Comparative Clinical Evaluation of NeoPlex RB-8 with Seeplex PneumoBacter ACE for Simultaneous Detection of Eight Respiratory Bacterial Pathogens.

There are several convenient and accurate molecular assays to detect respiratory bacterial infection. The NeoPlex RB-8 detection kit (NeoPlex RB-8) is a new multiplex real-time PCR assay that simultaneously detects Streptococcus pneumoniae, Mycoplasma pneumoniae, Chlamydophila pneumoniae, Legionella pneumophila, Haemophilus influenzae, Bordetella pertussis, Bordetella parapertussis, and Moraxella catarrhalis in a single test. This study compared the clinical concordance of NeoPlex RB-8 with another method, Seeplex PneumoBacter ACE detection assay (Seeplex PB ACE), which simultaneously detects S. pneumoniae, M. pneumoniae, C. pneumoniae, L. pneumophila, H. influenzae, and B. pertussis We tested 2,137 nasopharyngeal swab and sputum specimens using both assays. For discordant Bordetella parapertussis and M. catarrhalis specimens, we also performed bidirectional sequencing. For S. pneumoniae, M. pneumoniae, C. pneumoniae, L. pneumophila, H. influenzae, and B. pertussis, which are detected by both NeoPlex RB-8 and Seeplex PB ACE, the positive and negative agreement between the two assays ranged from 91.7 to 100% (κ = 0.918 to 1). S. pneumoniae and H. influenzae were the most discordant targets and measured with higher sensitivity and specificity by NeoPlex RB-8 than Seeplex PB ACE. For Bordetella parapertussis and M. catarrhalis, which are not detected by Seeplex PB ACE, NeoPlex RB-8 sensitivity and specificity were >99%. Overall, NeoPlex RB-8 was highly comparable to Seeplex PB ACE, but NeoPlex RB-8 was more clinically accurate, with higher throughput and more convenience.

USF2 enhances the osteogenic differentiation of PDLCs by promoting ATF4 transcriptional activities.

OBJECTIVE: Our study aimed to elucidate the regulatory molecules related to the osteogenic differentiation of periodontal ligament cells (PDLCs). BACKGROUND: Periodontal ligament cells are a favorable source for cell-based therapy in periodontal bone engineering and regeneration due to their potential multilineage differentiation ability. However, the molecular mechanism and signaling pathways related to the osteogenic differentiation of PDLCs are still unclear. METHODS: Osteoblast-specific protein expression levels were examined by ELISA in osteogenic-induced PDLCs (induced-PDLC group). A microarray assay and a bioinformatics analysis were carried out to reveal significantly expressed genes and the related pathways in induced-PDLCs, and these findings were then confirmed by qRT-PCR and a luciferase reporter assay. Finally, overexpressing and silencing gene systems were established to identify the specific transcriptional relationship and function of the target genes on the osteogenic differentiation of PDLCs. RESULTS: Osteogenically differentiated PDLCs with high levels of osteoblast-specific proteins were established. The upstream stimulatory factor 2 (USF2) and activating transcription factor 4 (ATF4) mRNA levels were upregulated the most through the MAPK signaling pathway in the induced-PDLC group. USF2 could bind to the transcriptional initiation region of ATF4 and regulate its transcriptional activities. Additionally, the overexpression of USF2 promoted osteoblast-specific gene expression and the Alizarin red staining of PDLCs, while simultaneously overexpressing USF2 and silencing ATF4 reversed the favorable osteogenic effect of the induced-PDLCs by reducing osteoblast-specific gene expression and the Alizarin red staining level. CONCLUSION: Our study demonstrated that USF2 could enhance the osteogenic differentiation of PDLCs by regulating ATF4 transcriptional activities, which provides a new strategy to utilize USF2 and ATF4 as potential target molecules for periodontal bone regeneration.

Assessment of respiratory and systemic toxicity of Benzalkonium chloride following a 14-day inhalation study in rats.

