In vitro degradation and cytocompatibility evaluation of novel soy and sodium caseinate-based membrane biomaterials

Soy- and casein-based membranes are newly proposed materials disclosing a combination of properties that might allow for their use in a range of biomedical applications. Two of the most promising applications are drug delivery carrier systems and wound dressing membranes. As for all newly proposed biomaterials, a cytotoxic scanning must be performed as a preliminary step in the process of the determination of the compatibility with biological systems (biocompatibility). In this study, the cytotoxicity of both soy- and casein-based protein biomaterials has been evaluated and correlated with the materials degradation behavior. It was possible to show, through morphological and biochemical tests that these natural origin materials do not exert any cytotoxic effect over cells, and in some cases can in fact enhance cell proliferation. The different treatments to which the membranes were subjected during their processing (that include crosslinking with glyoxal and tannic acid, and physical modification by thermal treatment) seemed to have a clear effect both on the materials mechanical properties and on their in vitro biological behavior.     

In vitro cytocompatibility and corrosion resistance of zinc-doped hydroxyapatite coatings on a titanium substrate

To improve biocompatibility and corrosion resistance during the initial implantation stage, zinc-substituted hydroxyapatite (ZnHAp) coating was fabricated on pure titanium by the electrolytic deposition method. The morphology, microstructure and chemical composition of the coating were investigated by X-ray diffraction, scanning electron microscopy, energy-dispersive X-ray analysis and Fourier transform infrared spectroscopy. The prepared ZnHAp crystals were calcium deficient and were carbonated owing to the incorporation of some Zn²⁺. This incorporation of Zn²⁺ into the HAp significantly reduced porosity and caused the coating to become noticeably denser. In addition, the Zn²⁺ ions were homogeneously distributed in the coating. The potentiodynamic polarisation test revealed that the ZnHAp-coated surface showed superior corrosion resistance over that of the HAp-coated surface and bare Ti. The in vitro bioactivity was evaluated in a simulated body fluid, which revealed that the ZnHAp coating can rapidly induce bone-like apatite formation of nuclear and growth features. In addition, the cell response tests showed that the MC3T3-E1 cells on the ZnHAp coating clearly enhanced the in vitro cytocompatibility of Ti compared with the same cells on HAp coating. ZnHAp coating was thus beneficial for improving biocompatibility.     

Cytocompatibility property evaluation of gas pressure sintered SiAlON-SiC composites with L929 fibroblast cells and Saos-2 osteoblast-like cells

Although the oxide ceramics have widely been investigated for their biocompatibility, non-oxide ceramics, such as SiAlON and SiC are yet to be explored in detail. Lack of understanding of the biocompatibility restricts the use of these ceramics in clinical trials. It is hence, essential to carry out proper and thorough study to assess cell adhesion, cytocompatibility and cell viability on the non-oxide ceramics for the potential applications. In this perspective, the present research work reports the cytocompatibility of gas pressure sintered SiAlON monolith and SiAlON-SiC composites with varying amount of SiC, using connective tissue cells (L929) and bone cells (Saos-2). The quantification of cell viability using MTT assay reveals the non-cytotoxic response. The cell viability has been found to be cell type dependent. An attempt has been made to discuss the cytocompatibility of the developed composites in the light of SiC content and type of sinter additives.     

胶原模拟多肽对L929细胞粘附的影响

设计合成了3种模拟胶原三螺旋结构或/和整合素识别位点的胶原模拟多肽(CMP),对其进行体外细胞相容性评价,研究其对小鼠成纤维细胞(L929)生长、粘附的影响。实验证实,3种CMP对成纤维细胞生长无明显的细胞毒性反应;3种包被胶原模拟多肽的基底均能在一定程度上促进细胞粘附、生长,具有良好的细胞粘附率和细胞附着形态,其中包含三螺旋结构和整合素识别位点的CMP27具有更好的促粘附效果,细胞粘附数量和形态与胶原接近。初步研究结果证实,胶原三螺旋结构与整合素识别位点共同作用促进L929细胞粘附。因此,CMP可以有效促进细胞粘附,有望作为粘附剂应用于生物医学领域,可为设计以多肽为基础的生物活性材料提供新的研究思路。     

Human monocyte-derived macrophages and dendritic cells as targets for biomaterial cytocompatibility studies using an improved in vitro culture system

