Adhesion and Activation of Blood Platelets on Laser-Structured Surfaces of Biomedical Metal Alloys.

The laser surface modification of metallic implants presents a promising alternative to other surface modification techniques. A total of four alloyed metallic biomaterials were used for this study: medical steel (AISI 316L), cobalt-chromium-molybdenum alloy (CoCrMo) and titanium alloys (Ti6Al4V and Ti6Al7Nb). Samples of metallic biomaterials after machining were subjected to polishing or laser modification in two different versions. The results of surface modification were documented using SEM imaging and roughness measurement. After modification, the samples were sterilized with dry hot air, then exposed to citrate blood, washed with PBS buffer, fixed with glutaraldehyde, sputtered with a layer of gold and imaged using SEM to enable the quantification of adhered, activated and aggregated platelets on the surface of biomaterial samples. The average total number, counted in the field of view, of adhered platelets on the surfaces of the four tested biomaterials, regardless of the type of modification, did not differ statistically significantly (66 ± 81, 67 ± 75, 61 ± 70 and 57 ± 61 for AISI 316L, CoCrMo, Ti6Al4V and Ti6Al7Nb, respectively) and the average number of platelet aggregates was statistically significantly higher (p < 0.01) on the surfaces of AISI 316L medical steel (42 ± 53) and of the CoCrMo alloy (42 ± 52) compared to the surfaces of the titanium alloys Ti6Al4V (33 ± 39) and Ti6Al7Nb (32 ± 37). Remaining blood after contact was used to assess spontaneous platelet activation and aggregation in whole blood by flow cytometry. An in-depth analysis conducted on the obtained results as a function of the type of modification indicates small but statistically significant differences in the interaction of platelets with the tested surfaces of metallic biomaterials.

Assessment of the influence of gold nanoparticles stabilized with PAMAM dendrimers on HUVEC barrier cells.

Civilization diseases, cancer, frequent mutations of viruses and other pathogens constitute the need to look for new drugs, as well as systems for their targeted delivery. One of the promising way of using drugs is supplying them by linking to nanostructures. One of the solution for the development of nanobiomedicine are metallic nanoparticles stabilized with various polymer structures. In this report, we present the synthesis of gold nanoparticles, their stabilization with polyamidoamine (PAMAM) dendrimers with ethylenediamine core and the characteristics of the obtained product (AuNPs/PAMAM). The presence, size and morphology of synthesized gold nanoparticles were evaluated by ultraviolet-visible light spectroscopy, transmission electron microscopy and atomic force microscopy. The hydrodynamic radius distribution of the colloids was analyzed by dynamic light scattering technique. Additionally, the cytotoxicity and changes in mechanical properties of human umbilical vein endothelial cell line (HUVEC) cells caused by AuNPs/PAMAM were assessed. The results of studies on the nanomechanical properties of cells suggest a two-step changes in cell elasticity as a response to contact with nanoparticles. When using AuNPs/PAMAM in lower concentrations, no changes in cell viability were observed and the cells were softer than untreated cells. When higher concentrations were used, a decrease in the cells viability to about 80 % were observed, as well as non-physiological stiffening of the cells. The presented results may play a significant role in the development of nanomedicine.

Investigation of HUVEC response to exposure to PAMAM dendrimers - changes in cell elasticity and vesicles release.

