medial femoral osteophyte mean 180.76, SD 43.48, CV 24.05; and medial tibial osteophyte mean 282.59, SD 69.28, CV 24.52. Analysis of the data at 1, 3, and 6 months showed that the mean PE (p = 0.598) and PEEK (p = 0.916) measurements did not change with the time of measurement. There was a decrease in bone mineral density in the lateral tibia at 3 months (p = 0.044). Otherwise, there was no significant change in bone density in other regions (p = 0.124-0.803). There was no overhang in all femoral prostheses, whereas there were two cases of overhang in tibial prostheses. Overhang measurements do not differ significantly across time points. The overhang measurements were not significantly different at all time points (p = 0.186-0.967). CONCLUSION: PEEK knee joint prosthesis has excellent CT compatibility. The change in periprosthetic bone volume during the follow-up period can be determined using the HU value after CT scan, while the prosthesis position can be assessed. This assessment may potentially guide future improvements in knee prosthesis alignment techniques and artificial knee prosthesis designs.

Hydrofluoric acid and nitric acid cotreatment for biofunctionalization of polyetheretherketone in M2 macrophage polarization and osteogenesis.

Due to its excellent mechanical and low-friction properties, polyetheretherketone (PEEK) has been widely investigated for use in orthopedic applications over the past decade. However, the bioinertness and poor osteogenic properties of PEEK have hampered its clinical application. In this study, the surface of PEEK was modified by co-treatment with hydrofluoric acid and nitric acid (AFN). The microstructures of the modified PEEK surfaces were investigated using scanning electron microscopy. The water contact angles of the surfaces were also measured. To evaluate their cytocompatibility, PEEK samples were used as substrates to culture rat bone mesenchymal stem cells, and cell adhesion, viability, and expression of specific marker genes were measured. Treatment of PEEK with AFN (PEEK-AFN) was found to enable better osteoblast adhesion, spreading, and proliferation; the activity of alkaline phosphatase (an early osteogenic differentiation marker) was also found to be enhanced post-treatment. Furthermore, PEEK-AFN was able to modulate macrophage polarization and down regulated the expression of proinflammatory factors via inhibiting the NF-κB pathway. Thus, treatment of PEEK with AFN could promote M2 polarization of the macrophages and stimulate the differentiation of osteoblasts. These results provide valuable information that could facilitate the use of PEEK-based composites as bone implant materials.

Curcumin Attenuation of Wear Particle-Induced Osteolysis via RANKL Signaling Pathway Suppression in Mouse Calvarial Model.

Wear particle-induced chronic inflammation and osteoclastogenesis are two critical factors in the osteolytic process. Curcumin (CUR) is an active compound of the medicinal herb Curcuma longa and has anti-inflammatory and antiosteoclastogenic properties. Our study tested the hypothesis that CUR might attenuate polymethylmethacrylate- (PMMA-) induced inflammatory osteolysis using mouse calvaria osteolysis model in vivo and in vitro. The mice were divided into four groups: phosphate-buffered saline group, CUR, PMMA, and PMMA + CUR groups. Three days before PMMA particle implantation, the mice were intraperitoneally injected with CUR (25 mg/kg/day). Ten days after the operation, the mouse calvaria was harvested for microcomputed tomography, histomorphometry, and molecular biology analysis. As expected, CUR markedly reduced the secretion of tumor necrosis factor-α, interleukin- (IL-) 1ß, and IL-6 in the calvarial organ culture. Moreover, CUR suppressed osteoclastogenesis and decreased bone resorption in vivo compared with PMMA-stimulated calvaria. Furthermore, CUR downregulated the osteoclast-specific gene expression and reversed the receptor activator of nuclear factor kappa-B ligand (RANKL)/osteoprotegerin messenger RNA and protein ratio in PMMA particle-stimulated mice. These results suggest that CUR attenuated PMMA particle-induced inflammatory osteolysis by suppressing the RANKL signaling pathway in the murine calvarium, which could be a candidate compound to prevent and treat AL.

Immunomodulatory Effects of Calcium and Strontium Co-Doped Titanium Oxides on Osteogenesis.