Since macrophage plays a key role in the biocompatibility process, neoplastic macrophage cell lines and human blood monocytes are commonly used as target cells for in vitro biomaterial tolerance evaluation. However, tumor cells profoundly differ from normal tissue cells and monocytes are only precursors of macrophages. It has become possible to generate recently, under adherent-free conditions, fully mature macrophages and dendritic cells from human blood monocytes in the presence of GM-CSF and GM-CSF + IL4 respectively. In the present work, we examined the effects of titanium-alloy on morphology, adhesion, cell phenotype and TNF- release activity of such differentiated cells grown in hydrophobic teflon bags. Scanning electron microscopy showed that macrophages substantially adhered and spread on titanium-alloy surface throughout the culture period, whereas only a few dendritic cells were adherent. The phenotype of both cell types remained unchanged in the presence of the tested material. However, titanium-alloy stimulated the secretion of TNF- by the macrophages of some donors. This model of culture may offer new insights into the biomaterial evaluation and may be useful for studying individual responses induced by biomaterials. © 2001 Kluwer Academic Publishers     

In vivo andin vitro evaluation of a biomachinable glassceramic

In this paper we describe the toxicological tests done on a machinable glassceramic, prepared at the Defence Metallurgical Research Laboratory. Tests show no toxicity and biocompatibility is inferred.     

In vitro evaluation of surface topographic changes and nickel release of lingual orthodontic archwires

The objective of the article is to study surface topographic changes and nickel release in lingual orthodontic archwires in vitro. Stainless steel (SS), nickel–titanium (NiTi) and copper–nickel–titanium (CuNiTi) lingual orthodontic archwires were studied using atomic absorption spectrometry for nickel release after immersion in a saline solution. Surface roughness changes were measured using atomic force microscopy. Differences between groups were analyzed using independent sample t-tests. Statistically significant changes in roughness were seen in all archwires except NiTi. Surface changes were most severe in the CuNiTi alloy. SS archwires released the highest amount of nickel. In conclusion, only roughness changes in CuNiTi archwires seemed to be clinically significant. The amount of nickel released for all archwires tested is below the levels known to cause cell damage.     

In vitro evaluation of betamethasone-loaded nanoparticles

The aim of the present work was to investigate the preparation of nanoparticles as a potential drug carrier in the treatment of various inflammatory diseases. A nanoprecipitation method was used to entrap betamethasone in a poly[ε-caprolactone] matrix. Process parameters such as the initial drug load, the surfactants (polyvinyl alcohol, PVA; sodium cholate, SC), and their concentration in the aqueous phase were analyzed for their influences on particle properties. Particle size changed with increasing surfactant concentrations (PVA: 250 to 400 nm; sodium cholate: 330 to 150 nm) due to changes in interface stability and viscosity of the aqueous phase. The zeta potential was around neutrality with PVA and between - 28 and - 42 mV with SC. Betamethasone encapsulation rates of about 75% and 90% slightly increased with higher surfactant concentration. Drug release profiles exhibited an initial burst release with both surfactants, PVA (8-18%) or SC (25-35%) followed by a sustained release delivering 15% to 40% of the entrapped drug within 48 hours. The present nanoparticulate formulations exhibit promising properties of a colloidal drug carrier for betamethasone. Although SC seems to be advantageous due to its biocompatibility, in terms of sustained drug release pattern, the use of PVA is favorable.     

In vitro evaluation and targeting of E. coli internalized by endothelial cells using albumin microspheres loaded with gentamicin

The purpose of this study was to evaluate the use of microencapsulated form of gentamicin and the traditional solution form for its intracellular bactericidal effect. Bovine serumalbumin (BSA) microspheres loaded with gentamicin were prepared by using Mini Spray Dryer. Human microvascular endothelial cells (HMECs) were exposed to increasing concentrations of Escherichia coli leading to internalization of E. coli. The internalized bacteria were targeted using either the microencapsulated or the solution form of gentamicin. The treatment groups using gentamicin solution form and microsphere form showed almost 46% and 86% inhibition in the growth of the internalized bacteria, respectively.     

In vitro evaluation of a new polymethylmethacrylate cement reinforced with hydroxyapatite

The nature of the orthopedic implant surface affects the interaction between cells and subsequent bone formation. The bone/cement interface in cement-held prostheses is considered to be the main cause of fracture leading to implant revision. It is thought that the introduction of a bioactive phase, such as hydroxyapatite (HA), to cement may permit a stronger implant by encouraging direct bone apposition rather than encapsulation of the implant by fibrous tissue. Thus, a poly(methylmethacrylate) (PMMA) cement incorporating 17.5% HA by weight has been investigated. In this study, in order to analyze the interaction at the cellular level, the in vitro biological response of the HA/PMMA to a similar PMMA without HA incorporation has been studied. Primary human osteoblast-like cells (HOB) were used as they are a model of the cell type the cements might encounter in vivo. Cell proliferation and growth were assessed by measurement of total cellular DNA and tritiated thymidine ([³H]-TdR) incorporation. Alkaline phosphatase (ALP) production was measured as an indicator of HOB phenotype upon the cements. The results showed that HA/PMMA was a better substrate for HOB cells, resulting in increased proliferation and ALP activity. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) showed that HOB cells cultured on the HA-filled PMMA preferentially anchored to HA particles exposed at the cement surface, with a close intimacy observed between HA and HOB cells.     

