Functionalization of Titanium with Chitosan via Silanation: Evaluation of Biological and Mechanical Performances

Complications in dentistry and orthopaedic surgery are mainly induced by peri-implant bacterial infections and current implant devices do not prevent such infections. The coating of antibacterial molecules such as chitosan on its surface would give the implant bioactive properties. The major challenge of this type of coating is the attachment of chitosan to a metal substrate. In this study, we propose to investigate the functionalization of titanium with chitosan via a silanation. Firstly, the surface chemistry and mechanical properties of such coating were evaluated. We also verified if the coated chitosan retained its biocompatibility with the peri-implant cells, as well as its antibacterial properties. FTIR and Tof-SIMS analyses confirmed the presence of chitosan on the titanium surface. This coating showed great scratch resistance and was strongly adhesive to the substrate. These mechanical properties were consistent with an implantology application. The Chitosan-coated surfaces showed strong inhibition of Actinomyces naeslundii growth; they nonetheless showed a non significant inhibition against Porphyromonas gingivalis after 32 hours in liquid media. The chitosan-coating also demonstrated good biocompatibility to NIH3T3 fibroblasts. Thus this method of covalent coating provides a biocompatible material with improved bioactive properties. These results proved that covalent coating of chitosan has significant potential in biomedical device implantation.     

Detailed evaluation of the agarose diffusion test in biocompatibility study with a microscopic image analysis system. Effect of plasma sterilization on biocompatible of an improved photoset polymer]

In addition to sterilizability, biocompatibility is a further necessary property of biomaterials. The agarose diffusion test is an established standard in vitro procedure for investigating this property. The usual method of evaluating the agarose diffusion test, based on macroscopic determination of the size of the zone of decoloration, and microscopic assessment of cell lysis, limits the power of the test. To obtain more information on the impact of plasma sterilization on the biocompatibility of a new polymer, a high-resolution computer-aided morphometric image analysis system for the quantification of the agarose diffusion test at the cellular level has been developed. This was able to detect statistically significant differences in the cell lysis rate of plasma sterilized polymer specimens before and after 4 weeks of incubation in a physiological solution. The new method provides highly detailed information on the interaction of soluble material elements and cells. In addition to cell damage by potentially toxic elements, the sensitivity of the L929 fibroblasts used in the agarose diffusion test to mechanical stress also needs to be considered and verified by further investigations.     

几丁聚糖在硅橡胶表面作涂层的实验研究

几丁聚糖具有良好的生物相容性和抗菌性,作为生物涂层已经引起了广泛的注意。研究了基体的不同表面处理方法、浸涂次数和几丁聚糖溶液浓度对涂层的表面形貌、结合强度等性能的影响,结果显示,几丁聚糖溶液浓度和硅橡胶表面粗糙度都存在一个最佳取值范围;增加浸涂次数可以改善涂层光洁度,但是对涂层附着力贡献不大;硅橡胶经紫外照射后可以改善几丁聚糖在其上的成膜性能。     

The Effect of Heparin-VEGF Multilayer on the Biocompatibility of Decellularized Aortic Valve with Platelet and Endothelial Progenitor Cells

The application of polyelectrolyte multilayer films is a new, versatile approach to surface modification of decellularized tissue, which has the potential to greatly enhance the functionality of engineered tissue constructs derived from decellularized organs. In the present study, we test the hypothesis that Heparin- vascular endothelial growth factor (VEGF) multilayer film can not only act as an antithrombotic coating reagent, but also induce proliferation of endothelial progenitor cells (EPCs) on the decellularized aortic heart valve. SEM demonstrated the adhesion and geometric deformation of platelets. The quantitative assay of platelet activation was determined by measuring the production of soluble P-selectin. Binding and subsequent release of heparin and VEGF from valve leaflets were assessed qualitatively by laser confocal scanning microscopy and quantitatively by ELISA methods. Human blood derived EPCs were cultured and the adhesion and growth of EPCs on the surface modified valvular scaffolds were assessed. The results showed that Heparin-VEGF multilayer film improved decellularized valve haemocompatibility with respect to a substantial reduction of platelet adhesion. Release of VEGF from the decellularized heart valve leaflets at physiological conditions was sustained over 5 days. In vitro biological tests demonstrated that EPCs achieved better adhesion, proliferation and migration on the coatings with Heparin-VEGF multilayer film. Combined, these results indicate that Heparin-VEGF multilayer film could be used to cover the decellularized porcine aortic valve to decrease platelet adhesion while exhibiting excellent EPCs biocompatibility.     