The aim of this study is to assess the effect of PAMAM dendrimers of second, fourth, and seventh generations on human umbilical vein endothelial cells. Primary endothelial cells were exposed to PAMAM dendrimers for 24 h, using concentrations reducing cellular viability to the levels of 90, 75, and 50%. We assumed, that changes in mechanical properties reflect toxicity of PAMAM dendrimers. The mechanical properties were investigated using atomic force spectroscopy (AFS) technique with the use of two approaches for measuring cell elasticity: global, where the tests were performed using a micrometer-hemispherical probe, and local, where a nanometer-sized probe was used. For the sharp probe, a reduction in the elasticity modulus was observed in comparison to untreated control cells, that is related to the depolymerization of the cytoskeleton and the processes leading to cell apoptosis. In the case of the hemispherical probe, cell softening was also observed in comparison to control cells, but with increasing PAMAM concentrations, the modulus of elasticity increases. It is related to the sensing of numerous intracellular vesicles with the use of this probe, e.g. endosomal and empty plasmalemmal which can also alter cell elasticity. The presence of external and intracellular vesicles was confirmed by scanning and transmission electron microscopy. The relationship between the elasticity of HUVEC cells exposed to PAMAM dendrimers of selected generations and their toxic effects was presented herein for the first time. In the transmission electron microscopy images of the cells exposed to PAMAM dendrimers, we have also observed distinctive vesicles with regular multilayer arranged structure.

Osteosarcoma cells in early and late stages as cancer in vitro progression model for assessing the responsiveness of cells to silver nanoparticles.

Understanding of biology of osteosarcoma malignant progression is indispensable for enhancement of conventional chemotherapy by the use of silver nanoparticles (AgNPs). We presented an in vitro model of cancer progression closely resembling processes occurring in vivo in terms of protein profile. A comparison of cytotoxic and genotoxic potential of AgNPs in Saos-2 cells in early stages of cancerous progression (early passages) with the cells in advanced stages (late passages) demonstrated significantly reduced responsiveness of the late passage cells to nanoparticles toxicity. It was also confirmed by proteome analysis as we identified considerably higher number of differentially expressed proteins in Saos-2 cells in early passages compared to the late passage cells. Our studies showed that the ability of AgNPs as potential drug carriers to deliver a medication and/or to evoke toxic effects might be significantly diminished in advanced stages of cancer progression.

Isolation, Biochemical Characterisation and Identification of Thermotolerant and Cellulolytic Paenibacillus lactis and Bacillus licheniformis.

RESEARCH BACKGROUND: Cellulose is an ingredient of waste materials that can be converted to other valuable substances. This is possible provided that the polymer molecule is degraded to smaller particles and used as a carbon source by microorganisms. Because of the frequently applied methods of pretreatment of lignocellulosic materials, the cellulases derived from thermophilic microorganisms are particularly desirable. EXPERIMENTAL APPROACH: We were looking for cellulolytic microorganisms able to grow at 50 °C and we described their morphological features and biochemical characteristics based on carboxymethyl cellulase (CMCase) activity and the API® ZYM system. The growth curves during incubation at 50 °C were examined using the BioLector® microbioreactor. RESULTS AND CONCLUSIONS: Forty bacterial strains were isolated from fermenting hay, geothermal karst spring, hot spring and geothermal pond at 50 °C. The vast majority of the bacteria were Gram-positive and rod-shaped with the maximum growth temperature of at least 50 °C. We also demonstrated a large diversity of biochemical characteristics among the microorganisms. The CMCase activity was confirmed in 27 strains. Hydrolysis capacities were significant in bacterial strains: BBLN1, BSO6, BSO10, BSO13 and BSO14, and reached 2.74, 1.62, 1.30, 1.38 and 8.02 respectively. Rapid and stable growth was observed, among others, for BBLN1, BSO10, BSO13 and BSO14. The strains fulfilled the selection conditions and were identified based on the 16S rDNA sequences. BBLN1, BSO10, BSO13 were classified as Bacillus licheniformis, whereas BSO14 as Paenibacillus lactis. NOVELTY AND SCIENTIFIC CONTRIBUTION: We described cellulolytic activity and biochemical characteristics of many bacteria isolated from hot environments. We are also the first to report the cellulolytic activity of thermotolerant P. lactis. Described strains can be a source of new thermostable cellulases, which are extremely desirable in various branches of circular bioeconomy.

Formation of Nitrogen Doped Titanium Dioxide Surface Layer on NiTi Shape Memory Alloy.