The effects of calcium (Ca) or strontium (Sr) on host osteogenesis and immune responses have been investigated separately. In clinical practice, these two elements may both be present around an orthopedic device, but their potential synergistic effects on osteogenesis and the immune response have not been explored to date. In this work, we investigated the immunomodulatory effects of Ca and Sr co-doped titanium oxides on osteogenesis in vitro using the mouse macrophage cell line RAW 264.7 alone and in co-culture with mouse bone mesenchymal stem cells (BMSCs), and in vivo using a mouse air-pouch model. Coatings containing Ca and Sr at different concentration ratios were fabricated on titanium substrates using micro-arc oxidation and electrochemical treatment. The in vitro and in vivo results demonstrated that the Ca and Sr concentration ratio has a marked influence on macrophage polarization. The coating with a Ca/Sr ratio of 2:1 was superior to those with other Ca and/or Sr concentrations in terms of modulating M2 polarization, which enhanced osteogenic differentiation of mouse BMSCs in co-culture. These findings suggest that the osteoimmunomodulatory effect of a titanium-oxide coating can be enhanced by modulating the concentration ratio of its components.

Anthocyanin suppresses CoCrMo particle-induced osteolysis by inhibiting IKKα/ß mediated NF-κB signaling in a mouse calvarial model.

Wear particle-induced osteolysis and bone resorption have been identified as critical factors of implant failure and total joint revision, in which nuclear factor kappa B (NF-κB) signaling and chronic inflammation have been shown to play key roles. Although anthocyanin is known to have anti-inflammatory function via blocking NF-κB pathway, it is still unclear whether anthocyanin has a protective effect on particle-induced osteolysis. In the present study, we aimed to investigate the detailed effects and the underlying mechanism of anthocyanin on CoCrMo particle-induced osteolysis in a mouse calvavial model. One hundred and twelve male BALB/c mice were divided randomly into four groups: sham group (sham operation and injection with PBS), vehicle group (CoCrMo particle treatment and injection with PBS), low-dose anthocyanin group (CoCrMo particle treatment and injecting anthocyanin with 0.1mg/g/day), and high-dose anthocyanin group (CoCrMo particle treatment and injecting anthocyanin with 0.4mg/g/day). Mice were sacrificed after two weeks, harvesting the calvariae tissue for in depth analysis by micro-CT, histomorphometry, immunohistochemical and molecular biology analysis. As expected, anthocyanin markedly inhibited CoCrMo particle-induced inflammatory infiltration and decreased bone loss in vivo. Anthocyanin also reversed the increase in the ratio of receptor activator of nuclear factor kappa B ligand (RANKL)/osteoproteger (OPG) and suppressed osteoclast formation in CoCrMo particle-stimulated calvaria. Additionally, anthocyanin significantly reduced the expression and secretion of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß) and interleukin-6 (IL-6) in the calvaria of CoCrMo-stimulated mice. Furthermore, we confirmed that anthocyanin attenuated osteolysis by blocking NF-κB pathway via inhibiting inhibitor of nuclear factor kappa-B kinase α/ß (IKKα/ß) phosphorylation. In conclusion, our study demonstrated that anthocyanin can protect against CoCrMo particle-induced inflammatory osteolysis via inhibiting the IKKα/ß-NF-κB pathway, and have a potential therapeutic effect on the treatment of wear particle-induced osteolysis.

Curcumin Attenuates Titanium Particle-Induced Inflammation by Regulating Macrophage Polarization In Vitro and In Vivo.

Periprosthetic inflammatory osteolysis and subsequent aseptic loosening are commonly observed in total joint arthroplasty. Other than revision surgery, few approved treatments are available for this complication. Wear particle-induced inflammation and macrophage polarization state play critical roles in periprosthetic osteolysis. We investigated the effects of curcumin, a polyphenol extracted from Curcuma longa, on titanium (Ti) particle-induced inflammation and macrophage polarization in vitro using the murine cell line RAW 264.7 and in vivo using a murine air pouch model. The expression of specific macrophage markers was qualitatively analyzed by immunofluorescence (inducible nitric oxide synthase and CD206) and quantitatively analyzed by flow cytometry (CCR7 and CD206), representing M1 and M2 macrophages, respectively. Our results show that curcumin induced a higher percentage of M2 macrophages together with a higher concentration of anti-inflammatory cytokine IL-10, and a lower percentage of M1 macrophages with a lower concentration of pro-inflammatory cytokines (TNF-α and IL-6). The genes encoding CD86 (M1) and CD163 (M2), two additional markers, were shifted by curcumin toward an M2 phenotype. C57BL/J6 mice were injected with air and Ti particles to establish an air pouch model. Curcumin reduced cell infiltration in the pouch membrane and decreased membrane thickness. The analysis of exudates obtained from pouches demonstrated that the effects of curcumin on macrophage polarization and cytokine production were similar to those observed in vitro. These results prove that curcumin suppresses Ti particle-induced inflammation by regulating macrophage polarization. Thus, curcumin could be developed as a new therapeutic candidate for the prevention and treatment of inflammatory osteolysis and aseptic loosening.

