On-Off Phagocytosis and Switchable Macrophage Activation Stimulated with NIR for Infected Percutaneous Tissue Repair of Polypyrrole-Coated Sulfonated PEEK.

Intelligent control of the immune response is essential for obtaining percutaneous implants with good sterilization and tissue repair abilities. In this study, polypyrrole (Ppy) nanoparticles enveloping a 3D frame of sulfonated polyether ether ketone (SP) surface are constructed, which enhance the surface modulus and hardness of the sulfonated layer by forming a cooperative structure of simulated reinforced concrete and exhibit a superior photothermal effect. Ppy-coated SP could quickly accumulate heat on the surface by responding to 808 nm near-infrared (NIR) light, thereby killing bacteria, and destroying biofilms. Under NIR stimulation, the phagocytosis and M1 activation of macrophages cultured on Ppy-coated SP are enhanced by activating complement 3 and its receptor, CD11b. Phagocytosis and M1 activation are impaired along with abolishment of NIR stimulation in the Ppy-coated SP group, which is favorable for tissue repair. Ppy-coated SP promotes Collagen-I, vascular endothelial growth factor, connective tissue growth factor, and α-actin (Acta2) expression by inducing M2 polarization owing to its higher surface modulus. Overall, Ppy-coated SP with enhanced mechanical properties could be a good candidate for clinical percutaneous implants through on-off phagocytosis and switchable macrophage activation stimulated with NIR.

A multifunctional cascade bioreactor based on a layered double oxides composite hydrogel for synergetic tumor chemodynamic/starvation/photothermal therapy.

The field of nanomedicine-catalyzed tumor therapy has achieved a lot of progress; however, overcoming the limitations of the tumor microenvironment (TME) to achieve the desired therapeutic effect remains a major challenge. In this study, a nanocomposite hydrogel (GH@LDO) platform combining the nanozyme CoMnFe-layered double oxides (CoMnFe-LDO) and natural enzyme glucose oxidase (GOX) was engineered to remodel the TME to enhance tumor catalytic therapy. The CoMnFe-LDO is a nanozyme that can convert endogenous H2O2 into reactive oxygen species (ROS) and O2 to achieve chemodynamic therapy (CDT) and alleviate the hypoxic microenvironment. Meanwhile, GOX can catalyze the conversion of glucose and O2 to gluconic acid and H2O2, which not only represses the ATP production of tumor cells to achieve starvation therapy (ST), but also decreases the pH value of TME and supplies extra H2O2 to enhance the CDT effect. Furthermore, this well-designed CoMnFe-LDO possessed a high photothermal conversion efficiency of GH@LDO (66.63%), which could promote the generation of ROS to enhance the CDT effect and achieve photothermal therapy (PTT) under near-infrared light irradiation. The GH@LDO hydrogel performes cascade reaction which overcomes the limitation of the TME and achieves satisfactory CDT/ST/PTT synergetic effects in vitro and in vivo. This work provides a new strategy for remodeling the TME using nanomedicine to achieve precise tumor cascaded catalytic therapy. STATEMENT OF SIGNIFICANCE: At present, the focus of tumor therapy has begun to shift from monotherapy to combination therapy for improving the overall therapeutic effect. In this study, we synthesized a CoMnFe-LDO nanozyme composed of multiple transition metal oxides, which demonstrated improved peroxidase and oxidase activities as well as favorable photothermal conversion capability. The CoMnFe-LDO nanozyme was compounded with an injectable GH hydrogel crosslinked by GOX and horseradish peroxidase (HRP). This nanocomposite hydrogel overcame the limitations of weak acidity, H2O2, and O2 levels in the TME and achieved synergetic CDT, ST, and PTT effects based on the cascaded catalytic actions of CoMnFe-LDO and GOX to H2O2 and glucose.

A Novel Stimuli-Responsive Injectable Antibacterial Hydrogel to Achieve Synergetic Photothermal/Gene-Targeted Therapy towards Uveal Melanoma.

