Susceptibility of Legionella gormanii Membrane-Derived Phospholipids to the Peptide Action of Antimicrobial LL-37-Langmuir Monolayer Studies.

LL-37 is the only member of the cathelicidin-type host defense peptide family in humans. It exhibits broad-spectrum bactericidal activity, which represents a distinctive advantage for future therapeutic targets. The presence of choline in the growth medium for bacteria changes the composition and physicochemical properties of their membranes, which affects LL-37's activity as an antimicrobial agent. In this study, the effect of the LL-37 peptide on the phospholipid monolayers at the liquid-air interface imitating the membranes of Legionella gormanii bacteria was determined. The Langmuir monolayer technique was employed to prepare model membranes composed of individual classes of phospholipids-phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), cardiolipin (CL)-isolated from L. gormanii bacteria supplemented or non-supplemented with exogenous choline. Compression isotherms were obtained for the monolayers with or without the addition of the peptide to the subphase. Then, penetration tests were carried out for the phospholipid monolayers compressed to a surface pressure of 30 mN/m, followed by the insertion of the peptide into the subphase. Changes in the mean molecular area were observed over time. Our findings demonstrate the diversified effect of LL-37 on the phospholipid monolayers, depending on the bacteria growth conditions. The substantial changes in membrane properties due to its interactions with LL-37 enable us to propose a feasible mechanism of peptide action at a molecular level. This can be associated with the stable incorporation of the peptide inside the monolayer or with the disruption of the membrane leading to the removal (desorption) of molecules into the subphase. Understanding the role of antimicrobial peptides is crucial for the design and development of new strategies and routes for combating resistance to conventional antibiotics.

Lipopolysaccharide of Legionella pneumophila Serogroup 1 Facilitates Interaction with Host Cells.

Legionella pneumophila is the primary causative agent of Legionnaires' disease. The mutant-type strain interrupted in the ORF7 gene region responsible for the lipopolysaccharide biosynthesis of the L. pneumophila strain Heysham-1, lacking the O-acetyl groups attached to the rhamnose of the core part, showed a higher surface polarity compared with the wild-type strain. The measurement of excitation energy transfer between fluorophores located on the surface of bacteria and eukaryotic cells showed that, at an early stage of interaction with host cells, the mutant exhibited weaker interactions with Acanthamoeba castellanii cells and THP-1-derived macrophages. The mutant displayed reduced adherence to macrophages but enhanced adherence to A. castellanii, suggesting that the O-acetyl group of the LPS core region plays a crucial role in facilitating interaction with macrophages. The lack of core rhamnose O-acetyl groups made it easier for the bacteria to multiply in amoebae and macrophages. The mutant induced TNF-α production more strongly compared with the wild-type strain. The mutant synthesized twice as many ceramides Cer(t34:0) and Cer(t38:0) than the wild-type strain. The study showed that the internal sugars of the LPS core region of L. pneumophila sg 1 can interact with eukaryotic cell surface receptors and mediate in contacting and attaching bacteria to host cells as well as modulating the immune response to infection.

Insight into the Mechanism of Interactions between the LL-37 Peptide and Model Membranes of Legionella gormanii Bacteria.

Legionella gormanii is a fastidious, Gram-negative bacterium known to be the etiological agent of atypical community-acquired pneumonia. The human cathelicidin LL-37 exhibits a dose-dependent bactericidal effect on L. gormanii. The LL-37 peptide at the concentration of 10 µM causes the bacteria to become viable but not cultured. The antibacterial activity of the peptide is attributed to its effective binding to the bacterial membrane, as demonstrated by the fluorescence lifetime imaging microscopy. In this study, to mimic the L. gormanii membranes and their response to the antimicrobial peptide, Langmuir monolayers were used with the addition of the LL-37 peptide to the subphase of the Langmuir trough to represent the extracellular fluid. The properties of the model membranes (Langmuir monolayers) formed by phospholipids (PL) isolated from the L. gormanii bacteria cultured on the non-supplemented (PL-choline) and choline-supplemented (PL+choline) medium were determined, along with the effect of the LL-37 peptide on the intermolecular interactions, packing, and ordering under the monolayer compression. Penetration tests at the constant surface pressure were carried out to investigate the mechanism of the LL-37 peptide action on the model membranes. The peptide binds to the anionic bacterial membranes preferentially, due to its positive charge. Upon binding, the LL-37 peptide can penetrate into the hydrophobic tails of phospholipids, destabilizing membrane integrity. The above process can entail membrane disruption and ultimately cell death. The ability to evoke such a great membrane destabilization is dependent on the share of electrostatic, hydrogen bonding and Lifshitz-van der Waals LL-37-PL interactions. Thus, the LL-37 peptide action depends on the changes in the lipid membrane composition caused by the utilization of exogenous choline by the L. gormanii.

