Development of sputtered nanoscale titanium oxide coating on osseointegrated implant devices and their biological evaluation

Nanoscale titanium oxide (TiO₂) coating was deposited on titanium (Ti) disks and Ti dental implants using r.f. magnetron sputtering technique. The coating was characterized using grazing angle X-ray diffraction (XRD), scanning electron microscopy (SEM), atomic force microscopy (AFM), contact angle measurement, profilometry and nano-scratch test. The coating also was evaluated with in vitro cell culture and in vivo dog femur model. Three groups of samples were prepared, including as-sputtered Ti (AS-Ti), sputtered Ti with a post-deposition heat treatment at 600 °C (SH-Ti) and machined Ti (MA-Ti) as controls. The AS-Ti and SH-Ti were dense surfaces consisting of nanoscale grains of 40 nm and 80 nm, respectively. Post-deposition heat treatment increased the coating adhesion. The SH-Ti and AS-Ti significantly decreased the water contact angles compared to the MA-Ti. The nanoscale AS-Ti and SH-Ti significantly improved cell adhesion within the first hour of incubation compared with the MA-Ti. No significant differences were observed in osteoblast proliferation and differentiation in vitro as well as reverse torque and histology in vivo among the three groups. In the present study, it was not observed that the nanoscale dense TiO₂ coating improved the osseointegration compared to the microscale dense Ti.     

Bone tissue reactions to biomimetic ion-substituted apatite surfaces on titanium implants

The aim of this study was to evaluate the bone tissue response to strontium- and silicon-substituted apatite (Sr-HA and Si-HA) modified titanium (Ti) implants. Sr-HA, Si-HA and HA were grown on thermally oxidized Ti implants by a biomimetic process. Oxidized implants were used as controls. Surface properties, i.e. chemical composition, surface thickness, morphology/pore characteristics, crystal structure and roughness, were characterized with various analytical techniques. The implants were inserted in rat tibiae and block biopsies were prepared for histology, histomorphometry and scanning electron microscopy analysis. Histologically, new bone formed on all implant surfaces. The bone was deposited directly onto the Sr-HA and Si-HA implants without any intervening soft tissue. The statistical analysis showed significant higher amount of bone–implant contact (BIC) for the Si-doped HA modification (P = 0.030), whereas significant higher bone area (BA) for the Sr-doped HA modification (P = 0.034), when compared with the non-doped HA modification. The differences were most pronounced at the early time point. The healing time had a significant impact for both BA and BIC (P < 0.001). The present results show that biomimetically prepared Si-HA and Sr-HA on Ti implants provided bioactivity and promoted early bone formation.     

The effects of vibration loading on adipose stem cell number,viability and differentiation towards bone-forming cells

Mechanical stimulation is an essential factor affecting the metabolism of bone cells and their precursors. We hypothesized that vibration loading would stimulate differentiation of human adipose stem cells (hASCs) towards bone-forming cells and simultaneously inhibit differentiation towards fat tissue. We developed a vibration-loading device that produces 3g peak acceleration at frequencies of 50 and 100 Hz to cells cultured on well plates. hASCs were cultured using either basal medium (BM), osteogenic medium (OM) or adipogenic medium (AM), and subjected to vibration loading for 3 h d^–1 for 1, 7 and 14 day. Osteogenesis, i.e. differentiation of hASCs towards bone-forming cells, was analysed using markers such as alkaline phosphatase (ALP) activity, collagen production and mineralization. Both 50 and 100 Hz vibration frequencies induced significantly increased ALP activity and collagen production of hASCs compared with the static control at 14 day in OM. A similar trend was detected for mineralization, but the increase was not statistically significant. Furthermore, vibration loading inhibited adipocyte differentiation of hASCs. Vibration did not affect cell number or viability. These findings suggest that osteogenic culture conditions amplify the stimulatory effect of vibration loading on differentiation of hASCs towards bone-forming cells.     