In vitro and in vivo responses of macrophages to magnesium-doped titanium.

Modulating immune response to biomaterials through changing macrophage polarization has been proven to be a promising strategy to elicit beneficial outcomes in tissue repair. The objective of this study was to evaluate the response of macrophage polarization to titanium doped with magnesium (0.1~0.35%), which was prepared through the magnesium plasma immersion ion implantation (Mg PIII) technique. The M1/M2 polarization profile of macrophages was investigated using a murine cell line RAW 264.7 in vitro and a murine air pouch model in vivo. Our results demonstrated that the Mg PIII-treated titanium induced a higher percentage of M2 macrophages and higher concentrations of the anti-inflammatory cytokines interleukin (IL)-4 and IL-10. Genes encoding two growth factors, bone morphogenetic protein 2 (BMP2) and vascular endothelial growth factor (VEGF) were up-regulated, thus indicating the ability of the M2 phenotype to promote wound healing. The nuclear factor κB (NF-κB) signalling pathway was down-regulated. In vivo the Mg PIII -treated titanium elicited a similar effect on macrophage polarization and induced thinner fibrous capsule formation and a decrease in infiltrated cells. These results indicate that Mg PIII treatment has the immunomodulatory potential to elicit the pro-healing M2-polarized macrophage phenotype, thus providing new insight into the development of immunomodulatory biomaterials.

Silver-nanoparticles-modified biomaterial surface resistant to staphylococcus: new insight into the antimicrobial action of silver.

Titanium implants are widely used clinically, but postoperative implant infection remains a potential severe complication. The purpose of this study was to investigate the antibacterial activity of nano-silver(Ag)-functionalized Ti surfaces against epidemic Staphylococcus from the perspective of the regulation of biofilm-related genes and based on a bacteria-cell co-culture study. To achieve this goal, two representative epidemic Staphylococcus strains, Staphylococcus epidermidis (S. epidermidis, RP62A) and Staphylococcus aureus (S. aureus, USA 300), were used, and it was found that an Ag-nanoparticle-modified Ti surface could regulate the expression levels of biofilm-related genes (icaA and icaR for S. epidermidis; fnbA and fnbB for S. aureus) to inhibit bacterial adhesion and biofilm formation. Moreover, a novel bacteria-fibroblast co-culture study revealed that the incorporation of Ag nanoparticles on such a surface can help mammalian cells to survive, adhere and spread more successfully than Staphylococcus. Therefore, the modified surface was demonstrated to possess a good anti-infective capability against both sessile bacteria and planktonic bacteria through synergy between the effects of Ag nanoparticles and ion release. This work provides new insight into the antimicrobial action and mechanism of Ag-nanoparticle-functionalized Ti surfaces with bacteria-killing and cell-assisting capabilities and paves the way towards better satisfying the clinical needs.

Increased Effects of Extracorporeal Shock Waves Combined with Gentamicin against Staphylococcus aureus Biofilms In Vitro and In Vivo.

An implant-associated bacterial infection is one of the most common and costly complications of orthopedic surgery. Once biofilms develop, it is extremely difficult to cure infections with antimicrobial agents. High-energy extracorporeal shock wave (ESW) treatment has been used for orthopedic-related diseases and has been found to be an effective bactericidal agent that is tolerable both in vitro and in vivo. The broad-spectrum antibiotic gentamicin exhibits bactericidal activity against Staphylococcus aureus, and bacterial resistance to gentamicin is lower. We tested the effectiveness of gentamicin in combination with ESW treatment against S. aureus biofilms in vivo and in vitro. The spread plate method, crystal violet staining, confocal laser scanning microscopy, scanning electron microscopy and microbiologic evaluation were used to compare the effects of combined treatment with those of either treatment alone. The results revealed statistically significant differences between the group treated with ESWs combined with gentamicin and all other groups. Our findings indicate that use of the combination of ESWs with gentamicin is more effective against S. aureus biofilms in vitro and in vivo.

Nano-thick calcium oxide armed titanium: boosts bone cells against methicillin-resistant Staphylococcus aureus.