An Insight into the Defects-Driven Plasticity in Ductile Cast Irons.

The microstructure and tensile behavior of two heavy section castings that had chemical compositions typical of GJS400 were investigated. Conventional metallography, fractography, and micro-Computer Tomography (µ-CT) were employed, enabling the quantification of the volume fractions of eutectic cells with degenerated Chunky Graphite (CHG), which was identified as the major defect in the castings. The Voce equation approach was exploited to evaluate the tensile behaviors of the defective castings for integrity assessment. The results demonstrated that the Defects-Driven Plasticity (DDP) phenomenon, which refers to an unexpected regular plastic behavior related to defects and metallurgical discontinuities, was consistent with the observed tensile behavior. This resulted in a linearity of Voce parameters in the Matrix Assessment Diagram (MAD), which contradicts the physical meaning of the Voce equation. The findings suggest that the defects, such as CHG, contribute to the linear distribution of Voce parameters in the MAD. Furthermore, it is reported that the linearity in the MAD of Voce parameters for a defective casting is equivalent to the existence of a pivotal point in the differential data of the tensile strain hardening data. This pivotal point was exploited to propose a new material quality index assessing the integrity of castings.

Physicochemical Characteristics of Model Membranes Composed of Legionella gormanii Lipids.

Legionella gormanii is one of the species belonging to the genus Legionella, which causes atypical community-acquired pneumonia. The most important virulence factors that enable the bacteria to colonize the host organism are associated with the cell surface. Lipids building the cell envelope are crucial not only for the membrane integrity of L. gormanii but also by virtue of being a dynamic site of interactions between the pathogen and the metabolites supplied by its host. The utilization of exogenous choline by the Legionella species results in changes in the lipids' composition, which influences the physicochemical properties of the cell surface. The aim of this study was to characterize the interfacial properties of the phospholipids extracted from L. gormanii cultured with (PL+choline) and without exogenous choline (PL-choline). The Langmuir monolayer technique coupled with the surface potential (SPOT) sensor and the Brewster angle microscope (BAM) made it possible to prepare the lipid monomolecular films (model membranes) and study their properties at the liquid/air interface at 20 °C and 37 °C. The results indicate the effect of the choline addition to the bacterial medium on the properties of the L. gormanii phospholipid membranes. The differences were revealed in the organization of monolayers, their molecular packing and ordering, degree of condensation and changes in the components' miscibility. These findings are the basis for further research on the mechanisms of adaptation of this pathogen, which by changing the native composition and properties of lipids, bypasses the action of antimicrobial compounds and avoids the host immune attack.

The Influence of Chemical Activity Models on the Description of Ion Transport through Micro-Structured Cementitious Materials.

The significance of ion activity in transport through a porous concrete material sample with steel rebar in its center and bathing solution is presented. For the first time, different conventions and models of ion activity are compared in their significance and influence on the ion fluxes. The study closes an interpretational gap between ion activity in a stand-alone (stagnant) electrolyte solution and ion transport (dynamic) through concrete pores. Ionic activity models developed in stationary systems, namely, the Debye-Hückel (DH), extended DH, Davies, Truesdell-Jones, and Pitzer models, were used for modeling the transport of ions driven through the activity gradient. The activities of ions are incorporated into a frame of the Nernst-Planck-Poisson (NPP) equations. Calculations were done with COMSOL software for a real concrete microstructure determined by X-ray computed tomography. The concentration profiles of four ions (Na+, Cl-, K+, OH-), the ionic strength, and the electric potential in mortar (with pores) and concrete samples (with aggregates and pores) are presented and compared. The Pitzer equation gave the most reliable results for all systems studied. The difference between the concentration profiles calculated with this equation and with the assumption of the ideality of the solution is negligible while the potential profiles are clearly distinguishable.