Remote automated multi-generational growth and observation of an animal in low Earth orbit

The ultimate survival of humanity is dependent upon colonization of other planetary bodies. Key challenges to such habitation are (patho)physiologic changes induced by known, and unknown, factors associated with long-duration and distance space exploration. However, we currently lack biological models for detecting and studying these changes. Here, we use a remote automated culture system to successfully grow an animal in low Earth orbit for six months. Our observations, over 12 generations, demonstrate that the multi-cellular soil worm Caenorhabditis elegans develops from egg to adulthood and produces progeny with identical timings in space as on the Earth. Additionally, these animals display normal rates of movement when fully fed, comparable declines in movement when starved, and appropriate growth arrest upon starvation and recovery upon re-feeding. These observations establish C. elegans as a biological model that can be used to detect changes in animal growth, development, reproduction and behaviour in response to environmental conditions during long-duration spaceflight. This experimental system is ready to be incorporated on future, unmanned interplanetary missions and could be used to study cost-effectively the effects of such missions on these biological processes and the efficacy of new life support systems and radiation shielding technologies.     

Modelling planar polarity of epithelia: the role of signal relay in collective cell polarization

Collective cell polarization is an important characteristic of tissues. Epithelia commonly display cellular structures that are polarized within the plane of the tissue. Establishment of this planar cell polarity requires mechanisms that locally align polarized structures between neighbouring cells, as well as cues that provide global information about alignment relative to an axis of a tissue. In the Drosophila ovary, the cadherin Fat2 is required to orient actin filaments located at the basal side of follicle cells perpendicular to the long axis of the egg chamber. The mechanisms directing this orientation of actin filaments, however, remain unknown. Here we show, using genetic mosaic analysis, that fat2 is not essential for the local alignment of actin filaments between neighbouring cells. Moreover, we provide evidence that Fat2 is involved in the propagation of a cue specifying the orientation of actin filaments relative to the tissue axis. Monte Carlo simulations of actin filament orientation resemble the results of the genetic mosaic analysis, if it is assumed that a polarity signal can propagate from a signal source only through a connected chain of wild-type cells. Our results suggest that Fat2 is required for propagating global polarity information within the follicle epithelium through direct cell–cell contact. Our computational model might be more generally applicable to study collective cell polarization in tissues.     

Microbial synthesis of core/shell gold/palladium nanoparticles for applications in green chemistry

We report a novel biochemical method based on the sacrificial hydrogen strategy to synthesize bimetallic gold (Au)–palladium (Pd) nanoparticles (NPs) with a core/shell configuration. The ability of Escherichia coli cells supplied with H₂ as electron donor to rapidly precipitate Pd(II) ions from solution is used to promote the reduction of soluble Au(III). Pre-coating cells with Pd(0) (bioPd) dramatically accelerated Au(III) reduction, with the Au(III) reduction rate being dependent upon the initial Pd loading by mass on the cells. Following Au(III) addition, the bioPd–Au(III) mixture rapidly turned purple, indicating the formation of colloidal gold. Mapping of bio-NPs by energy dispersive X-ray microanalysis suggested Au-dense core regions and peripheral Pd but only Au was detected by X-ray diffraction (XRD) analysis. However, surface analysis of cleaned NPs by cyclic voltammetry revealed large Pd surface sites, suggesting, since XRD shows no crystalline Pd component, that layers of Pd atoms surround Au NPs. Characterization of the bimetallic particles using X-ray absorption spectroscopy confirmed the existence of Au-rich core and Pd-rich shell type bimetallic biogenic NPs. These showed comparable catalytic activity to chemical counterparts with respect to the oxidation of benzyl alcohol, in air, and at a low temperature (90°C).     

Aerobic degradation of nitrobenzene by immobilization of Rhodotorula mucilaginosa in polyurethane foam

Rhodotorula mucilaginosa Z1 capable of degrading nitrobenzene was immobilized in polyurethane foam. The nitrobenzene-degrading capacity of immobilized cells was compared to free cells in batches in shaken culture. Effects of pH and temperature on the nitrobenzene degradation showed that polyurethane-immobilized Z1 had higher tolerances toward acid, alkali, and heat than those of free cells. Kinetic studies revealed that higher concentrations of nitrobenzene were better tolerated and more quickly degraded by polyurethane-immobilized Z1 than by free cells. Moreover, the ability of polyurethane-immobilized Z1 to resist nitrobenzene shock load was enhanced. Experiments on the nitrobenzene degradation in different concentrations of NaCl and in the presence of phenol or aniline demonstrated that polyurethane-immobilized Z1 exhibited higher tolerance toward salinity and toxic chemicals than those of free cells. Immobilization therefore could be a promising method for treating nitrobenzene industrial wastewater. This is the first report on the degradation of nitrobenzene by a polyurethane-immobilized yeast strain.     