BACKGROUND: Although biocides at low concentrations have been used to control pests, they can be more harmful than industrial chemicals as humans are directly and frequently exposed to such biocides. Benzalkonium chloride (BAC or BKC) is a non-toxic substance used to control pests. Recently, BAC has been increasingly used as a component in humidifier disinfectants in Korea, raising a serious health concern. Moreover, it poses significant health hazards to workers handling the chemical because of direct exposure. In the present study, we aimed to evaluate the respiratory toxicity of BAC due to its inhalation at exposure concentrations of 0.8 (T1 group), 4 (T2 group) and 20 (T3 group) mg/m3. RESULTS: In our previous study on the acute inhalational toxicity of BAC, bleeding from the nasal cavity was observed in all the rats after exposure to 50 mg/m3 BAC. Therefore, in this study, 20 mg/m3 was set as the highest exposure concentration, followed by 4 and 0.8 mg/m3 as the medium and low concentrations for 6 h/day and 14 days, respectively. After exposure, recovery periods of 2 and 4 weeks were provided. Additionally, alveolar lavage fluid was analyzed in males of the BAC-exposed groups at the end of exposure and 2 weeks after exposure to evaluate oxidative damage. In the T3 group exposed to BAC, deep breathing, hoarseness, and nasal discharge were observed along with a decline in feed intake and body weight, and nasal discharge was also observed in the T1 and T2 groups. ROS/RNS, IL-1ß, IL-6, and MIP-2 levels decreased in a concentration-dependent manner in the bronchoalveolar lavage fluid. Histopathological examination showed cellular changes in the nasal cavity and the lungs of the TI, T2, and T3 groups. CONCLUSIONS: As a result, it was confirmed that the target organs in the respiratory system were the nasal cavity and the lungs. The adverse effects were evaluated as reversible responses to oxidative damage. Furthermore, the no observed adverse effect level was found to be less than 0.8 mg/m3 and the lowest benchmark dose was 0.0031 mg/m3. Accordingly, the derived no-effect level of BAC was calculated as 0.000062 mg/m3.

Poly(acrylic acid)-regulated Synthesis of Rod-Like Calcium Carbonate Nanoparticles for Inducing the Osteogenic Differentiation of MC3T3-E1 Cells.

Calcium carbonate, especially with nanostructure, has been considered as a good candidate material for bone regeneration due to its excellent biodegradability and osteoconductivity. In this study, rod-like calcium carbonate nanoparticles (Rod-CC NPs) with desired water dispersibility were achieved with the regulation of poly (acrylic acid). Characterization results revealed that the Rod-CC NPs had an average length of 240 nm, a width of 90 nm with an average aspect ratio of 2.60 and a negative ζ-potential of -22.25 ± 0.35 mV. The degradation study illustrated the nanoparticles degraded 23% at pH 7.4 and 45% at pH 5.6 in phosphate-buffered saline (PBS) solution within three months. When cultured with MC3T3-E1 cells, the Rod-CC NPs exhibited a positive effect on the proliferation of osteoblast cells. Alkaline phosphatase (ALP) activity assays together with the osteocalcin (OCN) and bone sialoprotein (BSP) expression observations demonstrated the nanoparticles could induce the differentiation of MC3T3-E1 cells. Our study developed well-dispersed rod-like calcium carbonate nanoparticles which have great potential to be used in bone regeneration.

An antibacterial membrane based on Janus bacterial cellulose with nano-sized copper oxide through polydopamine conjugation for infectious wound healing.

Bacterial cellulose (BC) produced by Acetobacter xylinum has great advantages in wound dressing. However, the structural limitation under static culture, and lack of antibacterial properties restrict its application, especially for infectious wound healing. The present study reported an original wound dressing, which was composed of a Janus BC membrane with antibacterial nano-sized copper oxide (CuO) through polydopamine (PDA) conjugation to promote wound healing under infectious condition. The finished product (CuO/PDA/BC membrane) exhibited favorable air permeability, high hydrophilicity and good mechanical properties, as well as strong antibacterial effects by the sustained release of CuO and photothermal effect of CuO/PDA. Furthermore, CuO/PDA/BC membrane inhibited inflammatory response and promoted wound healing in an infectious wound model in vivo. These results suggested that our CuO/PDA/BC membrane had great potential as wound dressing for infectious wound healing.

Influence of nanocoated calcium phosphate on two different types of implant surfaces in different bone environment: an animal study.

