Decompression sickness bubbles: Are gas micronuclei formed on a flat hydrophobic surface?

It is a long-standing hypothesis that the bubbles which evolve as a result of decompression have their origin in stable gas micronuclei lodged in hydrophobic crevices, micelles of surface-active molecules, or tribonucleation. Recent findings supported by atomic force microscopy have indicated that tiny, flat nanobubbles form spontaneously on smooth, hydrophobic surfaces submerged in water. We propose that these nanobubbles may be the gas micronuclei responsible for the bubbles that evolve to cause decompression sickness. To support our hypothesis, we used hydrophilic and monolayer-covered hydrophobic smooth silicon wafers. The experiment was conducted in three main stages. Double distilled water was degassed at the low pressure of 5.60 kPa; hydrophobic and hydrophilic silicon wafers were placed in a bowl of degassed water and left overnight at normobaric pressure. The bowl was then placed in the hyperbaric chamber for 15 h at a pressure of 1013 kPa (=90 m sea water). After decompression, bubbles were observed and photographed. The results showed that bubbles only evolved on the hydrophobic surfaces following decompression. There are numerous hydrophobic surfaces within the living body (e.g., in the large blood vessels), which may thus be the sites where nanobubbles that serve as gas micronuclei for bubble evolution following decompression are formed.     

Quantitative PCR of pmoA using a novel reverse primer correlates with potential methane oxidation in Finnish fen

We report a new reverse primer (A621r) for use with A189f in PCR amplification of pmoA alleles in type II methanotrophs. The new primer combination was used to successfully amplify pmoA in peat monolith samples of various depths taken from fen-type peatlands in Finland. In quantitative PCR, pmoA amplicons produced from two sets of three replicate monoliths showed a significant Pearson correlation coefficient (r = 0.77 and 0.61) with methane oxidation potential. The maximum methane oxidation potential and number of pmoA amplicons ranged between 8.8–40.5 μmol g (dry weight)^?1 d^?1 and 5.5 × 10⁷–18.7 × 10⁷ g (wet weight)^?1, respectively, occurring in depths between 10 and 30 cm beneath the surface in the seven individual monoliths used in this study.     

A hairpin aptamer-based electrochemical biosensing platform for the sensitive detection of proteins

An aptamer-based electrochemical sensing platform for the direct protein detection has been developed using IgE and a specifically designed aptamer with hairpin structure as the model analyte and probe sequence, respectively. In the absence of IgE, the aptamer immobilized on an electrode surface forms a large hairpin due to the hybridization of the two complementary arm sequences, and peak currents of redox species dissolved in solution can be achieved. However, the target protein binding can not only cause the increase of the dielectric layer but also trigger the significant conformational switching of the aptamer due to the opening of the designed hairpin structure that pushes the biomolecule layer/electrolyte interface away from the electrode surface, suppressing substantially the electron transfer (eT) and resulting in a strong detection signal. The detection limit of 3.6 × 10^?11 M and linear response range of 5.4 × 10^?11 to 3.6 × 10^?8 M are achieved without any amplifier. The selectivity is confirmed by interference test. More importantly, an innovative concept of adapting intelligently a surface-confined aptamer sequence is introduced, and the limitations of the conventional electrochemical aptasensors have been overcome. The proposed sensing scheme is expected to become a promising strategy for the detection of proteins and other biomacromolecules.     

Clinico-hematological and micronuclear changes induced by cypermethrin in broiler chicks: Their attenuation with vitamin E and selenium