Ambiguities in Powder Indexing: the Impact of a Quaternary Lattice Metric Singularity on the Characterization of Mawsonite and Chatkalite

A lattice metric singularity occurs when unit cells defining two (or more) lattices yield the identical set of unique calculated d-spacings. The minerals Mawsonite and Chatkalite are of especial interest as both are characterized by tetragonal unit cells that correspond to the second member of a quaternary lattice metric singularity. This singularity includes lattices that are Cubic I, Tetragonal P, Orthorhombic F, and Orthorhombic P. The Mawsonite and Chatkalite lattices are unique in that they are highly specialized. In each case: (1) the determinative c/a ratio is very near 1/√2, (2) the symmetrical scalars of the reduced form [a · a : b · b : c · c = 1:2:2] have greater specialization than required for the given reduced form type, (3) the tetragonal lattice has derivative lattices of higher symmetry, and (4) the powder pattern is highly compressed. Mawsonite and Chatkalite serve as exemplar-type compounds. Their tetragonal structure has important implications in structure determination using powder diffraction data. First, any cubic I lattice — established solely on the basis of indexing procedures — may actually be tetragonal or orthorhombic. Second, in establishing the lattice of an unknown, results from powder data indexing require routine confirmation by other techniques (e.g., single crystal, optical, etc.).     

Effect of magnetic fields on cryptochrome-dependent responses in Arabidopsis thaliana

The scientific literature describing the effects of weak magnetic fields on living systems contains a plethora of contradictory reports, few successful independent replication studies and a dearth of plausible biophysical interaction mechanisms. Most such investigations have been unsystematic, devoid of testable theoretical predictions and, ultimately, unconvincing. A recent study, of magnetic responses in the model plant Arabidopsis thaliana, however, stands out; it has a clear hypothesis—that seedling growth is magnetically sensitive as a result of photoinduced radical-pair reactions in cryptochrome photoreceptors—tested by measuring several cryptochrome-dependent responses, all of which proved to be enhanced in a magnetic field of intensity 500 μT. The potential importance of this study in the debate on putative effects of extremely low-frequency electromagnetic fields on human health prompted us to subject it to the ‘gold standard’ of independent replication. With experimental conditions chosen to match those of the original study, we have measured hypocotyl lengths and anthocyanin accumulation for Arabidopsis seedlings grown in a 500 μT magnetic field, with simultaneous control experiments at 50 μT. Additionally, we have determined hypocotyl lengths of plants grown in 50 μT, 1 mT and approximately 100 mT magnetic fields (with zero-field controls), measured gene (CHS, HY5 and GST) expression levels, investigated blue-light intensity effects and explored the influence of sucrose in the growth medium. In no case were consistent, statistically significant magnetic field responses detected.     

Investigating the rheological properties of native plant latex

Plant latex, the source of natural rubber, has been of interest to mankind for millennia, with much of the research on its rheological (flow) properties focused towards industrial application. However, little is known regarding the rheology of the native material as produced by the plant, a key factor in determining latex''s biological functions. In this study, we outline a method for rheological comparison between native latices that requires a minimum of preparatory steps. Our approach provides quantitative insights into the coagulation mechanisms of Euphorbia and Ficus latex allowing interpretation within a comparative evolutionary framework. Our findings reveal that in laboratory conditions both latices behave like non-Newtonian materials with the coagulation of Euphorbia latex being mediated by a slow evaporative process (more than 60 min), whereas Ficus appears to use additional biochemical components to increase the rate of coagulation (more than 30 min). Based on these results, we propose two different primary defensive roles for latex in these plants: the delivery of anti-herbivory compounds (Euphorbia) and rapid wound healing (Ficus).     