NiTi shape memory alloys are increasingly being used as bone and cardiac implants. The oxide layer of nanometric thickness spontaneously formed on their surface does not sufficiently protect from nickel transition into surrounding tissues, and its presence, even in a small amount, can be harmful to the human organism. In order to limit this disadvantageous phenomenon, there are several surface engineering techniques used, including oxidation methods. Due to the usually complex shapes of implants, one of the most prospective methods is low-temperature plasma oxidation. This article presents the role of cathode sputtering in the formation of a titanium dioxide surface layer, specifically rutile. The surface of the NiTi shape memory alloy was modified using low-temperature glow discharge plasma oxidation processes, which were carried out in two variants: oxidation using an argon + oxygen (80% vol.) reactive atmosphere and the less chemically active argon + air (80% vol.), but with a preliminary cathode sputtering process in the Ar + N2 (1:1) plasma. This paper presents the structure (STEM), chemical composition (EDS, SIMS), surface topography (optical profilometer, Atomic Force Microscopy-AFM) and antibacterial properties of nanocrystalline TiO2 diffusive surface layers. It is shown that prior cathodic sputtering in argon-nitrogen plasma almost doubled the thickness of the produced nitrogen-doped titanium dioxide layers despite using air instead of oxygen. The (TiOxNy)2 diffusive surface layer showed a high level of resistance to E. coli colonization in comparison with NiTi, which indicates the possibility of using this surface layer in the modification of NiTi implants' properties.

Dysfunction of endothelial cells exposed to nanomaterials assessed by atomic force spectroscopy.

The study of the impact of nanomaterials on endothelial cell elasticity with the atomic force spectroscopy (AFS) can be a significant model for assessing nanomaterials toxic effects in vitro. The mechanical properties of cells exposed to nanostructures can provide information not only about cellular nano and micro-structure, but also about cell physiology. The toxicity of nanostructures is an important issue which must be carefully considered when the optimal nanomaterial is defined. There are no universal properties characterizing such a nanomaterial, i.e. depending on the intended use, the requirements can be diverse. For example, for biomedical use a nanomaterial should not negatively affect the cells or should cause the expected therapeutic or diagnostic effects in justified cases. The present study was devoted to the effects of silver nanoparticles (SNPs), multi-walled carbon nanotubes (MWCNTs) and poly(amidoamine) (PAMAM) dendrimers of 4th generation on functioning of endothelial cells. Immortalized endothelial cells were exposed for 24 h to the tested nanomaterials used in concentrations reducing cellular viability to the levels of 90 % and 75 %. The innovative nature of our work is the comparison of cell elasticity performed with various AFS probes, which enabled detection of local and global elasticity alteration caused by the nanostructures. The obtained results demonstrated changes in elasticity of endothelial cell induced by the nanostructures, which were closely correlated with the level of cellular viability, forming of actin stress fibres and elevated levels of reactive oxygen species. Trend of changes in local and global elasticity of cells exposed to nanostructures was similar, but the magnitude of the response was dependent on the selected probe. SNPs and MWCNTs evoked cells stiffening, which was correlated with changes in production levels of reactive oxygen species (ROS) and the cytoskeletal alteration. Softening of cells exposed to PAMAM dendrimers correlated with increased number of apoptotic cells and ROS production levels. Based on the obtained results we conclude, that the structure and the type of nanostructure (nanoparticle) is essential for their localization inside the cells and for the toxic effect on the endothelial cells.

Comprehensive Biological Evaluation of Biomaterials Used in Spinal and Orthopedic Surgery.