Uveal melanoma (UM) is the most prevalent primary intraocular malignant tumor with a high lethal rate. Patients who undergo conventional enucleation treatments consistently suffer permanent blindness, facial defects, and mental disorders, therefore, novel therapeutic modalities are urgently required. Herein, an injectable and stimuli-responsive drug delivery antibacterial hydrogel (CP@Au@DC_AC50) is constructed via a facile grinding method that is inspired by the preparation process of traditional Chinese medicine. The incorporation of gold nanorods can enhance the mechanical strength of the hydrogel and realize photothermal therapy (PTT) and thermosensitive gel-sol transformation to release the gene-targeted drug DC_AC50 on demand in response to low-density near-infrared (NIR) light. The orthotopic model of UM is built successfully and indicates the excellent efficiency of CP@Au@DC_AC50 in killing tumors without damage to normal tissue because of its synergistic mild temperature PTT and gene-targeted therapy. Moreover, the eyeball infection model reveals the remarkable antibacterial properties of the hydrogel which can prevent endophthalmitis in the eyeball. There is negligible difference between the CP@Au@DC_AC50+NIR group and normal group. This NIR light-triggered gene-targeted therapy/PTT/antibacterial treatment pattern provides a promising strategy for building multifunctional therapeutic platform against intraocular tumors and exhibits great potential for the clinical treatment of UM.

Supraciliary Incision as a New Double-Eyelid Approach for Asian Patients: Clinical Experience of 528 Cases.

BACKGROUND: The double-eyelid operation is the most requested cosmetic surgery in Asians. The incision is usually located at the pretarsal skin 6 mm to 8 mm above palpebral margin. The purpose of this paper is to report a novel approach of double-eyelid operation through a supraciliary incision (SCI). METHODS: Three transverse curved lines were drawn on the upper lid skin. The location of line 1 (SCI) was 1.5 mm above the eyelash, line 2 according to the amount of redundant skin excised and line 3 at 3 mm to 4 mm above line 2. After the incisions were made between line 1 and line 2, the subcutaneous dissection is carried out over 5 mm the line 3. Then, the redundant skin and a strip orbicularis oculi muscle were removed to open the orbital septum and to explore underside levator aponeurosis. Along the line 3, the internal buried fixation sutures between dermal tissue and the fusion line of the orbital septum and levator aponeurosis were placed. Finally, the wounds were closed between line 2 and line 1. RESULTS: There were 528 patients who underwent the double-eyelid operation through the supraciliary approach. In long-term follow-up, 288 patients were evaluated at 6 months to 78 months postoperatively. Of those, 266 patients were satisfactory for the result (92.36%) with natural shape and invisible surgical scar. In another 22 patients (7.63%), a revised blepharoplasty was performed in 22 eyelids. CONCLUSION: The double-eyelid surgery using the SCI has several advantages including less visibility of the incision, the protected subdermal vascular network, the intact continuity of the upper eyelid skin, the combination of the SCI and internal dermal buried suture method. The approach can be considered an efficient technique for Asian patients. LEVEL OF EVIDENCE IV: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Sodium butyrate-modified sulfonated polyetheretherketone modulates macrophage behavior and shows enhanced antibacterial and osteogenic functions during implant-associated infections.

Prevention of implant-associated infections and insufficient bone tissue integration is critical to exploit the immunomodulatory properties and antibacterial effects of implant materials, which have attracted considerable attention. Modulation of the functions of immune cells in different environments is crucial for managing infection and inferior bone integration via immunomodulation. In this work, sodium butyrate, a fermentation product of gut microbiota, was loaded onto 3D porous sulfonated polyetheretherketone (SP) to modulate the immune responses of cells in different environments. Evaluation of in vitro antibacterial effects showed that sodium butyrate-loaded SP exhibited superior antibacterial activity, especially in the samples containing high concentrations of sodium butyrate. Under bacterial stimulation, the phagocytic activity of macrophages increased with an increase in the sodium butyrate concentration via the production of reactive oxygen species (ROS), which favoured bactericidal activity in the implant-associated infection stage. For bacterial elimination, sodium butyrate-containing SP could polarize macrophages to the M2 phenotype and subsequently stimulate anti-inflammatory cytokine secretion, which is considered beneficial for bone regeneration in the tissue repair stage. In vitro osteogenesis was evaluated and the results demonstrated that treatment with sodium butyrate-containing SP increased the expression of osteogenic genes and proteins. An in vivo rat osteomyelitis model was used to evaluate the protective effect of the SP-loaded with sodium butyrate on bone destruction and osteolysis under infection conditions. To study osteogenesis in vivo, a rat femoral model without infection was used. The results indicated that the SP-B2 group exhibited superior anti-infection capacity and induced new bone formation around the implant in vivo. Treatment with sodium butyrate-containing porous SP modulated the macrophage response under different stimuli, thereby serving as a new approach for the design of smart implant materials with superior antibacterial and bone repair properties.