Surface wettability of plasma SiOx:H nanocoating-induced endothelial cells' migration and the associated FAK-Rho GTPases signalling pathways

Vascular endothelial cell (EC) adhesion and migration are essential processes in re-endothelialization of implanted biomaterials. There is no clear relationship and mechanism between EC adhesion and migration behaviour on surfaces with varying wettabilities. As model substrates, plasma SiO_x:H nanocoatings with well-controlled surface wettability (with water contact angles in the range of 98.5 ± 2.3° to 26.3 ± 4.0°) were used in this study to investigate the effects of surface wettability on cell adhesion/migration and associated protein expressions in FAK-Rho GTPases signalling pathways. It was found that EC adhesion/migration showed opposite behaviour on the hydrophilic and hydrophobic surfaces (i.e. hydrophobic surfaces promoted EC migration but were anti-adhesions). The number of adherent ECs showed a maximum on hydrophilic surfaces, while cells adhered to hydrophobic surfaces exhibited a tendency for cell migration. The focal adhesion kinase (FAK) inhibitor targeting the Y-397 site of FAK could significantly inhibit cell adhesion/migration, suggesting that EC adhesion and migration on surfaces with different wettabilities involve (p)FAK and its downstream signalling pathways. Western blot results suggested that the FAK-Rho GTPases signalling pathways were correlative to EC migration on hydrophobic plasma SiO_x:H surfaces, but uncertain to hydrophilic surfaces. This work demonstrated that surface wettability could induce cellular behaviours that were associated with different cellular signalling events.     

Logic programming to predict cell fate patterns and retrodict genotypes in organogenesis

Caenorhabditis elegans vulval development is a paradigm system for understanding cell differentiation in the process of organogenesis. Through temporal and spatial controls, the fate pattern of six cells is determined by the competition of the LET-23 and the Notch signalling pathways. Modelling cell fate determination in vulval development using state-based models, coupled with formal analysis techniques, has been established as a powerful approach in predicting the outcome of combinations of mutations. However, computing the outcomes of complex and highly concurrent models can become prohibitive. Here, we show how logic programs derived from state machines describing the differentiation of C. elegans vulval precursor cells can increase the speed of prediction by four orders of magnitude relative to previous approaches. Moreover, this increase in speed allows us to infer, or ‘retrodict’, compatible genomes from cell fate patterns. We exploit this technique to predict highly variable cell fate patterns resulting from dig-1 reduced-function mutations and let-23 mosaics. In addition to the new insights offered, we propose our technique as a platform for aiding the design and analysis of experimental data.     

Optical parameters of the tunable Bragg reflectors in squid

Cephalopods (e.g. octopus, squid and cuttlefish) dynamically tune the colour and brightness of their skin for camouflage and communication using specialized skin cells called iridocytes. We use high-resolution microspectrophotometry to investigate individual tunable Bragg structures (consisting of alternating reflectin protein-containing, high-refractive index lamellae and low-refractive index inter-lamellar spaces) in live and chemically fixed iridocytes of the California market squid, Doryteuthis opalescens. This subcellular, single-stack microspectrophotometry allows for spectral normalization, permitting use of a transfer-matrix model of Bragg reflectance to calculate all the parameters of the Bragg stack—the refractive indices, dimensions and numbers of the lamellae and inter-lamellar spaces. Results of the fitting analyses show that eight or nine pairs of low- and high-index layers typically contribute to the observed reflectivity in live cells, whereas six or seven pairs of low- and high-index layers typically contribute to the reflectivity in chemically fixed cells. The reflectin-containing, high-index lamellae of live cells have a refractive index proportional to the peak reflectivity, with an average of 1.405 ± 0.012 and a maximum around 1.44, while the reflectin-containing lamellae in fixed tissue have a refractive index of 1.413 ± 0.015 suggesting a slight increase of refractive index in the process of fixation. As expected, incremental changes in refractive index contribute to the greatest incremental changes in reflectivity for those Bragg stacks with the most layers. The excursions in dimensions required to tune the measured reflected wavelength from 675 (red) to 425 nm (blue) are a decrease from ca 150 to 80 nm for the high-index lamellae and from ca 120 to 50 nm for the low-index inter-lamellar spaces. Fixation-induced dimensional changes also are quantified, leading us to suggest that further microspectrophotometric analyses of this iridocyte system can be used as a model system to quantify the effects of various methods of tissue fixation. The microspectrophotometry technique described can be expected to provide deeper insights into the molecular and physical mechanisms governing other biophotonically active cells and structures.     