纳米结构Ti/TiN涂层对NiTi合金生物相容性的影响

目的：评价纳米结构Ti/TiN涂层对镍钛形状记忆合金（含镍50.6at%）生物相容性的影响,为生物医用纳米结构Ti/TiN涂层表面改性的NiTi合金材料生物安全性提供依据。方法：不影响基体的形状记忆性或超弹性效应的前提下,采用真空过滤电弧离子镀技术,在NiTi合金表面沉积一层纳米结构Ti/TiN涂层,分别对表面改性前和改性后的NiTi合金样品进行体外细胞毒性实验、溶血实验和血小板粘附实验,探索纳米结构Ti/TiN涂层对NiTi合金生物相容性的影响。结果：表面具有纳米结构Ti/TiN涂层的NiTi合金无细胞毒性,H9C2（2-1）细胞相容性优于涂层前,细胞形态典型,粘附数量明显大于涂层前。纳米结构Ti/TiN涂层有改善NiTi合金的血液相容性作用,其溶血率从2.1%降至涂层改性后的1.2%,同时,血小板黏附量和聚集程度小于处理前的NiTi合金。结论：纳米Ti/TiN涂层能够显著改善NiTi合金的生物相容性。     

Environment-friendly adhesives for surface bonding of wood-based flooring using natural tannin to reduce formaldehyde and TVOC emission

The objective of this research was to develop environment-friendly adhesives for face fancy veneer bonding of engineered flooring using the natural tannin form bark in the wood. The natural wattle tannin adhesive were used to replace UF resin in the formaldehyde-based resin system in order to reduce formaldehyde and volatile organic compound (VOC) emissions from the adhesives used between plywoods and fancy veneers. PVAc was added to the natural tannin adhesive to increase viscosity of tannin adhesive for surface bonding. For tannin/PVAc hybrid adhesives, 5%, 10%, 20% and 30% of PVAc to the natural tannin adhesives were added. tannin/PVAc hybrid adhesives showed better bonding than the commercial natural tannin adhesive with a higher level of wood penetration. The initial adhesion strength was sufficient to be maintained within the optimum initial tack range. The standard formaldehyde emission test (desiccator method), field and laboratory emission cell (FLEC) and VOC analyzer were used to determine the formaldehyde and VOC emissions from engineered flooring bonded with commercial the natural tannin adhesive and tannin/PVAc hybrid adhesives. By desiccator method and FLEC, the formaldehyde emission level of each adhesive showed the similar tendency. All adhesives satisfied the E(1) grade (below 1.5 mg/L) and E(0) grade (below 0.5 mg/L) with UV coating. VOC emission results by FLEC and VOC analyzer were different with the formaldehyde emission results. TVOC emission was slightly increased as adding PVAc.     

Biocompatibility and Favorable Response of Mesenchymal Stem Cells on Fibronectin-Gold Nanocomposites

A simple surface modification method, comprising of a thin coating with gold nanoparticles (AuNPs) and fibronectin (FN), was developed to improve the biocompatibility required for cardiovascular devices. The nanocomposites from FN and AuNPs (FN-Au) were characterized by the atomic force microscopy (AFM), UV-Vis spectrophotometry (UV-Vis), and Fourier transform infrared spectroscopy (FTIR). The biocompatibility of the nanocomposites was evaluated by the response of monocytes and platelets to the material surface in vitro. FN-Au coated surfaces demonstrated low monocyte activation and platelet activation. The behavior of human umbilical cord-derived mesenchymal stem cells (MSCs) on FN-Au was further investigated. MSCs on FN-Au nanocomposites particularly that containing 43.5 ppm of AuNPs (FN-Au 43.5 ppm) showed cell proliferation, low ROS generation, as well as increases in the protein expression levels of matrix metalloproteinase-9 (MMP-9) and endothelial nitric oxide synthase (eNOS), which may account for the enhanced MSC migration on the nanocomposites. These results suggest that the FN-Au nanocomposite thin film coating may serve as a potential and simple solution for the surface modification of blood-contacting devices such as vascular grafts.     

