Seznam abstraktů 2021: POSTEROVÁ SEKCE C
Datum: 21.10.2021
The study is focused on the wetting of nanofibrous layers of polycaprolactone. In the study, we proceeded to compare layers from direct current spinning and alternating current spinning. During the study, it was found that direct current spinning produces hydrophilic layers, and alternating current spinning produces hydrophobic layers. The study also found that the contact angle is affected by the solvent system. Due to the widespread use of polycaprolactone in tissue engineering, wetting of the fibrous layers with a gelatin hydrogel was also performed in the experiment. During the study, the Kwok-Neumann and Li-Neumann models were used to calculate the surface energy of the fibrous layers, and these models were also used to calculate the theoretical contact angles that the gelatin hydrogel will make on the fibrous layers. The study showed that the used models could predict how the fibre layers will be wetted with the prepared hydrogel.
Insulin is a peptide hormone produced by beta cells of pancreatic islets. In diabetes mellitus type 1, these islets are destroyed by the body’s own immune system, no insulin is produced and the blood glucose level is increased. Nowadays efforts in combatting diabetes type 1 focus on the transplantation of islets immunoprotected in microspheres made of non-covalently crosslinked hydrogels. The functionality of the encapsulated islets is retained, while the hydrogel matrix allows the permeation of the produced insulin into the bloodstream. The applicability of these microspheres has been extensively studied in vivo. However, a dynamic detection method to monitor the production of insulin and its diffusion through the microspheres is still missing. Herein we apply the technique of Surface Enhanced Raman Scattering (SERS) to detect physiologically relevant concentrations of insulin and its diffusion through alginate matrices using planar Ag SERS substrates. Insulin was detected in the concentration range of 10-3- 10-12 M. Preliminary in situ monitoring experiments resulted in the detection of insulin permeation through the microspheres membrane. As the secretion of insulin and its diffusion across the immunoprotective microspheres is a dynamic process, the development of such a detection method will lead to a better understanding of these processes as a function of time, matrix composition and glucose intake. ACKNOWLEDGEMENTS: Financial support of the projects APVV-14-0120 and APVV-18-0480 is acknowledged.
Hydrogels are presently under investigation due to their similarities with the native extracellular matrix (ECM). These unique materials could serve as a suitable platform for studying transport of soluble molecules. As is known, transport properties of the materials are given by their internal structure and could be greatly affected by its rearrangement. Thus the aim of this study is to investigate the influence of the lecithin nanodomains as a structure modifier on the transport properties of hydrogel systems. For this purpose physically and chemically crosslinked hydrogels, containing lecithin and cationic organic dye rhodamin 6G as a diffusion probe, were synthetized. The releasing of model active compound (organic dye) as well as structure modifier (lecithin) were determined via UV-VIS spectroscopy over the time. Based on the obtained data, effective diffusion coefficients and concentrations of the released dye were determined. Finally the limitations and advantages of the chosen hydrogels as a potential material for drug release were defined and discussed.
Design of the hydrogels with the tunable properties capable to imitate native extracellular matrix (ECM) is widely discussed topic in recent years. With respect to faithful imitation of ECM, this proposed study mostly utilizes biocompatible and biodegradable materials with ability to form crosslinked 3D networks (hydrogels) with further potential use as barrier materials, drug carrier in regenerative medicine, tissue engineering, etc. The inner structure (related to viscoelastic and transport properties) of the basic 3D matrix material were modified by phospholipid lecithin, which in water-based solutions forms organized nanodomain structures and consequently affects the final structural properties of the prepared hydrogels. These structural modifications then consequently lead to distinct mechanical properties.
Hydrogel systems, physically, chemically, and ionically crosslinked, were subjected to the structural characterization in the dried form of xerogel using direct visualization technique (scanning electron microscopy) and in swollen form of hydrogels by further evaluation of the viscoelastic properties using rheology. Complex viscoelastic characterization consisted in the comparison of strain sweep and frequency sweep tests of the prepared hydrogels and rehydrated hydrogels prepared by swelling of xerogels. Appropriate amount of lecithin, the type of material chosen for the hydrogel preparation, the state of the gel before and after drying and rehydration were discussed as well.