Label-free magnetic resonance imaging to locate live cells in three-dimensional porous scaffolds

Porous scaffolds are widely tested materials used for various purposes in tissue engineering. A critical feature of a porous scaffold is its ability to allow cell migration and growth on its inner surface. Up to now, there has not been a method to locate live cells deep inside a material, or in an entire structure, using real-time imaging and a non-destructive technique. Herein, we seek to demonstrate the feasibility of the magnetic resonance imaging (MRI) technique as a method to detect and locate in vitro non-labelled live cells in an entire porous material. Our results show that the use of optimized MRI parameters (4.7 T; repetition time = 3000 ms; echo time = 20 ms; resolution 39 × 39 µm) makes it possible to obtain images of the scaffold structure and to locate live non-labelled cells in the entire material, with a signal intensity higher than that obtained in the culture medium. In the current study, cells are visualized and located in different kinds of porous scaffolds. Moreover, further development of this MRI method might be useful in several three-dimensional biomaterial tests such as cell distribution studies, routine qualitative testing methods and in situ monitoring of cells inside scaffolds.     

Evidence for behavioural thermoregulation by the world's largest fish

Many fishes make frequent ascents to surface waters and often show prolonged surface swimming following descents to deep water. This affinity for the surface is thought to be related to the recovery of body heat lost at depth. We tested this hypothesis using data from time–depth recorders deployed on four whale sharks (Rhincodon typus). We summarized vertical movements into bouts of dives and classified these into three main types, using cluster analysis. In addition to day and night ‘bounce’ dives where sharks rapidly descended and ascended, we found a third type: single deep (mean: 340 m), long (mean: 169 min) dives, occurring in daytime with extremely long post-dive surface durations (mean: 146 min). Only sharks that were not constrained by shallow bathymetry performed these dives. We found a negative relationship between the mean surface duration of dives in the bout and the mean minimum temperature of dives in the bout that is consistent with the hypothesis that thermoregulation was a major factor driving use of the surface. The relationship broke down when sharks were diving in mean minimum temperatures around 25°C, suggesting that warmer waters did not incur a large metabolic cost for diving and that other factors may also influence surface use.     

Conventional Cells—The Last Step Toward General Acceptance of Standard Conventional Cells for the Reporting of Crystallographic Data

In 1969, a seminal section on reduced forms and conventional cells was published in the International Tables for X-Ray Crystallography. The section contains a table that gives a metric classification of the 44 reduced forms. In 2001, this table with appropriate revisions was republished in the Journal of Research of the National Institute of Standards and Technology. An especially valuable feature of the table is that it defines and allows the user to determine a standard conventional cell. Since 1969, there has been an evolution toward acceptance and widespread use of such conventional cells. An inspection of the articles in key crystallographic journals reveals that most cells follow the conventions. However, one major exception remains—the centered monoclinic lattices. In approximately one-third of these cases, non-conventional C-centered cells are used, apparently to avoid the use of I-centered cells. It is recommended that the crystallographic community routinely use the I-centered conventional cell in such cases.     