In vitro haematic proteins adsorption and cytocompatibility study on acrylic copolymer to realise coatings for drug-eluting stents

In the present paper, a preliminary in vitro analysis of biocompatibility of newly-synthesised acrylic copolymers is reported. In particular, with the aim to obtain coatings for drug-eluting stents, blood protein absorption and cytocompatibility were studied. For protein absorption tests, bovine serum albumin and bovine plasma fibrinogen were considered. Cytocompatibility was tested using C2C12 cell line as model, analysing the behaviour of polymeric matrices and of drug-eluting systems, obtained loading polymeric matrices with paclitaxel, an anti-mitotic drug, in order to evaluate the efficacy of a pharmacological treatment locally administered from these materials. Results showed that the amount of albumin absorbed was greater than the amount of fibrinogen (comprised in the range of 70%–85% and 10%–22% respectively) and it is a good behaviour in terms of haemocompatibility. Cell culture tests showed good adhesion properties and a relative poor proliferation. In addition, a strong effect related to drug elution and a correlation with the macromolecular composition were detected. In this preliminary analysis, tested materials showed good characteristics and can be considered possible candidates to obtain coatings for drug-eluting stents.     

In vitro cytotoxicity of surface modified bismuth nanoparticles

This paper describes in vitro cytotoxicity of bismuth nanoparticles revealed by three complementary assays (MTT, G6PD, and calcein AM/EthD-1). The results show that bismuth nanoparticles are more toxic than most previously reported bismuth compounds. Concentration dependent cytotoxicities have been observed for bismuth nanoparticles and surface modified bismuth nanoparticles. The bismuth nanoparticles are non-toxic at concentration of 0.5 nM. Nanoparticles at high concentration (50 nM) kill 45, 52, 41, 34 % HeLa cells for bare nanoparticles, amine terminated bismuth nanoparticles, silica coated bismuth nanoparticles, and polyethylene glycol (PEG) modified bismuth nanoparticles, respectively; which indicates cytotoxicity in terms of cell viability is in the descending order of amine terminated bismuth nanoparticles, bare bismuth nanoparticles, silica coated bismuth nanoparticles, and PEG modified bismuth nanoparticles. HeLa cells are more susceptible to toxicity from bismuth nanoparticles than MG-63 cells. The simultaneous use of three toxicity assays provides information on how nanoparticles interact with cells. Silica coated bismuth nanoparticles can damage cellular membrane yet keep mitochondria less influenced; while amine terminated bismuth nanoparticles can affect the metabolic functions of cells. The findings have important implications for caution of nanoparticle exposure and evaluating toxicity of bismuth nanoparticles.     

In vitro evaluation of hydroxyapatite reinforced polyhydroxybutyrate composite

Particulate hydroxyapatite (HA) was incorporated into polyhydroxybutyrate (PHB) to form a bioactive and biodegradable composite for applications in hard tissue replacement and regeneration. HA/PHB composite containing 10, 20, and 30 vol.% of HA was made for in vitro evaluation. In vitro studies were conducted using an acellular simulated body fluid (SBF). Composite specimens were immersed in SBF at 37 °C for various periods of time prior to surface analysis and mechanical testing. Results obtained from scanning electron microscopic (SEM) examination, thin film X-ray diffraction (TF-XRD) analysis, and Fourier transform infrared (FTIR) spectroscopy showed that a layer of bone-like apatite formed within a short period on HA/PHB composite after its immersion in SBF, demonstrating high in vitro bioactivity of the composite. The bioactivity and mechanical properties of the composite could be changed by varying the amount of HA in the composite. Dynamic mechanical analysis (DMA) revealed that the storage modulus (E′) of HA/PHB composite increased initially with immersion time in SBF, due to apatite formation on composite surface and decreased after prolonged immersion in SBF, indicating degradation of the composite in a simulated body environment. HA/PHB composite thus has the potential for its intended applications.     

Synthesis of a thermoresponsive platinum nanocomposite using a three-layer onion-like polymer as template

Using a well-designed three-layer onion-like polymer as template, a one-pot procedure that led to stable, narrow-sized and thermoresponsive Pt nanocomposites is described. The polymer consists of an outer shell of thermoresponsive poly(N-isopropylacrylamide), an inner shell of crosslinked poly(N,N-dimethylaminoethyl acrylate) and a hyperbranched polyglycerol core. The core is physically trapped by the shell, with a few thiol groups located on the interface between the core and the shell. The polymer is used as a template for the synthesis of platinum nanoparticles, ¹H NMR and TEM analyses suggest that the in-situ produced, narrow-sized Pt nanoparticle is loaded in the core part of the polymer so that the nanocomposite retains thermoresponsive activity.     