Nanostructural Surfaces with Different Elastic Moduli Regulate the Immune Response by Stretching Macrophages.

A proper immune response is key for the successful implantation of biomaterials, and designing and fabricating biomaterials to regulate immune responses is the future trend. In this work, three different nanostructures were constructed on the surface of titanium using a hydrothermal method, and through a series of in vitro and in vivo experiments, we found that the aspect ratio of nanostructures can affect the elastic modulus of a material surface and further regulate immune cell behaviors. This work demonstrates that nanostructures with a higher aspect ratio can endow a material surface with a lower elastic modulus, which was confirmed by experiments and theoretical analyses. The deflection of nanostructures under the cell adsorption force is a substantial factor in stretching macrophages to enhance cell adhesion and spreading, further inducing macrophage polarization toward the M1 phenotype and leading to intense immune responses. In contrast, a nanostructure with a lower aspect ratio on a material surface leads to a higher surface elastic modulus, making deflection of the material difficult and creating a surface that is not conducive to macrophage adhesion and spreading, thus reducing the immune response. Moreover, molecular biology experiments indicated that regulation of the immune response by the elastic modulus is primarily related to the NF-κB signaling pathway. These findings suggest that the immune response can be regulated by constructing nanostructural surfaces with the proper elastic modulus through their influence on cell adhesion and spreading, which provides new insights into the surface design of biomaterials.

Multifunctional sulfonated polyetheretherketone coating with beta-defensin-14 for yielding durable and broad-spectrum antibacterial activity and osseointegration.

To address periprosthetic joint infection (PJI), a formidable complication after joint arthroplasty, an implant with excellent osseointegration and effective antibacterial activity has being extensively pursued and developed. In this work, the mouse beta-defensin-14 (MBD-14) was immobilized on the polyetheretherketone (PEEK) surface with three-dimensional (3D) porous structure to improve its antibacterial activity and osseointegration. An in vitro antibacterial evaluation showed that the porous PEEK loaded with MBD-14 wages a durable and effective fight against both Staphylococcus aureus (gram-positive) and Pseudomonas aeruginosa (gram-negative). In addition to the superior antibacterial activity, we found that the enhanced proliferation and osteogenic differentiation of bone mesenchymal stem cells were verified through various in vitro analyses. To evaluate the in vivo bactericidal effect and osseointegration of the samples, the rat femoral models with infection and non-infection were established. The enhanced osseointegration of the MBD-14-loaded samples was found in both two in vivo models. And no bacteria survived on the surfaces of samples with a relatively high MBD-14 concentration. Above results indicate that the 3D porous PEEK coating loaded with MBD-14 simultaneously yields excellent osseointegration while exerting durable and broad-spectrum antibacterial activity. And it paves the way for PEEK to be applied clinically to address PJI. STATEMENT OF SIGNIFICANCE: (1). By using the physio-chemical technique including sulfonation and lyophilization etc., a three-dimensional porous network is developed on polyetheretherketone (PEEK) surface, in which mouse beta-defensin-14 (MBD-14, a broad-spectrum antimicrobial peptide) is then loaded. It endows PEEK with antibacterial activity and osseointegration. (2). Two in vivo animal models with infection and non-infection are used to prove the new bone formation around the samples. (3). Supplementary material also proves that MBD-14 promotes the osteogenic differentiation of BMSCs. However, its potential mechanism needs to be further studied in future. (4). The modified PEEK, including excellent osseointegration and a durable and broad-spectrum antibacterial activity, could be applied clinically to address PJI which is a hot potato for surgeons and patients undergoing total joint arthroplasty.

Controllable and durable release of BMP-2-loaded 3D porous sulfonated polyetheretherketone (PEEK) for osteogenic activity enhancement.

Polyetheretherketone (PEEK) is ideal for dental and orthopedic applications because its mechanical properties are similar to cortical bones. However, its inherent inert ability hinders its clinical applications. In this work, bone morphogenetic protein-2 (BMP-2) was immobilized onto the sulfonated PEEK (SPEEK) using lyophilization technology. The surface morphologies of the samples were analyzed by field-emission scanning electron microscopy (FE-SEM), and the chemical compositions were analyzed by energy-dispersive X-ray spectrometry (EDS). The release content of BMP-2 of the samples immersed in the PBS (pH = 7.4) was detected by a human BMP-2 ELISA kit. The results indicated that controllable and durable BMP-2 release was accomplished due to the three-dimensional (3D) network of sulfonated PEEK. The in vitro cellular experiments showed that the BMP-2-immobilized samples significantly enhanced the initial adhesion and spreading of rat bone mesenchymal stem cells (rBMSCs). Moreover, the collagen secretion, extracellular matrix mineralization and ALP activity were also improved. Thus, the BMP-2-immobilized samples greatly promoted the osteogenic differentiation of rBMSCs, which revealed that BMP-2 immobilization paves the way for the use of PEEK in clinical applications.