The biological seal of the implant–soft tissue interface evaluated in a tissue-engineered oral mucosal model

For dental implants, it is vital that an initial soft tissue seal is achieved as this helps to stabilize and preserve the peri-implant tissues during the restorative stages following placement. The study of the implant–soft tissue interface is usually undertaken in animal models. We have developed an in vitro three-dimensional tissue-engineered oral mucosal model (3D OMM), which lends itself to the study of the implant–soft tissue interface as it has been shown that cells from the three-dimensional OMM attach onto titanium (Ti) surfaces forming a biological seal (BS). This study compares the quality of the BS achieved using the three-dimensional OMM for four types of Ti surfaces: polished, machined, sandblasted and anodized (TiUnite). The BS was evaluated quantitatively by permeability and cell attachment tests. Tritiated water (HTO) was used as the tracing agent for the permeability test. At the end of the permeability test, the Ti discs were removed from the three-dimensional OMM and an Alamar Blue assay was used for the measurement of residual cells attached to the Ti discs. The penetration of the HTO through the BS for the four types of Ti surfaces was not significantly different, and there was no significant difference in the viability of residual cells that attached to the Ti surfaces. The BS of the tissue-engineered oral mucosa around the four types of Ti surface topographies was not significantly different.     

Modelling the emergence of polarity patterns for the intercellular transport of auxin in plants

The hormone auxin is actively transported throughout plants via protein machineries including the dedicated transporter known as PIN. The associated transport is ordered with nearby cells driving auxin flux in similar directions. Here, we provide a model of both the auxin transport and of the dynamics of cellular polarization based on flux sensing. Our main findings are: (i) spontaneous intracellular PIN polarization arises if PIN recycling dynamics are sufficiently nonlinear, (ii) there is no need for an auxin concentration gradient and (iii) ordered multi-cellular patterns of PIN polarization are favoured by molecular noise.     

Quantification of vital adherent <Emphasis Type="Italic">Streptococcus sanguinis</Emphasis> cells on protein-coated titanium after disinfectant treatment

The quantification of vital adherent bacteria is challenging, especially when efficacy of antimicrobial agents is to be evaluated. In this study three different methods were compared in order to quantify vital adherent Streptococcus sanguinis cells after exposure to disinfectants. An anaerobic flow chamber model accomplished initial adhesion of S. sanguinis on protein-coated titanium. Effects of chlorhexidine, Betadine^®, Octenidol^®, and ProntOral^® were assessed by quantifying vital cells using Live/Dead BacLight^?, conventional culturing and isothermal microcalorimetry (IMC). Results were analysed by Kruskal–Wallis one-way analysis of variance. Live/dead staining revealed highest vital cell counts (P < 0.05) and demonstrated dose-dependent effect for all disinfectants. Microcalorimetry showed time-delayed heat flow peaks that were proportioned to the remaining number of viable cells. Over 48 h there was no difference in total heat between treated and untreated samples (P > 0.05), indicating equivalent numbers of bacteria were created and disinfectants delayed growth but did not eliminate it. In conclusion, contrary to culturing, live/dead staining enables detection of cells that may be viable but non-cultivable. Microcalorimetry allows unique evaluation of relative disinfectant effects by quantifying differences in time delay of regrowth of remaining vital cells.     

Prediction of heat transfer coefficients for forced convective boiling of N2-hydrocarbon mixtures at cryogenic conditions using artificial neural networks

A key problem faced in the design of heat exchangers, especially for cryogenic applications, is the determination of convective heat transfer coefficients in two-phase flow such as condensation and boiling of non-azeotropic refrigerant mixtures. This paper proposes and evaluates three models for estimating the convective coefficient during boiling. These models are developed using computational intelligence techniques. The performance of the proposed models is evaluated using the mean relative error (mre), and compared to two existing models: the modified Granryd’s correlation and the Silver-Bell-Ghaly method. The three proposed models are distinguished by their architecture. The first is based on directly measured parameters (DMP-ANN), the second is based on equivalent Reynolds and Prandtl numbers (eq-ANN), and the third on effective Reynolds and Prandtl numbers (eff-ANN).The results demonstrate that the proposed artificial neural network (ANN)-based approaches greatly outperform available methodologies. While Granryd's correlation predicts experimental data within a mean relative error mre = 44% and the S-B-G method produces mre = 42%, DMP-ANN has mre = 7.4% and eff-ANN has mre = 3.9%. Considering that eff-ANN has the lowest mean relative error (one tenth of previously available methodologies) and the broadest range of applicability, it is recommended for future calculations. Implementation is straightforward within a variety of platforms and the matrices with the ANN weights are given in the appendix for efficient programming.