Biological acceptance is one of the most important aspects of a biomaterial and forms the basis for its clinical use. The aim of this study was a comprehensive biological evaluation (cytotoxicity test, bacterial colonization test, blood platelets adhesion test and transcriptome and proteome analysis of Saos-2 cells after contact with surface of the biomaterial) of biomaterials used in spinal and orthopedic surgery, namely, Ti6Al4V ELI (Extra Low Interstitials), its modified version obtained as a result of melting by electron beam technology (Ti6Al4V ELI-EBT), polyether ether ketone (PEEK) and polished medical steel American Iron and Steel Institute (AISI) 316L (the reference material). Biological tests were carried out using the osteoblasts-like cells (Saos-2, ATCC HTB-85) and bacteria Escherichia coli (DH5α). Results showed lack of cytotoxicity of all materials and the surfaces of both Ti6Al4V ELI and PEEK exhibit a significantly higher resistance to colonization with E. coli cells, while the more porous surface of the same titanium alloy produced by electron beam technology (EBT) is more susceptible to microbial colonization than the control surface of polished medical steel. None of the tested materials showed high toxicity in relation to E. coli cells. Susceptibility to platelet adhesion was very high for polished medical steel AISI 316L, whilst much lower for the other biomaterials and can be ranked from the lowest to the highest as follows: PEEK < Ti6Al4V ELI < Ti6Al4V ELI-EBT. The number of expressed genes in Saos-2 cells exposed to contact with the examined biomaterials reached 9463 genes in total (ranging from 8455 genes expressed in cells exposed to ELI to 9160 genes in cells exposed to PEEK). Whereas the number of differentially expressed proteins detected on two-dimensional electrophoresis gels in Saos-2 cells after contact with the examined biomaterials was 141 for PEEK, 223 for Ti6Al4V ELI and 133 for Ti6Al4V ELI-EBT. Finally, 14 proteins with altered expression were identified by mass spectrometry. In conclusion, none of the tested biomaterials showed unsatisfactory levels of cytotoxicity. The gene and protein expression analysis, that represents a completely new approach towards characterization of these biomaterials, showed that the polymer PEEK causes much more intense changes in gene and protein expression and thus influences cell metabolism.

Endothelial cell aging detection by means of atomic force spectroscopy.

Endothelial cell aging is related to changes not only in cell phenotype, such as luminal changes, intimal and medial thickening, and increased vascular stiffness, but encompasses different cell responses to various substances including drugs or nanomaterials. In the present work, time- and dose-dependent elasticity changes evoked by silver nanoparticles in endothelial cells in early (below 15) passages were analyzed. Silver nanoparticle concentrations of 3, 3.6, and 16 µg/mL were selected for elasticity measurements for long incubation (24 hours) and of 1 and 3 µg/mL for monitoring dynamic elasticity changes of 1-, 3-, and 6-hour incubations. Surprisingly, a significant reduction in the cells elasticity modulus at lower number of passages exposed to silver nanoparticles used at 3 µg/mL for 24 hours was demonstrated. These results are in contrast to those obtained for endothelial cells in late (33-43) passages that may result from cellular aging in response to nanosilver. Furthermore, for short incubation times (1 and 3 hours), SNP-induced significant increase in the cell elasticity modulus was detected. In current work, we also attempted to answer the question whether the changes in cell elasticity were induced by the silver nanoparticles stabilized with polyvinyl pyrrolidone or by stabilizer itself. Elasticity measurements were supplemented by observations made with transmission electron microscopy and scanning electron microscopy, which confirmed the presence of silver nanoparticles inside the cells and on the cell membrane. Additionally, activation of reactive oxygen species was detected for cells exposed to SNPs for 1 and 3 hours, which was accompanied by increased cell elasticity modulus suggesting a possible mechanism of observed phenomenon.

Designing laser-modified surface structures on titanium alloy custom medical implants using a hybrid manufacturing technology.

The hybrid technology combines an efficient material-removal process and implant surface treatment by the laser reducing time of manufacture process compared to currently used machining technologies. It also permits precise structuring of the implant material surface. Six structures of the Ti6Al4V ELI surface were designed and studied how the structure topography prepared with the hybrid technology affected the Escherichia coli adhesion to the surface and viability, as well as the growth, adhesion, and viability of human osteogenic Saos-2 cells cultured on the investigated surfaces. Results have confirmed that the microtopography of medical titanium alloy plays a beneficial role in bacterial adhesion and viability (number of bacteria found on reference surface: [5.9 ± 0.44] × 106 CFU/ml, sample no. 3: [8.8 ± 0.93] × 104 CFU/ml, and sample no. 5: [1.2 ± 0.23] × 107 CFU/ml; CFU - Colony Forming Unit). All tested structured surfaces enabled good cell attachment and proliferation of Saos-2 cells (viability of Saos-2 cells [% of control] for reference surface: 81.93%; sample no. 3: 75% and sample no. 5: 100%). Transcriptome analysis of genes commonly expressed in the process of osseointegration demonstrated that the use of hybrid technology allows designing structures that enhance osseointegration but it should be coupled with other methods of preventing bacterial growth, or with a different strategy to limit microbial colonization with the satisfactory osseointegration potential.