OBJECTIVE: The purpose of this study was to evaluate the osseointegration of two different types of surfaces, smooth and roughened surface implants nanocoated with calcium phosphate (CAP) around different bone environment. MATERIALS AND METHODS: Five male mongrel dogs were used in this study. The premolars and molars were extracted on both sides of the mandible. Eight weeks after extraction, implants were submerged on both sides of the mandible. On the left, CAP nanocoated roughened surface (RCAP) implants were installed whereas, the CAP nanocoated smooth surface (SCAP) implants were installed on the right side. The control group had no defect, on the other hand, three-wall intrabony defects were surgically created adjacent to the implant in the experimental group. The dogs were sacrificed after 12 weeks. RESULTS: Histological and histomorphometrical analysis were performed with the specimen. The SCAP and RCAP implants showed good osseointegration with no statistical significance in the control group. Histologically, the SCAP group showed little resolution of the defect compared with the RCAP group. In the experimental groups, there was a significant difference in defect fill between SCAP and RCAP. CONCLUSION: Within the limits of our study, it can be concluded that SCAP and RCAP implants show no difference in sufficient bone area whereas, CAP nanocoating on roughened implant surface may enhance osseointegration in deficient bone environment.

Functional engineering strategies of 3D printed implants for hard tissue replacement.

Three-dimensional printing technology with the rapid development of printing materials are widely recognized as a promising way to fabricate bioartificial bone tissues. In consideration of the disadvantages of bone substitutes, including poor mechanical properties, lack of vascularization and insufficient osteointegration, functional modification strategies can provide multiple functions and desired characteristics of printing materials, enhance their physicochemical and biological properties in bone tissue engineering. Thus, this review focuses on the advances of functional engineering strategies for 3D printed biomaterials in hard tissue replacement. It is structured as introducing 3D printing technologies, properties of printing materials (metals, ceramics and polymers) and typical functional engineering strategies utilized in the application of bone, cartilage and joint regeneration.

Osseointegration of dental implants installed without mechanical engagement: a histometric analysis in dogs.

OBJECTIVES: The purpose of this study was to elucidate the healing pattern of sand-blasted, large grid, acid-etched (SLA)-surfaced implants at two healing periods in a model that represents loosened implants (LIs) installed without mechanical engagement. MATERIAL AND METHODS: Five mongrel dogs were used, in which 20 dental implants were prepared. The implants were divided into two groups according to the absence or presence of initial mechanical engagement: LIs) and control, respectively. An oversized drill was used to prepare the implant area for the LI group. The implants were allowed to heal for 4 or 8 weeks. After the healing period, the experimental animals were sacrificed and block sections were obtained for histological analysis and histometric measurements. RESULTS: All implants were in intimate contact with the host bone and were without any inflammation after both 4 and 8 weeks of healing. While the mean amount of bone-to-implant contact (BIC) was constant in the control group, it tended to increase in the LI group with increasing healing period. However, neither BIC nor bone density differed significantly between the groups or with the healing period. CONCLUSION: From the results of the study, it can be conjectured that the submerged and unloaded SLA-surfaced implants could result in successful osseointegration, even if the mechanical engagement was not obtained at placement of the implants.

Apatite nanosheets inhibit initial smooth muscle cell proliferation by damaging cell membrane.

Understanding how nanostructured coatings interact with cells is related to how they manipulate cell behaviors and is therefore critical for designing better biomaterials. The apatite nanosheets were deposited on metallic substrates via biomimetic precipitation. Cell viability of apatite nanosheets towards to smooth muscle cells (SMCs) were investigated, and the underlying mechanism was proposed. Apatite nanosheets presented inhibitory activity on SMC growth, and caused rupture of cell membranes. On the basis of measuring changes in intracellular calcium ([Ca2+]i), observing cell contraction and apatite nanosheets - SMC interaction, it was found that calcium ions released from apatite led to rises in [Ca2+]i, which induced vigorous SMC contraction on apatite nanosheets. Consequently, the cell membrane of individual SMCs was cut/penetrated by the sharp edges of apatite nanosheets, resulting in cell inactivation. This damage of cell membranes suggests a novel mechanism to manipulate cell viability, and may offer insights for the better design of calcium-based nanostructured coatings or other biomedical applications.

Resolution of surgically created three-wall intrabony defects in implants using three different biomaterials: an in vivo study.