This study was carried out on 90 one-day-old broiler chicks to know clinico-hematological alterations, DNA damage caused by cypermethrin (CY), and attenuation of toxic effects by vitamin E (Vit E) and selenium (Se). Birds were randomly divided into five equal groups. Groups 1–4 received CY (600 ml kg^?1 b.wt) daily for 30 days by crop tubing. In addition to CY, groups 2, 3 and 4 received Vit E (150 mg kg^?1 b.wt), Se (0.25 mg kg^?1 b.wt), and Vit E (150 mg kg^?1 b.wt)+Se (0.25 mg kg^?1 b.wt), respectively. Group 5 served as control. Birds were monitored twice daily for clinical signs. They were weighed and blood samples were collected at experimental days 10, 20 and 30 for hematological studies. CY-treated birds showed more prominent signs of toxicity compared to CY+Vit E, CY+Se and CY+Vit E+Se birds. Body weight in groups 1–3 was significantly (P<0.05) smaller at days 20 and 30 when compared with the control group. Significantly (P<0.001) higher numbers of micronuclei appeared in chicks treated with CY compared to CY+Vit E- and CY+Se-treated birds. Significantly decreased total erythrocyte counts (TEC), hemoglobin (Hb) concentration and packed cell volume (PCV) in all treated groups were recorded. Treated birds suffered from macrocytic hypochromic anemia. Leukocytosis in early stage and later leucopenia was seen in treated birds. It can be concluded that CY induces toxic effects in broilers chicks; however, these toxic effects can be ameliorated by Vit E or Se. Combination of Vit E and Se was more effective to ameliorate toxic effects of cypermethrin.     

Adhesion,migration and mechanics of human adipose-tissue-derived stem cells on silk fibroin–chitosan matrix

Silk fibroin–chitosan (SFCS) scaffold is a naturally derived biocompatible matrix with potential reconstructive surgical applications. In this study, human adipose-derived mesenchymal stem cells (ASCs) were seeded on SFCS scaffolds and cell attachment was characterized by fluorescence, confocal, time-lapse, atomic force, and scanning electron microscopy (SEM) studies. Adhesion of ASCs on SFCS was 39.4 ± 4.8% at 15 min, increasing to 92.8 ± 1.5% at 120 min. ASC adhered at regions of architectural complexity and infiltrate into three-dimensional scaffold. Time-lapse confocal studies indicated a mean ASC speed on SFCS of 18.47 ± 2.7 μm h^?1 and a mean persistence time of 41.4 ± 9.3 min over a 2.75 h study period. Cytokinetic and SEM studies demonstrated ASC–ASC interaction via microvillus extensions. The apparent elastic modulus was significantly higher (p < 0.0001) for ASCs seeded on SFCS (69.0 ± 9.0 kPa) than on glass (6.1 ± 0.4 kPa). Also, cytoskeleton F-actin fiber density was higher (p < 0.05) for ASC seeded on SFCS (0.42 ± 0.02 fibers μm^?1) than on glass-seeded controls (0.24 ± 0.03 fibers μm^?1). Hence, SFCS scaffold facilitates mesenchymal stem cell attachment, migration, three-dimensional infiltration, and cell–cell interaction. This study showed the potential use of SFCS as a local carrier for autologous stem cells for reconstructive surgery application.     

Density–property relationships in collagen–glycosaminoglycan scaffolds

Collagen–glycosaminoglycan scaffolds for the regeneration of skin have previously been fabricated by freeze-drying a slurry containing a co-precipitate of collagen and glycosaminoglycan. The mechanical properties of the scaffold are low (e.g. the dry compressive Young’s modulus is roughly 30 kPa and the dry compressive strength is roughly 5 kPa). There is interest in using these scaffolds for tendon and ligament regeneration where there is a need for improved mechanical properties. Previous attempts to increase the mechanical properties of the scaffold by increasing the solid volume fraction of the scaffolds were limited by the increasing viscosity of the slurry, making it more difficult to mix and giving inhomogeneous scaffolds. Our recent work on mineralized collagen–glycosaminoglycan scaffolds used a vacuum filtration technique to increase the volume fraction of solids in the slurry, thereby increasing the density and mechanical properties of the scaffolds. In this work, we used this technique to fabricate collagen–glycosaminoglycan scaffolds with dry densities between 0.0076 and 0.0311 g cm^?3 and pore sizes between 250 and 350 μm, values appropriate for soft tissue growth. The compressive Young’s modulus and strength in the dry state increased from 32 to 127 kPa and from 5 to 19 kPa, respectively, with increasing density. The tensile Young’s modulus in the dry state increased from 295 to 3.1 MPa with increasing density. Finally, we showed that the attachment of cells onto the scaffold was directly proportional to the specific surface area of the scaffold, which defines the total internal surface area per volume of scaffold.     