The impact of the parameters of the constitutive model on the distribution of strain in the femoral head.

The rapid spread of the finite element method has caused that it has become, among other methods, the standard tool for pre-clinical estimates of bone properties. This paper presents an application of this method for the calculation and prediction of strain and stress fields in the femoral head. The aim of the work is to study the influence of the considered anisotropy and heterogeneity of the modeled bone on the mechanical fields during a typical gait cycle. Three material models were tested with different properties of porous bone carried out in literature: a homogeneous isotropic model, a heterogeneous isotropic model, and a heterogeneous anisotropic model. In three cases studied, the elastic properties of the bone were determined basing on the Zysset-Curnier approach. The tensor of elastic constants defining the local properties of porous bone is correlated with a local porosity and a second order fabric tensor describing the bone microstructure. In the calculations, a model of the femoral head generated from high-resolution tomographic scans was used. Experimental data were drawn from publicly available database "Osteoporotic Virtual Physiological Human Project." To realistically reflect the load on the femoral head, main muscles were considered, and their contraction forces were determined based on inverse kinematics. For this purpose, the results from OpenSim packet were used. The simulations demonstrated that differences between the results predicted by these material models are significant. Only the anisotropic model allowed for the plausible distribution of stresses along the main trabecular groups. The outcomes also showed that the precise evaluation of the mechanical fields is critical in the context of bone tissue remodeling under mechanical stimulations.

Simple anisotropic model of Bone Adaptation - SAMBA.

Bone presents the ability to adapt itself to the evolving mechanical environment. A simple anisotropic model for bone adaptation allowing reproducing the evolution of the elastic properties and the reorientation of the anisotropy frame is proposed is this work. The elastic properties are related to the value of the bone apparent density. The evolution law of the density is described via two functions reflecting the activities of the osteoclast and osteoblast cells. The anisotropy of the elastic properties of the bone is assumed evolving continuously between those of trabecular and compact tissues. The existence of a target material frame is assumed to describe its reorientation. The rate of rotation or spin of the material frame is supposed to be proportional to the target orientation angle and to the activity of the osteoclasts and osteoblasts. The mechanical stimulus governing the evolution of the apparent density is defined as the ratio between the current strain energy density and its critical value corresponding to the initiation of bone damage. The simulations showed that this simple model can reproduce some essential phenomena observed during bone adaptation process.

Investigation of Molten Metal Infiltration into Micropore Carbon Refractory Materials Using X-ray Computed Tomography.

The lifetime of a blast furnace (BF), and, consequently, the price of steel, strongly depends on the degradation of micropore carbon refractory materials used as lining materials in the BF hearth. One of the major degradation mechanisms in the BF hearth is related to the infiltration and dissolution of refractory materials in molten metal. To design new and more resilient materials, we need to know more about degradation mechanisms, which can be achieved using laboratory tests. In this work, we present a new investigation method of refractory materials infiltration resistance. The designed method combines a standard degradation test (hot metal penetration test) with X-ray computed tomography (XCT) measurements. Application of XCT measurements before and after molten metal infiltration allows observing changes in the micropore carbon refractory material's microstructure and identifying the elements of the open pore structure that are crucial in molten metal infiltration.

Decoding Rocks: An Assessment of Geomaterial Microstructure Using X-ray Microtomography, Image Analysis and Multivariate Statistics.

An understanding of the microstructure of geomaterials such as rocks is fundamental in the evaluation of their functional properties, as well as the decryption of their geological history. We present a semi-automated statistical protocol for a complex 3D characterization of the microstructure of granular materials, including the clustering of grains and a description of their chemical composition, size, shape, and spatial properties with 44 unique parameters. The approach consists of an X-ray microtomographic image processing procedure, followed by measurements using image analysis and statistical multivariate analysis of its results utilizing freeware and widely available software. The statistical approach proposed was tested out on a sandstone sample with hidden and localized deformational microstructures. The grains were clustered into distinctive groups covering different compositional and geometrical features of the sample's granular framework. The grains are pervasively and evenly distributed within the analysed sample. The spatial arrangement of grains in particular clusters is well organized and shows a directional trend referring to both microstructures. The methodological approach can be applied to any other rock type and enables the tracking of microstructural trends in grains arrangement.