OBJECTIVES: To histomorphometrically analyze bone formation on amorphous calcium phosphate (ACP), micro-macroporous biphasic calcium phosphate (MBCP), and freeze-dried bone allograft (FDBA) in three-wall defects adjacent to structured surface with calcium phosphate nanocoating implants in dogs. MATERIALS AND METHODS: Five male mixed-breed dogs were used in this study. The premolars and molars were extracted on both sides of the mandible. Eight weeks after extraction, four implants were submerged on each side of the mandible. Three-wall intrabony defects (5 × 3 × 3 mm) were surgically created adjacent to the implants before installation. No grafts were placed in the control group. At the experimental sites, each intrabony defect was grafted with either ACP, MBCP, or FDBA. The dogs were sacrificed after 12 weeks, and histological and histomorphometrical analyses of the implant sites were performed. RESULTS: All of the three experimental groups exhibited defect resolution and osseointegration that showed a statistically significant difference compared with the control group in terms of remaining defect depth and bone-to-implant contact (BIC). However, there were no statistical significances among the three experimental groups. MBCP had the highest BIC (63.57 ± 21.57%), followed by ACP and FDBA. The control group showed the least bone area and the greatest remaining defect depth. CONCLUSION: Grafts with the synthetic biomaterials ACP and MBCP showed bone regeneration that was similar to FDBA in surgically created three-wall intrabony defects adjacent to implants. Within the limitations of this study, it can be concluded that ACP and MBCP synthetic biomaterials are as effective as FDBA at osteoconduction.

Silk Fibroin/Collagen Blended Membrane Fabricated via a Green Papermaking Method for Potential Guided Bone Regeneration Application: In Vitro and In Vivo Evaluation.

Guided bone regeneration (GBR) technology is a commonly used surgical procedure for the repair of damaged periodontal tissues. Poor mechanical property and rapid degradation rate are the major reasons for GBR membrane failure in clinical applications. Herein, we applied a green papermaking method to fabricate silk fibroin (SF) membranes blended with collagen and tested their performance. The results showed that the blended SF75 (SF and collagen in a weight ratio of 75:25) membranes are biocompatible with good mechanical properties in the wet condition and appropriate biodegradation rate. MC3T3-E1 osteoblast cell adhesion and proliferation on the membranes were improved by the hybrid biological functions of SF and collagen. Subcutaneous implantation in rats for 9 weeks demonstrated that the membranes induced a less severe inflammatory response. The biodegradation time of the SF75 membranes was appropriate for tissue regeneration. This research, for the first time, reports a blended membrane prepared from silk fibroin and collagen with an ecofriendly method, which shows promise for application in guided bone regeneration.

Synergistic Effects on Incorporation of ß-Tricalcium Phosphate and Graphene Oxide Nanoparticles to Silk Fibroin/Soy Protein Isolate Scaffolds for Bone Tissue Engineering.

In bone tissue engineering, an ideal scaffold is required to have favorable physical, chemical (or physicochemical), and biological (or biochemical) properties to promote osteogenesis. Although silk fibroin (SF) and/or soy protein isolate (SPI) scaffolds have been widely used as an alternative to autologous and heterologous bone grafts, the poor mechanical property and insufficient osteoinductive capability has become an obstacle for their in vivo applications. Herein, ß-tricalcium phosphate (ß-TCP) and graphene oxide (GO) nanoparticles are incorporated into SF/SPI scaffolds simultaneously or individually. Physical and chemical properties of these composite scaffolds are evaluated using field emission scanning electron microscope (FESEM), X-ray diffraction (XRD) and attenuated total reflectance Fourier transformed infrared spectroscopy (ATR-FTIR). Biocompatibility and osteogenesis of the composite scaffolds are evaluated using bone marrow mesenchymal stem cells (BMSCs). All the composite scaffolds have a complex porous structure with proper pore sizes and porosities. Physicochemical properties of the scaffolds can be significantly increased through the incorporation of ß-TCP and GO nanoparticles. Alkaline phosphatase activity (ALP) and osteogenesis-related gene expression of the BMSCs are significantly enhanced in the presence of ß-TCP and GO nanoparticles. Especially, ß-TCP and GO nanoparticles have a synergistic effect on promoting osteogenesis. These results suggest that the ß-TCP and GO enhanced SF/SPI scaffolds are promising candidates for bone tissue regeneration.

Electrical stimulation as a novel tool for regulating cell behavior in tissue engineering.