Effects of Type I Collagen Degradation on the Durability of Three Adhesive Systems in the Early Phase of Dentin Bonding

ObjectiveThis study was designed to evaluate the effects of type I collagen degradation on the durability of three adhesive systems in the early phase of dentin bonding.MethodsBonded dentin specimens were prepared using three different types of adhesive systems. Micro-tensile bond strength and degradation of collagen were tested before, and after 1 month or 4 months of aging in artificial saliva. The relationship between micro-tensile bond strength and collagen degradation was analyzed by calculating their Pearson’s correlation coefficient.ResultsAging induced time-dependent reduction in micro-tensile bond strengths for all the tested adhesive systems, although such reduction for the single-step self-etching adhesive G-Bond (GB) was not statistically significant. The bond strength of the two-step self-etching primer adhesive system Clearfil SE Bond (SEB) was similar to that of the two-step etch-and-rinse self-priming adhesive system Single Bond 2 (SB), and they were both significantly reduced after one or four months of aging. A negative correlation was found between the degree of collagen degradation and magnitude of micro-tensile bond strength (r = - 0.65, p = 0.003). The Pearson’s correlation coefficient was 0.426, indicating that 42.6% of the aging-induced reduction in bond strength can be explained by the degradation of collagen.ConclusionsIn the early phase of dentin bonding, there was a negative correlation between the degree of collagen degradation and the magnitude of micro-tensile bond strength. The reduction of bond strength was accompanied by the degradation of collagen. These results provide evidence for the causative relationship between the degradation of collagen and the deterioration of dentin-adhesive interface.     

Development of biodegradable scaffolds based on patient-specific arterial configuration

Biodegradable scaffolds are of great value in tissue engineering. We have developed a method for fabricating patient-specific vascular scaffolds from a biocompatible and biodegradable polymer, poly(L-lactide-co-epsilon-caprolactone). This method's usefulness is due to flexibility in the choice of materials and vascular configurations. Here, we present a way to fabricate scaffolds of human carotid artery by combining processes of rapid prototyping, lost wax, dip coating, selective dissolution, and salt leaching. The result was the successful development of porous biodegradable scaffolds, with mechanical strength covering the range of human blood vessels (1-3 MPa). Human umbilical vein endothelial cells were also cultured on the scaffolds and their biocompatibility was confirmed by cell growth. The Young's modulus of scaffolds could be controlled by changing polymer concentration and porosity. The wall thickness of the tubular scaffold was also controllable by adjusting polymer concentration and pull-up velocity during dip coating. We believe that this fabrication technique can be applied to patient-specific regeneration of blood vessels.     

In vitro biocompatibility evaluation of a heat‐resistant 3D printing material for use in customized cell culture devices

Additive manufacturing (3D printing) enables the fabrication of highly customized and complex devices and is therefore increasingly used in the field of life sciences and biotechnology. However, the application of 3D‐printed parts in these fields requires not only their biocompatibility but also their sterility. The most common method for sterilizing 3D‐printed parts is heat steam sterilization—but most commercially available 3D printing materials cannot withstand high temperatures. In this study, a novel heat‐resistant polyacrylate material for high‐resolution 3D Multijet printing was evaluated for the first time for its resistance to heat steam sterilization and in vitro biocompatibility with mouse fibroblasts (L929), human embryonic kidney cells (HEK 293E), and yeast (Saccharomyces cerevisiae (S. cerevisiae)). Analysis of the growth and viability of L929 cells and the growth of S. cerevisiae confirmed that the extraction media obtained from 3D‐printed parts had no negative effect on the aforementioned cell types, while, in contrast, viability and growth of HEK 293E cells were affected. No different effects of the material on the cells were found when comparing heat steam sterilization and disinfection with ethanol (70%, v/v). In principle, the investigated material shows great potential for high‐resolution 3D printing of novel cell culture systems that are highly complex in design, customized and easily sterilizable—however, the biocompatibility of the material for other cell types needs to be re‐evaluated.     