How to Obtain the Maximum Properties Flexibility of 3D Printed Ketoprofen Tablets Using Only One Drug-Loaded Filament?

The flexibility of dose and dosage forms makes 3D printing a very interesting tool for personalized medicine, with fused deposition modeling being the most promising and intensively developed method. In our research, we analyzed how various types of disintegrants and drug loading in poly(vinyl alcohol)-based filaments affect their mechanical properties and printability. We also assessed the effect of drug dosage and tablet spatial structure on the dissolution profiles. Given that the development of a method that allows the production of dosage forms with different properties from a single drug-loaded filament is desirable, we developed a method of printing ketoprofen tablets with different dose and dissolution profiles from a single feedstock filament. We optimized the filament preparation by hot-melt extrusion and characterized them. Then, we printed single, bi-, and tri-layer tablets varying with dose, infill density, internal structure, and composition. We analyzed the reproducibility of a spatial structure, phase, and degree of molecular order of ketoprofen in the tablets, and the dissolution profiles. We have printed tablets with immediate- and sustained-release characteristics using one drug-loaded filament, which demonstrates that a single filament can serve as a versatile source for the manufacturing of tablets exhibiting various release characteristics.

A 3D model of the renal vasculature - a joined result of the corrosion casting technique, micro-CT imaging and rapid prototyping technology.

Three-dimensional (3D) printed model of the renal vasculature shows a high level of accuracy of subsequent divisions of both the arterial and the venous tree. However, minor artifacts appeared in the form of oval endings to the terminal branches of the vascular tree, contrary to the anticipated sharply pointed segments. Unfortunately, selective laser sintering process does not currently permit to present the arterial, venous and urinary systems in distinct colors, hence topographic relationship between the vascular and the pelvicalyceal systems is difficult to attain. Nonetheless, the 3D printed model can be used for educational purposes to demonstrate the vast renal vasculature and may also serve as a reference model whilst evaluating morphological anomalies of the intrarenal vasculature in a surgical setting.

Characterization of mechanical behaviour of healthy and injured human incus by eigenfrequency evaluation.

The usage of finite element method techniques gives a possibility to replace time-consuming experiments or imitate physical process in the ear by numerical simulation. Especially, the research of spatial motion of ossicular chain in the middle ear is of high interest for the oto-surgeons and engineers. It is known that the most affected bone from the ossicular chain is the incus. After the cholesteatoma operation and tympanoplasty, the affected incus is removed or sacrificed; thus, the possibility of transducing noise lays on the stapes, new titanium or other material prosthesis. In this case, the affected incus was removed because of the cholesteatoma that was lying in front of it in the tympanic cavity. The removed incus with the affected long process passed micro-computed tomography. The computer-aided design systems allowed redesigning a 'healthy' incus with an intact long process. In this way, it was possible to evaluate the influence of damaged long process of incus in the vibrational analysis. This article analyses the problems of mechanical behaviour of injured and healthy human incus. The numerical simulation has demonstrated that the features of healthy incus and analysed injured incus do not differ significantly, especially at low (about 500 Hz) frequencies. It explains why there is no impact of cholesteatoma on hearing for a long time in the audiogram.

Micro-CT analysis of the internal acoustic meatus angles as a method of sex estimation in skeletal remains.

The aim of the present study was to verify the lateral angle method for sex estimation by using computed micro-tomography. Two measurements of the anterior lateral angle of the internal acoustic meatus were assessed. The performed analysis revealed that the mean angle at the level of transverse crest of the fundus of the internal acoustic meatus in adults is significantly greater in females (45.58° vs. 39.68°; p < 0.05). A 45° sectioning point was applied, and sex allocation using this measurement was correct in 81.81% of adult samples. In turn, when utilizing the measurement at the level of modiolus of the cochlea, we did not obtain a satisfactory result in sex classification (59.09%). In subadults (male and female samples), the anterior lateral angles were undifferentiated. Thus, the conducted micro-CT analysis shows that the anterior lateral angle measurement method can be used as a preliminary indication of sex in adult individuals.