Recently, electrical stimulation as a physical stimulus draws lots of attention. It shows great potential in disease treatment, wound healing, and mechanism study because of significant experimental performance. Electrical stimulation can activate many intracellular signaling pathways, and influence intracellular microenvironment, as a result, affect cell migration, cell proliferation, and cell differentiation. Electrical stimulation is using in tissue engineering as a novel type of tool in regeneration medicine. Besides, with the advantages of biocompatible conductive materials coming into view, the combination of electrical stimulation with suitable tissue engineered scaffolds can well combine the benefits of both and is ideal for the field of regenerative medicine. In this review, we summarize the various materials and latest technologies to deliver electrical stimulation. The influences of electrical stimulation on cell alignment, migration and its underlying mechanisms are discussed. Then the effect of electrical stimulation on cell proliferation and differentiation are also discussed.

Patterned iridium oxide film as neural electrode interface: Biocompatibility and improved neurite outgrowth with electrical stimulation.

Iridium (Ir) thin film was deposited on patterned titanium substrate by direct-current (DC) magnetron sputtering, and then activated in sulfuric acid (H2SO4) through repetitive potential sweeps to form iridium oxide (IrOx) as neural electrode interface. The resultant IrOx film showed a porous and open morphology with aligned microstructure, exhibited superior electrochemical performance and excellent stability. The IrOx film supported neural stem cells (NSCs) attachment, proliferation and improved processes without causing toxicity. The patterned IrOx films offered a unique system to investigate the synergistic effects of topographical cue and electrical stimulation on neurite outgrowth. Electrical stimulation, when applied through patterned IrOx films, was found to further increase the neurite extension of neuron-like cells and significantly reorient the neurite alignment towards to the direction of stimulation. These results indicate that IrOx film, as electrode-tissue interface is highly stable and biocompatible with excellent electrochemical properties.

Development of novel oxygen carriers by coupling hemoglobin to functionalized multiwall carbon nanotubes.

Preparation of stable and effective artificial oxygen carriers (AOCs) is a promising strategy to temporarily replace transfused blood and solve tissue hypoxia. Developing hemoglobin (Hb) loaded particles is one of the main ways to prepare suitable AOCs. Particles with a hierarchical micro/nanostructure can be loaded with plenty of proteins and have attracted great attention. Therefore, multiwall carbon nanotubes (MWCNTs) were chosen to fabricate AOCs. To improve the Hb-loading capacity of MWCNTs, functionalized MWCNTs, including carboxyl-functionalized MWCNTs (MWCNT-COOH), amino-functionalized MWCNTs (MWCNT-NH2), and heparin-conjugated MWCNTs (MWCNT-Hep), were prepared. Then, in this study, Hb was coupled to the functionalized MWCNTs to fabricate the AOCs. The functionalized MWCNTs and the AOCs were characterized by FTIR, SEM, TEM, and zeta potential analysis. The oxygen/Hb-loading capacity of the AOCs was also measured. The adverse effects of the AOCs on human umbilical vein endothelial cells (HUVECs) and human red blood cells (RBCs) were evaluated. The results showed that (1) the functional groups were grafted on the surface of the MWCNTs, and Hb was bound to the functionalized MWCNTs, thus the AOCs were successfully prepared; (2) MWCNT-Hep-Hb had the most stable dispersibility (i.e., the most negative zeta potential) in 0.9 wt% NaCl solution (MWCNT-Hep-Hb < MWCNT-COOH-Hb < MWCNT-Hb < MWCNT-NH2-Hb < 0); (3) MWCNT-Hep had the best Hb-loading capability, which was three times that of purified MWCNTs; (4) with concentrations increased up to 400 µg mL-1, MWCNT-Hep-Hb still had the highest cell viability (97.63% > 80%, ISO 10993-5:2009) and excellent blood biocompatibility. Therefore, MWCNT-Hep-Hb might be a satisfactory candidate as a blood substitute.

Osteogenesis of 3D printed macro-pore size biphasic calcium phosphate scaffold in rabbit calvaria.

To investigate the osteogenesis of macro-pore sized bone scaffolds, biphasic calcium phosphate scaffolds with accurately controlled macro-pore size (0.8, 1.2, and 1.6 mm) and identical porosity of 70% were fabricated by the 3D printing technology. Eight New Zealand rabbits were selected in the present study, while four 8-mm-diameter calvarial defects were created in each rabbit to place BCP scaffolds with different macro-pore size. The harvested specimens of four and eight weeks were used to evaluate the bone forming ability by micro CT and histological examination. All 3D-printed BCP scaffolds exhibited excellent mechanical properties and had better bone-forming ability than the control at both four and eight weeks. Among them, scaffold with 0.8 mm pore size was superior for initial bone formation and maturation, resulting in the highest value of total bone formation.