Since the use of systemic antibiotics for preventing acute biomaterial-associated infections (BAIs) may build up bacterial resistance and result in huge medical costs and unpredictable mortality, new precaution strategies are required. Here, it demonstrated that titanium armed with a nano-thick calcium oxide layer was effective on averting methicillin-resistant Staphylococcus aureus (MRSA) infections in rabbits. The calcium oxide layer was constructed by, firstly, injecting of metallic calcium into titanium via a plasma immersion ion implantation process, and then transforming the outer most surface into oxide by exposing to the atmosphere. Although the calcium oxide armed titanium had a relative low reduction rate (~74%) in growth of MRSA in vitro, it could markedly promote the osteogenic differentiation of bone marrow stem cells (BMSCs), restore local bone integration against the challenge of MRSA, and decrease the incidence of MRSA infection with a rate of 100% (compared to the titanium control). This study demonstrated for the first time that calcium, as one of the major elements in a human body, could be engineered to avert MRSA infections, which is promising as a safe precaution of disinfection for implantable biomedical devices.

Influence of sulfur content on bone formation and antibacterial ability of sulfonated PEEK.

Polyetheretherketone (PEEK) is desirable in orthopedic and dental applications because its mechanical properties are similar to those of natural bones but the bioinertness and inferior osteoconduction of PEEK have hampered many clinical applications. In this work, PEEK is sulfonated by concentrated sulfuric acid to fabricate a three-dimensional (3D) network. A hydrothermal treatment is subsequently conducted to remove the residues and the temperature is adjusted to obtain different sulfur concentrations. In vitro cell proliferation and real-time PCR analyses disclose enhanced proliferation and osteogenic differentiation of rat bone mesenchymal stem cells (rBMSCs) on the samples with small sulfur concentrations. The in vitro antibacterial evaluation reveals that all the sulfonated samples possess excellent resistance against Staphylococcus aureus and Escherichia coli. The in vivo rat femur implantation model is adopted and X-ray, micro-CT, and histological analyses indicate that not only the premeditated injected bacteria cells are sterilized, but also new bone forms around the samples with small sulfur concentrations. The in vitro and in vivo results reveal that the samples subjected to the hydrothermal treatment to remove excess sulfur have better osseointegration and antibacterial ability and PEEK modified by sulfonation and hydrothermal treatment is promising in orthopedic and dental applications.

Zn/Ag micro-galvanic couples formed on titanium and osseointegration effects in the presence of S. aureus.

Titanium implants possessing simultaneous osseointegration and antibacterial ability are desirable. In this work, three types of Zn/Ag micro-galvanic couples are fabricated on titanium by plasma immersion ion implantation to investigate the osseointegration and antibacterial effects as well as the involved mechanisms. The in vitro findings disclose enhanced proliferation, osteogenic differentiation, and gene expressions of the rat bone mesenchymal stem cells (rBMSCs), as well as good antibacterial ability on all three micro-galvanic couples. Excellent antimicrobial ability is also observed in vivo and the micro-CT and histological results reveal notable osseointegration in vivo despite the presence of bacteria. The Zn/Ag micro-galvanic couple formed on Zn/Ag dual-ion co-implanted titanium shows the best osseointegration as well as good antibacterial properties in vivo obtained from a rabbit tibia model. The difference among the three Zn/Ag micro-galvanic couples can be ascribed to the contact between the Ag NPs and Zn film, which affects the corrosion process. Our results indicate that the biological behavior can be controlled by the corrosion process of the Zn/Ag micro-galvanic couples.

The in vitro and in vivo performance of a strontium-containing coating on the low-modulus Ti35Nb2Ta3Zr alloy formed by micro-arc oxidation.