预酸蚀乳牙牙本质对自酸蚀粘接系统粘接强度的影响

目的:探讨预酸蚀乳牙牙本质对自酸蚀粘接系统粘接强度的影响。方法:随机选取28颗健康乳磨牙,磨除颊舌面釉质,暴露牙本质粘接面,沿近远中向劈开形成56个样本,随机分为7组(n=8)。直接涂布组(A1组,A2组和A3组)分别涂布AdperTM Easy One(AEO),Xeno-V(XV)和OptiBond All In One(AIO)三种自酸蚀粘接剂,预酸蚀组(B1组,B2组和B3组)在涂布三种自酸蚀粘接剂前先使用35%磷酸酸蚀乳牙牙本质15 s,对照组(C组)使用Prime&Bond NTTM(NT)全酸蚀粘接剂,每个样本用Z350复合树脂堆砌成直径为3 mm的树脂小柱,通过剪切试验测试剪切粘接强度,并通过扫描电子显微镜观察断裂表面形态。结果:B1组,B2组的剪切粘接强度值明显高于A1组,A2组(P<0.001);B3组与A3组的剪切粘接强度值比较无明显差别(P=0.94)。A2组的剪切粘接强度值低于C组(P<0.05);B1组的剪切粘接强度值明显高于C组(P<0.001)。扫描电镜观察结果显示各组试件断裂面形态多为牙本质和复合树脂界面破坏。直接涂布组(A1组,A2组和A3组)断裂多发生在混合层的底部,树脂突较少且低于小管口。B1组和B2组试件断裂面可见多数牙本质小管被树脂突填满,断裂多发生在混合层的中上部。B3组试件断裂面可见牙本质小管空虚,树脂突较少。结论:预酸蚀乳牙牙本质可以提高AEO,XV两种自酸蚀粘接剂的剪切粘接强度。自酸蚀粘接剂处理乳牙牙本质可以达到全酸蚀粘接剂处理的粘接强度,但应用自酸蚀粘接剂前预酸蚀乳牙牙本质可以获得更高的粘接强度。     

Data on the push–out bond strength of three different root canal treatment sealers

It is of interest to document data on the push – out bond strength of three different root canal treatment sealers such as MTA Fillapex (MTA based), AH plus (Epoxy Resin based) and Apexit plus (Calcium hydroxide based). Forty-five freshly extracted human maxillary central incisors with closed apices were selected randomly. All the teeth were sectioned at cement-enamel junction using a diamond disc before starting the root canal preparation to obtain root length of 12 mm. All teeth were instrumented using ProTaper rotary instruments. 5.25% sodium hypochlorite was used for irrigation between instrumentation followed by 17% EDTA, and final rinse by saline. Obturation procedures were done using the gutta-percha single cone technique. 45 roots were randomly assigned to 3 groups of 15 for obturation with gutta-percha cones and 1 of the 3 sealers (n=15). Group 1 = MTA Fillapex sealer + gutta-percha: Group 2 = AH plus sealer + gutta-percha:Group 3 = Apexit plus sealer + gutta-percha. The roots were sectioned horizontally to its canal into 3 sections: Coronal, Mid-root and Apical-thirds using a precision cutting machine, with a thickness of 3 mm. The specimens were subjected to push-out test using a universal testing machine that carried a plunger. The loading speed was 1mm/min until the dislodgment of the material occurred. The independent t- test was used to compare the mean scores among the study groups. The level of significance was set at 5% for all tests. After the push-out bond strength test, each sample was evaluated under stereomicroscope (40x) to determine the mode of failure and recorded as one of the following categories: adhesive, cohesive or mixed. The observations thus obtained were subjected to statistical analysis using Student - t test. AH Plus showed significantly higher values than MTA Fillapex and Apexit plus (p < 0.05). Amongst the push-out bond strength AH Plus sealer showed significant difference from MTA Fillapex and Apexit plus groups. There was no significant difference between MTA Fillapex and Apexit plus however (p>0.05). Microscopic analysis displayed that the majority of the modes were cohesive failures for AH Plus, adhesive failures for MTA Fillapex and mixed failures for Apexit Plus. . Thus, AH Plus had the highest bond strength and MTA Fillapex had the lowest bond strength to root dentin. Mean push-out bond strength values were ranked as follows; AH Plus >Apexit Plus > MTA Fillapex. Microscopic analysis displayed that the majority of the modes were cohesive failures of AH Plus, adhesive failures for MTA Fillapex and mixed failures for Apexit Plus.     