The biocompatibility of nanostructured calcium phosphate coated on micro-arc oxidized titanium.

To achieve improved osseointegration, there have been many efforts to modify the surface composition and topography of dental implants. Recently, the anodic oxidation treatment of titanium (Ti) has attracted a great deal of attention. Meanwhile, calcium phosphate is commonly applied to metallic implants as a coating material for fast fixation and firm implant-bone attachment on the account of its demonstrated bioactive and osteoconductive properties. In the present study, anodized surface and calcium phosphate deposition by electron beam evaporation were combined. Nanostructured calcium phosphate film was deposited on the micro-arc oxidized Ti. New apatite layer formed easily on the coated film when incubating in DPBS solution at 37 degrees C. By adding basic fibroblast growth factor (bFGF) in the DPBS solution, the bFGF could be immobilized in the newly formed apatite layer. The coated film enhanced osseointegration of Ti implants in vivo.

Laminin functionalized biomimetic apatite to regulate the adhesion and proliferation behaviors of neural stem cells.

BACKGROUND: Functionalizing biomaterial substrates with biological signals shows promise in regulating neural stem cell (NSC) behaviors through mimicking cellular microenvironment. However, diverse methods for immobilizing biological molecules yields promising results but with many problems. Biomimetic apatite is an excellent carrier due to its non-toxicity, good biocompatibility, biodegradability, and favorable affinity to plenty of molecules. Therefore, it may provide a promising alternative in regulating NSC behaviors. METHODS: Biomimetic apatite immobilized with the extracellular protein - laminin (LN) was prepared through coprecipitation process in modified Dulbecco's phosphate-buffered saline (DPBS) containing LN. The amount of coprecipitated LN and their release kinetics were examined. The adhesion and proliferation behaviors of NSC on biomimetic apatite immobilized with LN were investigated. RESULTS: The coprecipitation approach provided well retention of LN within biomimetic apatite up to 28 days, and supported the adhesion and proliferation of NSCs without cytotoxicity. For long-term cultivation, NSCs formed neurosphere-like aggregates on non-functionalized biomimetic apatite. A monolayer of proliferated NSCs on biomimetic apatite with coprecipitated LN was observed and even more stable than the positive control of LN coated tissue-culture treated polystyrene (TCP). CONCLUSION: The simple and reproducible method of coprecipitation suggests that biomimetic apatite is an ideal carrier to functionalize materials with biological molecules for neural-related applications.

Biomimetic apatite formed on cobalt-chromium alloy: A polymer-free carrier for drug eluting stent.

In this study, sirolimus (SRL) was loaded within biomimetic apatite formed on cobalt-chromium (Co-Cr) alloy, which has been reported for the first time, to inhibit the in-stent restenosis. Two different groups of loading SRL within biomimetic apatite were prepared: Group A (mono-layer of apatite/SRL) and Group B (bi-layer of apatite/SRL). Group A and Group B showed the biphasic pattern of SRL release up to 40 and 90days, respectively. The attachment of human artery smooth muscle cell (HASMC) for both Group A and Group B was significantly inhibited, and proliferation dramatically decreased with the release of SRL. Noteworthily, biomimetic apatite alone also suppressed the SMC proliferation. The porous biomimetic apatite uniformly covered Co-Cr stent without crack or webbings. After balloon expansion, the integrity of biomimetic apatite was sufficient to resist delamination or destruction. Thus, this study demonstrated that biomimetic apatite is a promising drug carrier for potential use in stents.

Silk fibroin membrane used for guided bone tissue regeneration.

With the aim to develop a novel membrane with an appropriate mechanical property and degradation rate for guided bone tissue regeneration, lyophilized and densified silk fibroin membrane was fabricated and its mechanical behavior as well as biodegradation property were investigated. The osteoconductive potency of the silk fibroin membranes were evaluated in a defect rabbit calvarial model. Silk fibroin membrane showed the modulated biodegradable and mechanical properties via ethanol treatment with different concentration. The membrane could prevent soft tissue invasion from normal tissue healing, and the amounts of new bone and defect closure with silk fibroin membrane were similar to those of commercially available collagen membrane.