Simultaneous transcriptome and proteome analysis of EA.hy926 cells under stress conditions induced by nanomaterials.

Today, the extensive and constantly growing number of applications in the field of nanotechnology poses a lot of questions about the potential toxicity of nanomaterials (NMs) toward cells of different origins. In our work we employed the tools of molecular biology to evaluate changes that occur in human endothelial cells at the transcriptomic and proteomic level, following 24 h of exposure to three different classes of NMs. Using microarray technology, we demonstrated that 24 h of exposure to silver nanoparticles (SNPs), multiwalled carbon nanotubes (MWCNTs) and polyamidoamine dendrimers (PAMAMs) leads to changes in 299, 1271, and 431 genes, respectively, influencing specific molecular pathways. The 2D-DIGE and mass spectrometry analysis revealed that differentially expressed proteins were involved in numerous cellular processes, for example, cytoskeletal reorganization, cell growth and proliferation, or response to stress. Both, transcriptome and proteome alterations indicate reorganization of mechanism regulating cell functioning. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1024-1034, 2019.

Sensing of silver nanoparticles on/in endothelial cells using atomic force spectroscopy.

Endothelial cells, due to their location, are interesting objects for atomic force spectroscopy study. They constitute a barrier between blood and vessel tissues located deeper, and therefore they are the first line of contact with various substances present in blood, eg, drugs or nanoparticles. This work intends to verify whether the mechanical response of immortalized human umbilical vein endothelial cells (EA.hy926), when exposed to silver nanoparticles, as measured using force spectroscopy, could be effectively used as a bio-indicator of the physiological state of the cells. Silver nanoparticles were characterized with transmission electron microscopy and dynamic light scattering techniques. Tetrazolium salt reduction test was used to determine cell viability after treatment with silver nanoparticles. An elasticity of native cells was examined in the Hanks' buffer whereas fixed cells were softly fixed with formaldehyde. Additional aspect of the work is the comparative force spectroscopy utilizing AFM probes of ball-shape and conical geometries, in order to understand what changes in cell elasticity, caused by SNPs, were detectable with each probe. As a supplement to elasticity studies, cell morphology observation by atomic force microscopy and detection of silver nanoparticles inside cells using transmission electron microscopy were also performed. Cells exposed to silver nanoparticles at the highest selected concentrations (3.6 µg/mL, 16 µg/mL) are less elastic. It may be associated with the reorganization of the cellular cytoskeleton and the "strengthening" of the cell cortex caused by presence of silver nanoparticles. This observation does not depend on cell fixation. Agglomerates of silver nanoparticles were observed on the cell membrane as well as inside the cells.

Toxicity of silver nanoparticles, multiwalled carbon nanotubes, and dendrimers assessed with multicellular organism Caenorhabditis elegans.

Nematode Caenorhabditis elegans (C. elegans) was used to investigate the impact of silver nanoparticles (SNP), multiwalled carbon nanotubes (MWCNT), and polyamidoamine dendrimers (PAMAM) used in concentration of 1010 particle/mL. Population-based observations and gene expression analysis were employed in this study. SNP and PAMAM caused decrease in the number of live nematodes and their body length, but MWCNT did not affect the population of nematodes. Gene expression analysis revealed significant changes caused by the presence of all studied nanomaterials, and the results strongly suggest a specific metabolic response of the nematode organism to exposure to various nanomaterials. It was shown that C. elegans is a very sensitive organism capable to respond specifically to the exposure to some nanomaterials and therefore could be considered as a possible biosensor for early warning of presence of some nanoparticles.