Three-dimensional quantification of susceptibility artifacts from various metals in magnetic resonance images

Susceptibility artifacts generated in magnetic resonance (MR) images were quantitatively evaluated for various metals using a three-dimensional (3-D) artifact rendering to demonstrate the correlation between magnetic susceptibility and artifact volume. Ten metals (stainless steel, Co–Cr alloy, Nb, Ti, Zr, Mo, Al, Sn, Cu and Ag) were prepared, and their magnetic susceptibilities measured using a magnetic balance. Each metal was embedded in a Ni-doped agarose gel phantom and the MR images of the metal-containing phantoms were taken using 1.5 and 3.0 T MR scanners under both fast spin echo and gradient echo conditions. 3-D renderings of the artifacts were constructed from the images and the artifact volumes were calculated for each metal. The artifact volumes of metals decreased with decreasing magnetic susceptibility, with the exception of Ag. Although Sn possesses the lowest absolute magnetic susceptibility (1.8 × 10^?6), the artifact volume from Cu (?7.8 × 10^?6) was smaller than that of Sn. This is because the magnetic susceptibility of Cu was close to that of the agarose gel phantom (?7.3 × 10^?6). Since the difference in magnetic susceptibility between the agarose and Sn is close to that between the agarose and Ag (?17.5 × 10^?6), their artifact volumes were almost the same, although they formed artifacts that were reversed in all three dimensions.     

Porous polycaprolactone scaffold for cardiac tissue engineering fabricated by selective laser sintering

An advanced manufacturing technique, selective laser sintering (SLS), was utilized to fabricate a porous polycaprolactone (PCL) scaffold designed with an automated algorithm in a parametric library system named the “computer-aided system for tissue scaffolds” (CASTS). Tensile stiffness of the sintered PCL strut was in the range of 0.43 ± 0.15 MPa when a laser power of 3 W and scanning speed of 150 in s^?1 was used. A series of compressive mechanical characterizations was performed on the parametric scaffold design and an empirical formula was presented to predict the compressive stiffness of the scaffold as a function of total porosity. In this work, the porosity of the scaffold was selected to be 85%, with micropores (40–100 μm) throughout the scaffold. The compressive stiffness of the scaffold was 345 kPa. The feasibility of using the scaffold for cardiac tissue engineering was investigated by culturing C2C12 myoblast cells in vitro for 21 days. Fluorescence images showed cells were located throughout the scaffold. High density of cells at 1.2 × 10⁶ cells ml^?1 was recorded after 4 days of culture. Fusion and differentiation of C2C12 were observed as early as 6 days in vitro and was confirmed with myosin heavy chain immunostaining after 11 days of cell culture. A steady population of cells was then maintained throughout 21 days of culturing. This work demonstrated the feasibility of tailoring the mechanical property of the scaffold for soft tissue engineering using CASTS and SLS. The macroarchitecture of the scaffold can be modified efficiently to fabricate scaffolds with different macropore sizes or changing the elemental cell design in CASTS. Further process and design optimization could be carried out in the future to fabricate scaffolds that match the tensile strength of native myocardium, which is of the order of tens of kPa.     

Discovery of low mucus adhesion surfaces

Mucus secretion from the body is ubiquitous, and finding materials that resist mucus adhesion is a major technological challenge. Here, using a high throughput platform with photo-induced graft polymerization, we first rapidly synthesized, screened and tested a library of 55 different surfaces from six functional monomer classes to discover porcine intestinal low mucus adhesion surfaces using a 1 h static mucus adsorption protocol. From this preliminary screen, two chemistries, a zwitterionic ([2-(acryloyloxy)ethyl] trimethylammonium chloride) and a multiple hydroxyl (N-[tris(hydroxymethyl)methyl]acrylamide) surface, exhibited significantly low mucus adhesion from a Langmuir-type isotherm when exposed to increasing concentrations of mucus for 24 h. Apolar or hydrophobic interactions were likely the dominant attractive forces during mucus binding since many polar or hydrophilic monomers reduced mucus adhesion. Hansen solubility parameters were used to illustrate the importance of monomer polarity and hydrogen bonding in reducing mucus adsorption. For a series of polyethylene glycol (PEG) monomers with changing molecular weight from 144 g mol^?1 to 1100 g mol^?1, we observed an excellent linear correlation (R² = 0.998) between relative amount adsorbed and the distance from a water point in a specialized Hansen solubility parameter plot, emphasizing the role of surface–water interactions for PEG modified surfaces.     