Biocompatibility of nano-hydroxyapatite/Mg-Zn-Ca alloy composite scaffolds to human umbilical cord mesenchymal stem cells from Wharton’s jelly in vitro

Seeding cells and scaffolds play pivotal roles in bone tissue engineering and regenerative medicine.Wharton’s jelly-derived mesenchymal stem cells(WJCs)from human umbilical cord represent attractive and promising seeding cells in tissue regeneration and engineering for treatment applications.This study was carried out to explore the biocompatibility of scaffolds to seeding cells in vitro.Rod-like nano-hydroxyapatite(RN-HA)and flake-like micro-hydroxyapatite(FM-HA)coatings were prepared on Mg-Zn-Ca alloy substrates using micro-arc oxidation and electrochemical deposition.WJCs were utilized to investigate the cellular biocompatibility of Mg-Zn-Ca alloys after different surface modifications by observing the cell adhesion,morphology,proliferation,and osteoblastic differentiation.The in vitro results indicated that the RN-HA coating group was more suitable for cell proliferation and cell osteoblastic differentiation than the FM-HA group,demonstrating better biocompatibility.Our results suggested that the RN-HA coating on Mg-Zn-Ca alloy substrates might be of great potential in bone tissue engineering.     

A new integrated concept for the improved preparation of sections of fresh or frozen tissue for light microscope histochemistry

In order to obtain thin sections of plant tissues which combined good morphological preservation and the preservation of the substances and enzyme activities in the tissues, a concept of section preparation by external stabilization was developed. The main components are as follows: appropriate supporting medium; surface coating before each sectioning process, the coating being either non-permanent, permanent, or semi-permanent; suitable techniques for affixing the coated sections to the slides using either pressure-sensitive adhesive or solvent-based adhesive; and mounting media with defined refractive indices (preferably UV-curable, water-soluble monomers). By this approach, sections exhibiting excellent morphological and physiological preservation were obtained using either a cryostat at -30 degrees C or a rotary microtome at room temperature.     

Sterilization of sealed PVDF pouches containing decellularized scaffold by electrical stimulation

A terminal sterilization process for tissue engineering products, such as allografts and biomaterials is necessary to ensure complete removal of pathogenic microorganisms such as the bacteria, fungi, and viruses. However, it can be difficult to sterilize allografts and artificial tissue models packaged in wet conditions without deformation. In this study, we investigated the sterilization effects of electrical stimulation (ES) and assessed its suitability by evaluating sterility assurance levels in pouches at a constant current. Stability of polyvinylidene fluoride pouches was determined by a sterility test performed after exposure to five microorganisms (Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa, Escherichia coli, and Candida albicans) for 5 days; the sterility test was also performed with decellularized human dermal tissues inoculated with the five microorganisms. Sterilization using ES inactivated microorganisms both inside and outside of sealed pouches and caused no damage to the packaged tissue. Our results support the development of a novel system that involves ES sterilization for packaging of implantable biomaterials and human derived materials.     

Effect of methods of sterilization on thermoplastics with special reference to modified surfaces]

For materials intended for use in the medical setting their sterilizability is an indispensable prerequisite. In the case of most polymers the usual sterilization methods result in changes that even extend to cleavage of the polymer chains. A particular problem in this respect are the surfaces modified for improved biocompatibility investigated in the present study, which are characterised by enlarged contact areas. For this reason, possible changes to three different thermoplastics commonly used for medical applications (polyethylene, thermoplastic polyurethane, polycarbonate) were investigated. Steam, gas and radiation were used for sterilization. Tensile tests were employed to identify changes in mucosal characteristics caused by different sterilization techniques irrespective of the surface modification. Sterilization-related changes to the structure of the modified surfaces were investigated with the scanning electron microscope (SEM). Differential thermo analysis (DTA) was used to determine changes in the thermal characteristics of the plastics. Clear tendencies with regard to the behaviour of the plastics after sterilization with various techniques were found. A general statement about the compatibility of plastic materials with a specific sterilization method is not possible on the basis of this study. For every new polymeric product used for medical purposes, the characteristics required must first be defined and compliance with the permissible variations of these characteristics investigated for each of the various sterilization techniques available.     