The ß-titanium alloy is thought to be a promising alloy using as orthopedic or dental implants owing to its characteristics, which contains low elastic modulus, high corrosion resistance and well biocompatibility. Our previous study has reported that a new ß-titanium alloy Ti35Nb2Ta3Zr showed low modulus close to human bone, equal tissue compatibility to a traditional implant alloy Ti6Al4V. In this study, micro-arc oxidation (MAO) was applied on the Ti35Nb2Ta3Zr alloy to enhance its surface characteristics and biocompatibility and osseointegration ability. Two different coatings were formed, TiO2 doped with calcium-phosphate coating (Ca-P) and calcium-phosphate-strontium coating (Ca-P-Sr). Then we evaluated the effects of the MAO coatings on the Ti35Nb2Ta3Zr alloy through in vitro and in vivo tests. As to the characteristics of the coatings, the morphology, chemical composition, surface roughness and contact angle of MAO coatings were tested by scanning electron microscopy, energy dispersive spectroscopy, atomic force microscopy, and video contact-angle measurement system respectively. Besides, we performed MTT assay, ALP test and cell morphology-adhesion test on materials to evaluate the MAOed coating materials' biocompatibility in vitro. The in vivo experiment was performed through rabbit model. Alloys were implanted into rabbits' femur shafts, then we performed micro-CT, histological and sequential fluorescent labeling analysis to evaluate implants' osseointegration ability in vivo. Finally, the Ca-P specimens and Ca-P-Sr specimens exhibited a significant enhancement in surface roughness, hydrophilicity, cell proliferation, cell adhesion. More new bone was found around the Ca-P-Sr coated alloy than Ca-P coated alloy and Ti35Nb2Ta3Zr alloy. In conclusion, the MAO treatment improved in vitro and in vivo performance of Ti35Nb2Ta3Zr alloy. The Ca-P-Sr coating may be a promising modified surface formed by MAO for the novel ß-titanium alloy Ti35Nb2Ta3Zr.

Balancing the Osteogenic and Antibacterial Properties of Titanium by Codoping of Mg and Ag: An in Vitro and in Vivo Study.

To simultaneously enhance the osteogenic and antibacterial properties of titanium, we introduced magnesium (Mg), silver (Ag), or both by using the plasma immersion ion implantation (PIII) technique, producing three PIII sample groups, namely, Mg-doped titanium (Mg-PIII), Ag-doped titanium (Ag-PIII), and Mg and Ag codoped titanium (Mg/Ag-PIII). The in vitro antibacterial efficacy of Mg/Ag-PIII group was about 7-10% higher than that of Ag-PIII. In vitro and in vivo results demonstrated that osteogenic property of Mg/Ag PIII group was better than that of Ag-PIII or Mg-PIII. It was believed that the galvanic effects between Mg and Ag NPs played a key role in facilitating the release of Mg but reducing the release of silver, answering for the selective performances of the Mg/Ag-PIII group over bacterial and mammalian cells. This study demonstrated that the integration of multiple functional elements could be realized by the dual-source PIII technique, and in this case, the antibacterial properties and osteogenic property of titanium could be balanced.

Antimicrobial and osteogenic properties of silver-ion-implanted stainless steel.

Prevention of implant loosening and infection is crucial to orthopedic and dental surgeries. In this work, the surface of stainless steel (SS) was modified by silver-sourced plasma immersion ion implantation (Ag-PIII). Metallic silver nanoparticles with various diameters and distributions were fabricated on the SS surfaces after treatment with Ag-PIII for 0.5 and 1.5 h, respectively. The osteogenic activity and antimicrobial properties of SS before and after Ag-PIII treatment were evaluated using in vitro and in vivo tests. The results demonstrated that Ag-PIII treatment not only promoted the antibacterial activity of SS but also enhanced the osteogenic differentiation of human bone marrow stromal cells.

Calcium Plasma Implanted Titanium Surface with Hierarchical Microstructure for Improving the Bone Formation.

Introducing hierarchical microstructure and bioactive trace elements simultaneously onto the surface of titanium implant is a very effective way to improve the osseointegration between bone and implant. In this work, hierarchical topography was prepared on Ti surface via acid etching and sandblasting (SLA) to form micropits and microcavities then underwent Ca plasma immersion ion implantation (Ca-PIII) process. The surface wettability and roughness did not change obviously before and after Ca-PIII process. The in vitro evaluations including cell adhesion, activity, alkaline phosphatase (ALP), osteogenic genes (Runx2, OSX, ALP, BSP, Col1a1, OPN, and OC), and protein (BSP, Col1a1, OPN, and OC) expressions revealed that the introduction of Ca ions onto the surface of SLA-treated Ti can promote greater osteoblasts adhesion, spread and proliferation, which in return further accelerated the maturation and mineralization of osteoblasts. More importantly, in vivo evaluations including Micro-CT evaluation, histological observations, push-out test, sequential fluorescent labeling and histological observations verified that Ca-SLA-treated Ti implants could efficiently promote new bone formation in early times. These promising results suggest that Ca-SLA-treated Ti has the potential for future application in orthopedic field.