Smart release of doxorubicin loaded on polyetheretherketone (PEEK) surface with 3D porous structure.

It is important to fabricate an implant possessing environment sensitive drug delivery. In this work, the construction of 3D porous structure on polyetheretherketone (PEEK) surface and pH sensitive polymer, chitosan, was introduced. The smart release of doxorubicin can be realized on the 3D porous surface of PEEK loading chitosan. We give a feasible explanation for the effect of chitosan on smart drug release according to Henderson-Hasselbalch equation. Furthermore, the intracellular drug content of the cell cultured on the samples with highest chitosan is significantly higher at pH 4.0, whereas lower at pH 7.4 than other samples. The smart release of doxorubicin via modification with chitosan onto 3D porous PEEK surface paves the way for the application of PEEK in drug loading platform for recovering bone defect caused by malignant bone tumor.

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.

Variation analysis of the severe acute respiratory syndrome coronavirus putative non-structural protein 2 gene and construction of three-dimensional model.

BACKGROUND: The rapid transmission and high mortality rate made severe acute respiratory syndrome (SARS) a global threat for which no efficacious therapy is available now. Without sufficient knowledge about the SARS coronavirus (SARS-CoV), it is impossible to define the candidate for the anti-SARS targets. The putative non-structural protein 2 (nsp2) (3CL(pro), following the nomenclature by Gao et al, also known as nsp5 in Snidjer et al) of SARS-CoV plays an important role in viral transcription and replication, and is an attractive target for anti-SARS drug development, so we carried on this study to have an insight into putative polymerase nsp2 of SARS-CoV Guangdong (GD) strain. METHODS: The SARS-CoV strain was isolated from a SARS patient in Guangdong, China, and cultured in Vero E6 cells. The nsp2 gene was amplified by reverse transcription-polymerase chain reaction (RT-PCR) and cloned into eukaryotic expression vector pCI-neo (pCI-neo/nsp2). Then the recombinant eukaryotic expression vector pCI-neo/nsp2 was transfected into COS-7 cells using lipofectin reagent to express the nsp2 protein. The expressive protein of SARS-CoV nsp2 was analyzed by 7% sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). The nucleotide sequence and protein sequence of GD nsp2 were compared with that of other SARS-CoV strains by nucleotide-nucleotide basic local alignment search tool (BLASTN) and protein-protein basic local alignment search tool (BLASTP) to investigate its variance trend during the transmission. The secondary structure of GD strain and that of other strains were predicted by Garnier-Osguthorpe-Robson (GOR) Secondary Structure Prediction. Three-dimensional-PSSM Protein Fold Recognition (Threading) Server was employed to construct the three-dimensional model of the nsp2 protein. RESULTS: The putative polymerase nsp2 gene of GD strain was amplified by RT-PCR. The eukaryotic expression vector (pCI-neo/nsp2) was constructed and expressed the protein in COS-7 cells successfully. The result of sequencing and sequence comparison with other SARS-CoV strains showed that nsp2 gene was relatively conservative during the transmission and total five base sites mutated in about 100 strains investigated, three of which in the early and middle phases caused synonymous mutation, and another two base sites variation in the late phase resulted in the amino acid substitutions and secondary structure changes. The three-dimensional structure of the nsp2 protein was successfully constructed. CONCLUSIONS: The results suggest that polymerase nsp2 is relatively stable during the phase of epidemic. The amino acid and secondary structure change may be important for viral infection. The fact that majority of single nucleotide variations (SNVs) are predicted to cause synonymous, as well as the result of low mutation rate of nsp2 gene in the epidemic variations, indicates that the nsp2 is conservative and could be a target for anti-SARS drugs. The three-dimensional structure result indicates that the nsp2 protein of GD strain is high homologous with 3CL(pro) of SARS-CoV urbani strain, 3CL(pro) of transmissible gastroenteritis virus and 3CL(pro) of human coronavirus 229E strain, which further suggests that nsp2 protein of GD strain possesses the activity of 3CL(pro).