Validation and Substantiation of 25 kGy as Sterilization Dose for Lyophilized Human Amnion Membrane

The validation and substantiation of sterilization dose for lyophilized human amnion membrane by gamma irradiation delivered by Co⁶⁰ source were investigated. The validation experiments were conducted according to ISO 13409 method B. A total of 120 human amnion membranes were collected. Of these, 10 membranes were used for estimation of bioburden and 20 membranes were used for the individual sterility test at verification dose. The average bioburden per product unit with sample item portion (SIP = 1) for lyophilized human amnion membrane was 572 cfu. The verification dose experiments were done at dose of 8.1 kGy and the results of sterility tests showed that human amnion membrane got one positive. Consequently, the sterilization dose of 25 kGy was confirmed and substantiated.     

Optimization of an Aqueous Tablet-Coating Process Containing Carboxymethylated Cassia fistula Gum

The present investigation was aimed at developing and optimizing a simple aqueous tablet-coating formulation and its process. 5-Fluorouracil (5-FU) was used to ascertain the relative lipophilic/hydrophilic behavior of the coating system. Optimization was performed by evaluating the adhesive force strength and cohesive force strength of the tablet coat using a texture analyzer. The in vitro release of 5-FU was found to decrease with an increase in (tablet surface-coat) adhesive force strength. The (tablet-tablet) cohesive force strength was reduced by the addition of magnesium silicate to the coating solution. The addition of magnesium silicate (0.2% w/v) to the carboxymethyl Cassia fistula gum-chitosan (CCG-CH) coating surface significantly inhibited the release of 5-FU possibly due to an increase in the hydrophobic character of the coated tablet surface. This was possible by coating cohesive force strength reduction coating compositions (CCG-CH (70:30) and 0.3% magnesium silicate). Further, the FTIR-ATR and DSC analyses suggested the pivotal role of magnesium silicate in modifying the release of 5-FU from CCG-CH-coated tablets due to hydrogen bonding of its Si-O-Si or Mg-O groups with -OH moieties of CCG-CH.     

Direct identification of disulfide bond linkages in human insulin-like growth factor I (IGF-I) by chemical synthesis

The primary structure of human IGF-I, except for the disulfide bond system, has been reported by Rinderknecht and Humbel. IGF-I afforded the corresponding characteristic peptide fragment on V8 protease digestion, which contained Cys6, Cys47, Cys48, and Cys52. Two possible fragments, Type I with Cys6-Cys47 and Cys48-Cys52, and Type II with Cys6-Cys48 and Cys47-Cys52, were synthesized. The disulfide bond system of IGF-I was unequivocally determined to be the Type II form along with Cys18-Cys61. Interestingly, the Type I system was included in the disulfide bond isomer produced as the main by-product in the refolding step on IGF-I synthesis by the recombinant DNA method.     

骨髓基质细胞与交联明胶-聚羟基丁酸酯膜的生物相容性研究

目的研究生物材料交联明胶-聚羟基丁酸酯膜与骨髓基质细胞的生物相容性,探讨新型材料在骨组织工程中的应用前景。方法体外培养兔骨髓基质细胞,分别接种于G-PHB(交联明胶-聚羟基丁酸酯)、PHB(聚羟基丁酸酯)和G(交联明胶)材料膜片。采用MTT法检测细胞增殖活性,体视学方法检测细胞粘附能力,荧光双染法检测细胞完整性,扫描电镜观察细胞-材料界面。结果MTT检测发现G-PHB组增殖活性最强,而且表现为最佳的细胞粘附特性,与对照组比较差异有显著性意义。各组细胞完整性分析没有发现显著性差异。扫描电镜观察显示,G-PHB组细胞粘附及铺展良好,优于其他各组。结论交联后的生物降解膜材料G-PHB与BMSCs细胞的体外相容性明显优于单纯膜材料PHB和明胶,在骨组织工程学领域具有良好的研究价值和应用潜力。