Peri-implant reactivity and osteoinductive potential of immobilized rhBMP-2 on titanium carriers

Recombinant human BMP-2 (rhBMP-2) was immobilized non-covalently and covalently as a monolayer on plasma vapour deposited (PVD) porous commercially pure titanium surfaces in amounts of 5–8 μg cm^?2, providing a ca. 10-fold increase vs. previously reported values [37]. Dissociation of the immobilized [¹²⁵I]rhBMP-2 from the surface occurred in a two-phase exponential decay: a first rapid phase (ca. 15% of immobilized BMP-2) with a half-life of 1–2 days and a second slow sustained release phase (ca. 85% of immobilized BMP-2) with a half-life of 40–60 days. Dissociation rate constants of sustained release of k_?1 = 1.3–1.9 × 10^?7 s^?1 were determined, allowing an estimation of the binding constants (K_A) for the adsorbed rhBMP-2 monolayer, to be around 10¹² M^?1. The rhBMP-2-coated surfaces showed a high level of biological activity, as demonstrated by in vitro epifluorescence tests for alkaline phosphatase with MC3T3-E1 cells and in vivo experiments. In vivo osteoinductivity of rhBMP-2-coated implants was investigated in a gap-healing model in the trabecular bone of the distal femur condylus of sheep. Healing occurred without inflammation or capsule formation. The calculated concentration of released rhBMP-2 in the 1 mm gap ranged from 20 to 98 nM – well above the half-maximal response concentration (K_0.5) for inducing alkaline phosphatase in MC3T3-E1 cells. After 4, 9 and 12 weeks the bone density (BD) and bone-to-implant contact (BIC) of the explanted implants were assessed histomorphometrically. Implants with immobilized rhBMP-2 displayed a significant (2- to 4-fold) increase in BD and BIC values vs. negative controls after 4–9 weeks. Integration of implants by trabecular bone was achieved after 4 weeks, indicating a mean “gap-filling rate” of ～250 μm week^?1. Integration of implants by cortical bone was observed after 9 weeks. Control implants without rhBMP-2 were not osseointegrated. This study demonstrates the feasibility of enhancing peri-implant osseointegration and gap bridging by immobilized rhBMP-2 on implant surfaces which may serve as a model for future clinical applications.     

Development of a shear measurement sensor for measuring forces at human–machine interfaces

Measuring shear force is crucial for investigating the pathology and treatment of pressure ulcers. In this study, we introduced a bi-axial shear transducer based on strain gauges as a new shear sensor. The sensor consisted of aluminum and polyvinyl chloride plates placed between quadrangular aluminum plates. On the middle plate, two strain gauges were placed orthogonal to one another. The shear sensor (54 mm × 54 mm × 4.1 mm), which was validated by using standard weights, displayed high accuracy and precision (measurement range, −50 to 50 N; sensitivity, 0.3 N; linear relationship, R² = 0.9625; crosstalk error, 0.635% ± 0.031%; equipment variation, 4.183). The shear force on the interface between the human body and a stand-up wheelchair was measured during sitting or standing movements, using two mats (44.8 cm × 44.8 cm per mat) that consisted of 24 shear sensors. Shear forces on the sacrum and ischium were almost five times higher (15.5 N at last posture) than those on other sites (3.5 N on average) during experiments periods. In conclusion, the proposed shear sensor may be reliable and useful for measuring the shear force on human–machine interfaces.     

The effect of screw insertion angle and thread type on the pullout strength of bone screws in normal and osteoporotic cancellous bone models

Screw fixation can be extremely difficult to achieve in osteoporotic (OP) bone because of its low strength. This study determined how pullout strength is affected by placing different bone screws at varying angles in normal and OP bone models. Pullout tests of screws placed axially, and at angles to the pullout axis (ranging from 10° to 40°), were performed in 0.09 g cm^?3, 0.16 g cm^?3 and 0.32 g cm^?3 polyurethane (PU) foam. Two different titanium alloy bone screws were used to test for any effect of thread type (i.e. cancellous or cortical) on the screw pullout strength. The cancellous screw required a significantly higher pullout force than the cortical screw (p < 0.05). For both screws, pullout strength significantly increased with increasing PU foam density (p < 0.05). For screws placed axially, and sometimes at 10°, the observed mechanism of failure was stripping of the internal screw threads generated within the PU foam by screw insertion. For screws inserted at 10°, 20°, 30° and 40°, the resistance to pullout force was observed to be by compression of the PU foam material above the angled screw; clinically, this suggests that compressed OP bone is stronger than unloaded OP bone.     