Carbon nanotube/iron oxide hybrid particles and their PCL-based 3D composites for potential bone regeneration.

This study describes the preparation, and evaluates the biocompatibility, of hydroxylated multi-walled carbon nanotubes (fCNTs) functionalized with magnetic iron oxide nanoparticles (IONs) creating hybrid nanoparticles. These nanoparticles were used for preparing a composite porous poly(ε-caprolactone) scaffolds for potential utilization in regenerative medicine. Hybrid fCNT/ION nanoparticles were prepared in two mass ratios - 1:1 (H1) and 1:4 (H4). PCL scaffolds were prepared with various concentrations of the nanoparticles with fixed mass either of the whole nanoparticle hybrid or only of the fCNTs. The hybrid particles were evaluated in terms of morphology, composition and magnetic properties. The cytotoxicity of the hybrid nanoparticles and the pure fCNTs was assessed by exposing the SAOS-2 human cell line to colloids with a concentration range from 0.01 to 1 mg/ml. The results indicate a gradual increase in the cytotoxicity effect with increasing concentration. At low concentrations, interestingly, SAOS-2 metabolic activity was stimulated by the presence of IONs. The PCL scaffolds were characterized in terms of the scaffold architecture, the dispersion of the nanoparticles within the polymer matrix, and subsequently in terms of their thermal, mechanical and magnetic properties. A higher ION content was associated with the presence of larger agglomerates of particles. With exception of the scaffold with the highest content of the H4 nanoparticle hybrid, all composites were superparamagnetic. In vitro tests indicate that both components of the hybrid nanoparticles may have a positive impact on the behavior of SAOS-2 cells cultivated on the PCL composite scaffolds. The presence of fCNTs up to 1 wt% improved the cell attachment to the scaffolds, and a content of IONs below 1 wt% increased the cell metabolic activity.

Biocompatible Nanocomposite Implant with Silver Nanoparticles for Otology-In Vivo Evaluation.

The aim of this work was to investigate of biocompatibility of polymeric implants modified with silver nanoparticles (AgNPs). Middle ear prostheses (otoimplants) made of the (poly)acrylonitrile butadiene styrene (ABS) and ABS modified with silver nanoparticles were prepared through extrusion and injection moulding process. The obtained prostheses were characterized by SEM-EDX, micro-CT and mechanical tests, confirming their proper shape, good AgNPs homogenization and mechanical parameters stability. The biocompatibility of the implants was evaluated in vivo on rats, after 4, 12, 24 and 48 weeks of implantation. The tissue-healing process and cytotoxicity of the implants were evaluated on the basis of microscopic observations of the materials morphology after histochemical staining with cytochrome c oxidase (OCC) and acid phosphatase (AP), as well as via micro-tomography (ex vivo). The in vivo studies confirmed biocompatibility of the implants in the surrounding tissue environment. Both the pure ABS and nanosilver-modified ABS implants exhibited a distinct decrease in the area of granulation tissue which was replaced with the regenerating muscle tissue. Moreover, a slightly smaller area of granulation tissue was observed in the surroundings of the silver-doped prosthesis than in the case of pure ABS prosthesis. The kinetics of silver ions releasing from implants was investigated by ICP-MS spectrometry. The measurement confirmed that concentration of the silver ions increased within the implant's immersion period. Our results showed that middle ear implant with the nanoscale modification is biocompatible and might be used in ossicular reconstruction.

The impact of polymers on 3D microstructure and controlled release of sildenafil citrate from hydrophilic matrices.