The nanofibrous material with an incorporated antibiotic was developed and tested. The introduced structure could serve as promising prevention of the life-threatening postoperative in gastrointestinal surgery associated with the occurrence of bacteria. Polymer solutions of poly-ε-caprolactone (PCL) with antibiotic Gentamicin sulfate (GS), in concentrations 0; 1; 3; 5; 7,5 and 10 wt. % were electrospun via needleless NanospiderTM device. The produced layers were characterized by scanning electron microscopy and their wettability was measured. The amount of antibiotic in the layers was tested by UV-spectrophotometry and high-performance liquid chromatography (HPLC). The cytotoxicity of the material extracts was evaluated on the 3T3 mouse fibroblast cell line. The layers were evaluated as cytocompatible. Antibacterial tests were performed to confirm the presence release and efficiency of the antibiotic GS from the nanofiber layers. The effect of ethylene oxide sterilization on nanofibrous layers with incorporated antibiotic was also studied throughout the experiment part. Sterilization did not cause any significant changes in the nanofibrous materials with an incorporated antibiotic, and the sterile layers had antibacterial properties. This research was supported by Czech Health Research Council project AZV NU20J-08-00009 Prevention of intestinal anastomotic leakage and postoperative adhesions by using nanofibrous biodegradable materials.
The prevalence of chronic wounds is increasing due to population ageing and specific illnesses like diabetes mellitus and vascular diseases. Nanofibrous membranes fabricated using synthetic polymers are promising materials to enhance skin wounds healing. PCL and PVA membranes are being studied to be used as skin scaffolds and hydrogels, respectively. The present study considers the development of a multi-layered membrane made of PCL and PVA loaded with platelet lysate. PCL nanofibers allowed cell adhesion and growth, whereas PVA acted as a hydrogel that releases the bioactive compounds of platelet lysate. The cytocompatibility of the membrane containing platelet lysate and without it was demonstrated on two cell types involved in wound healing, keratinocytes and fibroblasts. Both cell types were able to adhere and proliferate on the membranes. In addition, the membrane containing platelet lysate enhanced the proliferation of fibroblasts. A co-culture study was also performed by seeding each cell type on one side of the membrane. Cells were co-cultured for 7 days and results showed that the platelet lysate increased the proliferation of cells achieving a monolayer of keratinocytes and fibroblasts on each side of the membrane. Thus, the beneficial effect of PCL-PVA+PL membranes on monocultures and co-cultures was demonstrated and can be considered a potential scaffold to treat chronic wounds.
Electrospun PLCL nanofibrous materials produced by direct current (DC) or alternating current (AC) electrospinning are good candidates for absorbable reinforcement for cardiovascular surgery composite bandages. For an appropriate description of the fiber layers and prediction of its relevant biomechanical characteristics, the knowledge of fiber orientation in the electrospun layers is essential. In this study, the different orientation of fibers in AC and DC electrospun materials was achieved only by changing the rewinding speed of the supporting material, as the easy way for the production of relatively large samples with controlled fiber orientation. The fiber orientation was measured on scanning electron microscope images at different magnification using an open-source image processing software FIJI/Image J with the implementation of the "OrientationJ" plugin. Subsequently, the fiber layers were compared in terms of fiber orientation. Optimal image parameters for the measurement and the best unidirectional oriented-aligned samples were determined.
Vascular diseases usually destroy native vessels of patients, and very often, it is not possible to restore the original vessel or replace the damaged vessel with an autologous transplant. That is why nowadays there is a big need for artificial small-diameter vascular grafts. These grafts should have sufficient mechanical and morphological properties. In this study, small-diameter (6 mm) tubular scaffolds were fabricated using needleless electrospinning process. Two groups of scaffolds with different wall thickness were made, specifically 300 and 400 micrometers. Needleless electrospinning is a very new technology for producing tubular samples which is not mentioned in many articles yet. As a polymer was used polycaprolactone (PCL). PCL is one of the representatives of synthetic biodegradable polyesters, it has good mechanical properties and hemocompatibility, and that is why PCL is commonly used in tissue engineering. After production the scaffolds in both groups were analyzed morphologically and mechanically. At least 400 diameters were measured from the outer and inner layer separately and compared together. Tensile strength was investigated in the longitudinal direction, and burst pressure was investigated in the radial direction. Than also the contact angle and porosity was measured. Results of these measurements were compared and the difference between two groups of samples was investigated.