The mechanical properties of individual,electrospun fibrinogen fibers

We used a combined atomic force microscopic (AFM)/fluorescence microscopic technique to study the mechanical properties of individual, electrospun fibrinogen fibers in aqueous buffer. Fibers (average diameter 208 nm) were suspended over 12 μm-wide grooves in a striated, transparent substrate. The AFM, situated above the sample, was used to laterally stretch the fibers and to measure the applied force. The fluorescence microscope, situated below the sample, was used to visualize the stretching process. The fibers could be stretched to 2.3 times their original length before breaking; the breaking stress was 22 × 10⁶ Pa. We collected incremental stress–strain curves to determine the viscoelastic behavior of these fibers. The total stretch modulus was 17.5 × 10⁶ Pa and the relaxed elastic modulus was 7.2 × 10⁶ Pa. When held at constant strain, electrospun fibrinogen fibers showed a fast and slow stress relaxation time of 3 and 55 s.Our fibers were spun from the typically used 90% 1,1,1,3,3,3-hexafluoro-2-propanol (90-HFP) electrospinning solution and re-suspended in aqueous buffer. Circular dichroism spectra indicate that α-helical content of fibrinogen is ～70% higher in 90-HFP than in aqueous solution.These data are needed to understand the mechanical behavior of electrospun fibrinogen structures. Our technique is also applicable to study other nanoscopic fibers.     

In vitro and in vivo evaluation of ultrananocrystalline diamond as an encapsulation layer for implantable microchips

Thin ultrananocrystalline diamond (UNCD) films were evaluated for use as hermetic and bioinert encapsulating coatings for implantable microchips, where the reaction to UNCD in vitro and in vivo tissue was investigated. Leakage current tests showed that depositing UNCD coatings, which were conformally grown in (1% H₂) Ar/CH₄ plasma, on microchips rendered the surface electrochemically inactive, i.e. with a very low leakage current density (2.8 × 10⁻⁵ A cm⁻² at −1 V and 1.9 × 10⁻³ A cm⁻² at ± 5 V) ex vivo. The impact of UNCD with different surface modifications on the growth and activation of macrophages was compared to that of standard-grade polystyrene. Macrophages attached to oxygen-terminated UNCD films down-regulated their production of cytokines and chemokines. Moreover, with UNCD-coated microchips, which were implanted subcutaneously into BALB/c mice for up to 3 months, the tissue reaction and capsule formation was significantly decreased compared to the medical-grade titanium alloy Ti–6Al–4V and bare silicon. Additionally, the leakage current density, elicited by electrochemical activity, on silicon chips encapsulated in oxygen-terminated UNCD coatings remained at the low level of 2.5 × 10⁻³ A cm⁻² at 5 V for up to 3 months in vivo, which is half the level of those encapsulated in hydrogen-terminated UNCD coatings. Thus, controlling the surface properties of UNCDs makes it possible to manipulate the in vivo functionality and stability of implantable devices so as to reduce the host inflammatory response following implantation. These observations suggest that oxygen-terminated UNCDs are promising candidates for use as encapsulating coatings for implantable microelectronic devices.     