Human brain endothelial barrier cells are distinctly less vulnerable to silver nanoparticles toxicity than human blood vessel cells: A cell-specific mechanism of the brain barrier?

The blood-brain barrier (BBB) constitutes a distinctive and tightly regulated interface between the brain and the peripheral circulation. The objective of studies was to compare responses of human endothelial cells representing the model of blood vessels - EA.hy926 and HUVEC cells and the model of the brain endothelial barrier - HBEC5i cells to silver nanoparticles (SNPs). A contact of SNPs with endothelial cells resulted in a formation of SNP agglomerates. Consequently, the SNPs uptake by endothelial cells affected cell viability and membrane integrity however observed responses were different. Brain endothelial barrier HBEC5i cells were much less vulnerable to SNPs toxicity comparing to EA.hy926 and HUVEC cells. It can be ascribed to the presence of specialized cellular components of the brain barrier, protecting HBEC5i cells against toxic SNPs. Fundamental understanding of SNPs inducing the BBB dysfunction may initiate engineering novel SNPs which are safe for the BBB and thereby safe for the brain.

The effect of carbon nanoparticles on biological properties of polyester nanocomposites.

The aim of present study was to determine the hemocompatibility, cellular response of endothelial cells and bacterial adhesion to a new polyester nanocomposite. The carbon nanoparticle nanocomposite was prepared via in situ polymerization of monomers to obtain material of hardness 55 Sh D similar to polyurethanes used in medical applications, for example, in heart-assisting devices. The carbon nanoparticle-containing polyester exhibits markedly reduced bacterial colonization, as compared to commercially available polyurethanes. Further the nanocomposite possesses markedly improved hemocompatibility, as determined by flow cytometry, and robust endothelialization. Possible explanations for these beneficial properties include surface nanoroughness of carbon nanoparticle-containing nanocomposites and presence of fatty acid sequences within polymer structure.

Radio-opaque polyethylene for personalized craniomaxillofacial implants.

OBJECTIVE: This study aimed to present a new possibility to create radio-opaque implant material for craniomaxillofacial reconstruction. MATERIALS AND METHODS: The test disks made of the own compound of polyethylenes with addiction of 2, 4, and 6 % of weight TiO2 was investigated for cytotoxicity [each group 15 disks respectively]. Next, computed tomography of the disks was performed in environment of muscle and fat. Hardness, tensile modulus and strength, and compressive modulus and strength were tested too. RESULTS: Deterioration of mechanical properties of the composites containing titanium dioxide was observed [hardness, tensile modulus and strength, compressive modulus and strength, respectively: 56.7 ± 1.6 shore D, 354 ± 52, 22.5 ± 1.3, 21.8 ± 1.1, and 2995 ± 327 MPa as addiction of 2 % TiO2; 52.0 ± 0.9 shore D, 347 ± 66, 18.0 ± 0.7, 14.2 ± 0.9, and 1396 ± 477 MPa as 4 % TiO2; 51.3 ± 1.3 shore D, 316 ± 9, 17.4 ± 0.2, 13.6 ± 0.6, and 1100 ± 144 MPa as 6 % TiO2 added]. The test disks revealed no cytotoxicity effect on human osteoblasts. The new material presents mild radio-opacity which was enough to observe the implant in relation to fat and muscle, but with no visible effect of beam hardening. CONCLUSION: In view of the performed tests, the polyethylene enriched by titanium dioxide seems to be a proper material to consider manufacturing of craniomaxillofacial implants. CLINICAL RELEVANCE: Maxilloafacial surgery is still looking for new implantologic materials. The proposed one is a new way to manufacture an implant visible in computed tomography which does not interfere with its shape in radiological examination and makes it possible to observe the surrounding soft tissues.

Early cell response to contact with biomaterial's surface.