Sildenafil citrate has short biological half-life in humans. Thus, matrix tablets of controlled release were designed and prepared by compaction on the basis of hydrophilic polymers, i.e. HPMC, sodium alginate, carbomer, poloxamer and their mixtures. The impact of these polymers on sildenafil release in vitro and its pharmacokinetics in vivo was evaluated. Since drug release rate from hydrophilic matrices can be govern by the porosity of the matrix, the microstructure of tablets was studied using X-ray microcomputed tomography. 3D network of either open (percolating) or closed (non-percolating) pores was reconstructed. The tortuosity and the diameter of both kinds of pores were determined. Their spatial distribution within the matrix was analyzed in linear and radial direction. Polymer-dependent characteristics of the open pores (Ø > 2 µm) architecture was shown. The release profiles of sildenafil from matrix tablets fitted to Korsmeyer-Peppas model (r2: 0.9331-0.9993) with either Fickian diffusion or anomalous transport involved. Mean dissolution time (MDT) from tablets made of HPMC, carbomer or a mixture of HPMC and sodium alginate (2:1) was ca. 100 min, which was more than twelve times longer as compared to matrices prepared of silicified microcrystalline cellulose (MDT = 8 min). MDT correlated with the number of the open pores (Pearson's r = 0.94). Sustained release of sildenafil from ground carbomer tablets reflected in the slow absorption of the drug (tmax = 5.0 ±â€¯1.2 h) in vivo and the relative bioavailability of 151%. Interestingly, the relative bioavailability of sildenafil from binary matrices composed of HPMC and sodium alginate (2:1) was almost four times higher than that of sildenafil alone.

Oval Window Size and Shape: a Micro-CT Anatomical Study With Considerations for Stapes Surgery.

BACKGROUND: The oval window is an important structure with regard to stapes surgeries, including stapedotomy for the treatment of otosclerosis. Recent study of perioperative imaging of the oval window has revealed that oval window niche height can indicate both operative difficulty and subjective discomfort during otosclerosis surgery. With regard to shape, structures incorporated into the oval window niche, such as cartilage grafts, must be compatible with the shape of the oval window. Despite the clinical importance of the oval window, there is little information regarding its size and shape. METHODS: This study assessed oval window size and shape via micro-computed tomography paired with modern morphometric methodology in the fetal, infant, child, and adult populations. Additionally, the study compared oval window size and shape between sexes and between left- and right-sided ears. RESULTS: No significant differences were found among traditional morphometric parameters among age groups, sides, or sexes. However, geometric morphometric methods revealed shape differences between age groups. Further, geometric morphometric methods provided the average oval window shape and most-likely shape variance. CONCLUSION: Beyond demonstrating oval window size and shape variation, the results of this report will aid in identifying patients among whom anatomical variation may contribute to surgical difficulty and surgeon discomfort, or otherwise warrant preoperative adaptations for the incorporation of materials into and around the oval window.

Internal and external morphology of mandibular molars: An original micro-CT study and meta-analysis with review of implications for endodontic therapy.

The aim of this radiological micro-CT study and meta-analysis was to determine the morphological features of the root canal anatomy of the mandibular molars. The radiological study included micro-CT scans of 108 mandibular first, 120 mandibular second, and 146 mandibular third molars. For our meta-analysis, an extensive search was conducted through PubMed, Embase, and Web of Science to identify articles eligible for inclusion. Data extracted included investigative method (cadaveric, intraoperative, or imaging), Vertucci type of canal configuration, presence/number of canals, roots, apical foramina, apical deltas, and intercanal communications. In the mesial roots of mandibular molars, the most frequent Vertucci type of canal configuration was type IV, except for the mandibular third molar where type I was most common. Type I was most common in the distal root. There were usually two canals in the mesial root and one in the distal root. Two was the most common number of roots, and a third root was most prevalent in Asia. One apical foramen was most common in the distal root and two apical foramina in the mesial root. Intercanal communications were most frequent in the mesial root. Knowledge of the complex anatomy of the mandibular molars can make root canal therapy more likely to succeed. We recommend the use of cone-beam computed tomography before and after endodontic treatment to enable the root anatomy to be accurately described and properly diagnosed, and treatment outcome to be assessed. Clin. Anat. 31:797-811, 2018. © 2018 Wiley Periodicals, Inc.

Corrigendum to "Metformin Decreases Reactive Oxygen Species, Enhances Osteogenic Properties of Adipose-Derived Multipotent Mesenchymal Stem Cells In Vitro, and Increases Bone Density In Vivo".

[This corrects the article DOI: 10.1155/2016/9785890.].