Poroelastic response of articular cartilage by nanoindentation creep tests at different characteristic lengths

Nanoindentation is an experimental technique which is attracting increasing interests for the mechanical characterization of articular cartilage. In particular, time dependent mechanical responses due to fluid flow through the porous matrix can be quantitatively investigated by nanoindentation experiments at different penetration depths and/or by using different probe sizes. The aim of this paper is to provide a framework for the quantitative interpretation of the poroelastic response of articular cartilage subjected to creep nanoindentation tests. To this purpose, multiload creep tests using spherical indenters have been carried out on saturated samples of mature bovine articular cartilage achieving two main quantitative results. First, the dependence of indentation modulus in the drained state (at equilibrium) on the tip radius: a value of 500 kPa has been found using the large tip (400 μm radius) and of 1.7 MPa using the smaller one (25 μm). Secon, the permeability at microscopic scale was estimated at values ranging from 4.5 × 10⁻¹⁶ m⁴/N s to 0.1 × 10⁻¹⁶ m⁴/N s, from low to high equivalent deformation. Consistently with a poroelastic behavior, the size-dependent response of the indenter displacement disappears when characteristic size and permeability are accounted for. For comparison purposes, the same protocol was applied to intrinsically viscoelastic homogeneous samples of polydimethylsiloxane (PDMS): both indentation modulus and time response have been found size-independent.     

Biodegradable nanocomposite hydrogel structures with enhanced mechanical properties prepared by photo-crosslinking solutions of poly(trimethylene carbonate)–poly(ethylene glycol)–poly(trimethylene carbonate) macromonomers and nanoclay particles

Soft hydrogels with elasticity modulus values lower than 100 kPa that are tough and biodegradable are of great interest in medicine and in tissue engineering applications. We have developed a series of soft hydrogel structures from different methacrylate-functionalized triblock copolymers of poly(ethylene glycol) (PEG) with poly(trimethylene carbonate) (PTMC) by photo-crosslinking aqueous solutions of the macromonomers in 2.5 and 5 wt.% colloidal dispersions of clay nanoparticles (Laponite XLG). The length of the PTMC blocks of the macromonomers and the clay content determined the physicomechanical properties of the obtained hydrogels. While an increase in the PTMC block length in the macromonomers from 0.2 to 5 kg/mol resulted in a decrease in the gel content, the addition of 5 wt.% Laponite nanoclay to the crosslinking solution lead to very high gel contents of the hydrogels of more than 95%. The effect of PTMC block length on the mechanical properties of the hydrogels was not as pronounced, and soft gels with a compressive modulus of less than 15 kPa and toughness values of 25 kJ m^?3 were obtained. However, the addition of 5 wt.% Laponite nanoclay to the formulations considerably increased the compressive modulus and resilience of the hydrogels; swollen nanocomposite networks with compressive modulus and toughness values of up to 67 kPa and 200 kJ m^?3, respectively, could then be obtained. The prepared hydrogels were shown to be enzymatically degradable by cholesterol esterase and by the action of macrophages. With an increase in PTMC block length in the hydrogels, the rates of mass loss increased, while the incorporated Laponite nanoclay suppressed degradation. Nanocomposite hydrogel structures with a designed gyroid pore network architecture were prepared by stereolithography. Furthermore, in the swollen state the porous gyroid structures were mechanically stable and the pore network remained fully open and interconnected.     

Anterior tibiofemoral intersegmental forces during landing are predicted by passive restraint measures in women

BackgroundPassive restraint capabilities may influence sagittal plane knee joint mechanics during activity. This study aimed to determine if measures associated with passive restraint of anterior translation of the tibia are predictive of peak anterior knee shear force during landing.MethodsPassive restraint measures were assessed via joint arthrometry and during 40% body weight simulated weight acceptance using recreationally active students (73 F, 42 M; 21.8 ± 2.9 yr, 1.69 ± 0.1 m, 68.9 ± 14.1 kg). Anterior knee laxity (mm) at 133 N and initial (0–20 N) and terminal (100–130 N) anterior stiffnesses (N/mm) were calculated from arthrometer data. Peak anterior tibial acceleration (m?s^?2) relative to the femur was assessed via electromagnetic position sensors during 40% body weight acceptance trials. Peak knee shear force was assessed during double-leg drop jumps.ResultsSex specific linear stepwise regressions revealed that in females, increasing peak tibial acceleration (5.1 ± 1.8 m·s^? 2) (R²? = 7.3%, P? = 0.021), increasing initial anterior stiffness (31.0 ± 14.0 N/mm) (R²? = 5.9%, P? = 0.032), and decreasing terminal anterior stiffness (43.4 ± 17.4 N/mm) (R²? = 4.9%, P? = 0.046) collectively predicted greater peak knee shear forces (66.6 ± 12.03% BW) (multiple R² = 18.1%). No male regressions were significant.ConclusionsSagittal laxity measures are associated with anterior knee shear loads during landing in females. Greater tibial acceleration during early axial load along with greater initial and lesser terminal anterior stiffnesses predicted increasing anterior knee shear forces. Future work should investigate the combined contribution of passive and active restraints to high-risk ACL biomechanics.     