Most biomaterials at present have sufficient mechanical properties; however compliance with standards for biocompatibility is often not sufficient in clinical practice. This may be due to the complexity of biological systems in general and the diversity of individual responses to these materials by implant recipients. Significant improvement of biocompatibility must involve surface modification of implants, which in the future will make it possible to introduce individually selected types of surface modification for individual recipients. The key to this technology seems to be understanding the processes occurring at the site of contact of the implant with the tissue. Processes resulting from the stress generated by the contact of the biomaterial surfaces were observed with endothelial cells line EA.hy926, and it was demonstrated that differently modified surfaces of medical steel (polished medical steel and medical steel coated with Parylene C and nanocrystalline diamond) cause diverse cellular response in cells grown on these surfaces, on both the cellular (cell morphology and cell survival) and molecular (transcriptome and proteome profiles) levels. The herein presented observations are a good starting point not only for further research and the development of far-reaching personalization of medical implants, but also to study the potential use of cells as a specific sensor capable of recognizing different surfaces with which these cells come into contact. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 104B: 880-893, 2016.

Physicochemical and Biological Investigation of Different Structures of Carbon Coatings Deposited onto Polyurethane

The aim of this study was to examine the thrombogenic properties of polyurethane that was surface modified with carbon coatings. Physicochemical properties of manufactured coatings were investigated using transmission electron microscopy (TEM), atomic force microscopy (AFM), X-ray Photoelectron Spectroscopy (XPS), Raman spectroscopy and contact angle measurement methods. Samples were examined by the Impact-R method evaluating the level of platelets activation and adhesion of particular blood cell elements. The analysis of antimicrobial resistance against E. coli colonization and viability of endothelial cells showed that polyurethane modified with use of carbon layers constituted an interesting solution for biomedical application.

Resistance of oxidative stress in biofilm and planktonic cells

This work studied the susceptibility of biofilm produced by E. coli to oxidative stress, and compared the components of free radicals defences: level of glutathione, catalase and dismutase activities in planktonic and biofilm located cells. Results showed the diversity of responses to oxidative stress in bacterial cells in log or stationary phases in both planktonic and biofilm forms. The bacteria were exposed to free-radical donors (H2O2, tBOOH, menadione, SIN-1 or peroxynitrite) in a wide range of final concentrations, from 0.5 to 10mM. Different level of toxicity of individual donors, independence of cell type (planktonic forms or biofilm) and phases of growth were observed. The highest oxidative stress resistance was observed for the cells in logarithmic phase of growth treated with H2O2, both in planktonic and biofilm forms, whereas for the cells in stationary phase, the highest resistance was observed for menadione. These results showed higher efficiency of agents based on superoxide anion donors in combating bacteria colonizing abiotic surfaces stainless steel (AISI 316L).

Proteomic analysis of proteins engaged in α-methylene-δ-lactone cytotoxic effects in hormone-independent breast cancer MDA-MB-231 cells.

A simple synthetic α-methylene-δ-lactone, 1-isopropyl-2-methylene-1,2-dihydrobenzochromen-3-one, designated DL-3, was shown previously to induce apoptosis and significantly suppress cell metastatic potential in MDA-MB-231 breast cancer cells. The mechanisms through which DL-3 exerts its effects are poorly understood. The purpose of this study was to investigate the protein expression profiles in MDA-MB-231 cells exposed to the DL-3 treatment. Using 2D differential gel electrophoresis, a set of eight differentially expressed proteins (spot intensities which showed ≥1.25-fold change and statistical significance, p < 0.05, between the control and DL-3-treated group) were found and successfully identified by mass spectrometry (MALDI-TOF/MS). The proteomic results revealed that the presence of DL-3 in MDA-MB-231 cells led to the differential regulation of some proteins that are involved in the cell cycle progression, apoptosis, cytokinesis, modulation of transcription, cellular signaling, and vesicular trafficking. The function of other identified proteins is still unknown. Therefore, our data indicate new directions for the further studies of the pathways engaged in the anticancer action exerted by α-methylene-δ-lactones in cancer cells.