In vitro evaluation of artificial ageing on surface properties and early Candida albicans adhesion to prosthetic resins

Objective The aim of this in vitro study was to examine potential changes and influences of prosthetic resin surface properties on Candida albicans adhesion after surface treatment or artificial ageing. Materials and methods Standardized specimens of a denture base resin and a veneering composite were prepared, polished, and randomly subjected to different surface roughness treatments or artificial ageing protocols (storage in ethanol or artificial saliva for 7/90 d, thermocycling). Surface roughness (Ra) and surface free energy were determined prior and after each treatment. Specimens were incubated with phosphate buffered saline or whole saliva for 2 h at 37°C, and later with Candida albicans suspension (2.5 h, 37°C). Adherent viable fungi were quantified using a bioluminescence assay. Results Artifical ageing did not affect substratum surface roughness, yet slight increases in substratum surface free energy and significant increases in Candida albicans adhesion were observed. Saliva coating marginally influenced Candida albicans adherence to reference and surface treated specimens, yet more pronounced differences in Candida albicans adhesion between the various artificially aged specimens were found. Conclusion No correlation between substratum surface roughness or surface free energy and Candida albicans adhesion could be established.     

In vitro bioactivity, cytocompatibility, and antibiotic release profile of gentamicin sulfate-loaded borate bioactive glass/chitosan composites

Borate bioactive glass-based composites have been attracting interest recently as an osteoconductive carrier material for local antibiotic delivery. In the present study, composites composed of borate bioactive glass particles bonded with a chitosan matrix were prepared and evaluated in vitro as a carrier for gentamicin sulfate. The bioactivity, degradation, drug release profile, and compressive strength of the composite carrier system were studied as a function of immersion time in phosphate-buffered saline at 37 °C. The cytocompatibility of the gentamicin sulfate-loaded composite carrier was evaluated using assays of cell proliferation and alkaline phosphatase activity of osteogenic MC3T3-E1 cells. Sustained release of gentamicin sulfate occurred over ~28 days in PBS, while the bioactive glass converted continuously to hydroxyapatite. The compressive strength of the composite loaded with gentamicin sulfate decreased from the as-fabricated value of 24 ± 3 MPa to ~8 MPa after immersion for 14 days in PBS. Extracts of the soluble ionic products of the borate glass/chitosan composites enhanced the proliferation and alkaline phosphatase activity of MC3T3-E1 cells. These results indicate that the gentamicin sulfate-loaded composite composed of chitosan-bonded borate bioactive glass particles could be useful clinically as an osteoconductive carrier material for treating bone infection.     

涤纶材料表面固定水蛭素及其血液相容性研究

利用紫外辐照仪在聚对苯二甲酸乙二醇酯(涤纶,PET)膜表面接枝聚丙烯酸(PET-AA)长链分子,再通过化学方法进一步固定水蛭素,获得的改性表面的X光电子能谱分析表明水蛭素分子被有效地固定在PET表面.改性表面的水接触角降低,亲水性得到改善.体外血小板粘附实验的结果显示,固定了水蛭素的PET薄膜的血液相容性比未改性的PET膜有明显的改善.     

In vitro evaluation of bioactive strontium-based ceramic with rabbit adipose-derived stem cells for bone tissue regeneration

The development of bone replacement materials is an important objective in the field of orthopaedic surgery. Due to the drawbacks of treating bone defects with autografts, synthetic bone graft materials have become optional. So in this work, a bone tissue engineering approach with radiopaque bioactive strontium incorporated calcium phosphate was proposed for the preliminary cytocompatibility studies for bone substitutes. Accumulating evidence indicates that strontium containing biomaterials promote enhanced bone repair and radiopacity for easy imaging. Hence, strontium calcium phosphate (SrCaPO₄) and hydroxyapatite scaffolds have been investigated for its ability to support and sustain the growth of rabbit adipose-derived mesenchymal stem cells (RADMSCs) in vitro. They were characterized via Micro-CT for pore size distribution. Cells used were isolated from New Zealand White rabbit adipose tissue, characterized by FACS and via differentiation into the osteogenic lineage by alkaline phosphatase, Masson’s trichome, Alizarin Red and von Kossa staining on day 28. Material-cell interaction was observed by SEM imaging of cell morphology on contact with material. Live–Dead analysis was done by confocal laser scanning microscopy and cell cluster analysis via μCT. The in vitro biodegradation, elution and nucleation of apatite formation of the material was evaluated using simulated body fluid and phosphate buffered saline in static regime up to 28 days at 37 °C. These results demonstrated that SrCaPO₄ is a good candidate for bone tissue engineering applications and with osteogenically-induced RADMSCs, they may serve as potential implants for the repair of critical-sized bone defects.