Differential localization and bacteriostasis of Vibrio campbellii among tissues of the Eastern oyster,Crassostrea virginica

In bivalve mollusks the roles of individual tissues in antimicrobial defense remain unclear. In this study, Crassostrea virginica were injected in the adductor muscle with 10⁵ live Vibrio campbellii. Major tissues were dissected at 10, 30, 60 or 120 min postinjection (PI); in each tissue undegraded (intact) bacteria were quantified by real-time PCR and culturable bacteria were enumerated by selective plating. At 10 min PI, accumulation of bacteria varied among tissues from approximately 2.4 × 10³ (labial palps, digestive gland) to 24.2 × 10³ (gonads) intact Vibrio g^?1. Neither distribution nor accumulation of intact bacteria changed with time except in the hemolymph. In most tissues, more than 80% of intact bacteria were culturable at 10 min PI and culturability decreased with time. In contrast, only 19% of intact bacteria in gonadal tissue could be cultured at 10 min PI, pointing to a major role for the gonadal tissues in antibacterial defense of molluscs.     

In vitro cytotoxicity of porous silicon microparticles: Effect of the particle concentration,surface chemistry and size

We report here the in vitro cytotoxicity of mesoporous silicon (PSi) microparticles on the Caco-2 cells as a function of particle size fractions (1.2–75 μm), particle concentration (0.2–4 mg ml^?1) and incubation times (3, 11 and 24 h). The particle size (smaller PSi particles showed higher cytotoxicity) and the surface chemistry treatment of the PSi microparticles were considered to be the key factors regarding the toxicity aspects. These effects were significant after the 11 and 24 h exposure times, and were explained by cell–particle interactions involving mitochondrial disruption resulting from ATP depletion and reactive oxygen species production induced by the PSi surface. These events further induced an increase in cell apoptosis and consequent cell damage and cell death in a dose-dependent manner and as a function of the PSi particle size. These effects were, however, less pronounced with thermally oxidized PSi particles. Under the experimental conditions tested and at particle sizes >25 μm, the non-toxic threshold concentration for thermally hydrocarbonized and carbonized PSi particles was <2 mg ml^?1, and for thermally oxidized PSi microparticles was <4 mg ml^?1.     

Elastic modulus and collagen organization of the rabbit cornea: Epithelium to endothelium

The rabbit is commonly used to evaluate new corneal prosthetics and study corneal wound healing. Knowledge of the stiffness of the rabbit cornea would better inform the design and fabrication of keratoprosthetics and substrates with relevant mechanical properties for in vitro investigations of corneal cellular behavior. This study determined the elastic modulus of the rabbit corneal epithelium, anterior basement membrane (ABM), anterior and posterior stroma, Descemet’s membrane (DM) and endothelium using atomic force microscopy (AFM). In addition, three-dimensional collagen fiber organization of the rabbit cornea was determined using nonlinear optical high-resolution macroscopy. The elastic modulus as determined by AFM for each corneal layer was: epithelium, 0.57 ± 0.29 kPa (mean ± SD); ABM, 4.5 ± 1.2 kPa, anterior stroma, 1.1 ± 0.6 kPa; posterior stroma, 0.38 ± 0.22 kPa; DM, 11.7 ± 7.4 kPa; and endothelium, 4.1 ± 1.7 kPa. The biophysical properties, including the elastic modulus, are unique for each layer of the rabbit cornea and are dramatically softer in comparison to the corresponding regions of the human cornea. Collagen fiber organization is also dramatically different between the two species, with markedly less intertwining observed in the rabbit vs. human cornea. Given that the substratum stiffness considerably alters the corneal cell behavior, keratoprosthetics that incorporate mechanical properties simulating the native human cornea may not elicit optimal cellular performance in rabbit corneas that have dramatically different elastic moduli. These data should allow for the design of substrates that better mimic the biomechanical properties of the corneal cellular environment.