Applications of cyclodextrins, being cyclic oligomers containing an internal hydrophobic cavity, have been frequently studied for uses in medicine, chemistry, pharmaceuticals, water treatment, and food processing. The highly polar cyclodextrin compound containing an inherent chiral and hydrophobic cavity give possibilities for its application in wide array of areas. Generally, the hydrophobic cavity is exploited to form host-guest interactions to stabilise molecules in aqueous environments, for example, in pharmaceuticals whereby the incorporation of poorly water-soluble drugs increases both their bioavailability and stability.
Electrospun nanofibers have continually attracted increasing amounts of attention, predominantly due to the high specific surface area as well as their controllable surface chemistry and treatment. Melding the benefits from nanofibers with that of cyclodextrins has recently become a focus of much research. Cyclodextrin can be electrospun into nanofibers, forming fibers solely through non-covalent intermolecular interactions, however, these fibers dissolve rapidly in contact with aqueous solvents and hence their applications are highly limited. Conversely, the formation of nanofibers with cyclodextrin being covalently attached or incorporated with the fibers are often formed using a carrier polymer or copolymer during the spinning process, with the carrier/copolymer often not being compatible for the desired uses. To overcome these dependencies, the formation of specific disubstituted cyclodextrins could allow for new avenues for cyclodextrin polymerization and nanofiber formation to be explored without any of the common problems associated with using copolymers. Through capping of β-cyclodextrin, a technique to produce a stereo-isomerically pure product, followed directly by substitution to one of various reactive groups can be controlled to produce di-substituted functionalised β-cyclodextrin. Therefore, the functional groups can then be further reacted through a site-specific known mechanism to incorporate groups which can then be polymerised, via processes such as sol-gel, for the formation of polymeric β-cyclodextrin chains. Subsequently, these chains can be electrospun which could drastically improve the applicability of cyclodextrin nanofibers in controlled drug delivery, wound healing, bioenegineering, and water treatment.
Biodegradable electrospun materials are widely used for medical application. Polycaprolatone is polymer suitable for electrospinning technology and is very often used to create nanofibrous scaffolds for tissue engineering. The time to disintegration or biodegradation of such materials is very important here. However, testing is not entirely easy. It is not possible to create exactly body-like conditions in vitro. Moreover, it is not easy to find suitable analytical methods that would show exactly what happens in the nanofibrous polycaprolactone electrospun samples at certain stages of degradation, ie how the internal structure of decaying nanofibers changes. This paper describes the traditional use of methods for testing polycaprolatone nanofibers by enzymatically catalyzed degradation. Morphological changes are studied using scanning electron microscope images. However, it also offers a non-traditional analysis of polycaprolactone electrospun materials using the ssNMR method.
Threads are the basic textile linear structure used in many areas. The variability of the thread structure made possible by special constructions brings great possibilities of variability. It is also possible to introduce nanofiber material into the yarn construction as a cover for central microfiber yarns, and these can be further intertwined into more complex yarn constructions. Continuous production of so-called composite yarns containing central microfiber yarns and nanofiber wrappers is made possible by alternating current electrospinning. Subsequent braiding of these composite yarns allows the protection of nanofiber packages and the variability of the overall construction of the resulting yarns. This paper introduces the creation of yarns and various types of basic as well as braided yarns with nanofibers. Selected sutures were further subjected to in-vitro testing to verify cytocompatibility for application in hygiene such as dental sutures or medicine as surgical sutures.
Anastomotic leakage and peritoneal adhesions belong to major complications after colorectal resection. These life-threatening complications are diagnosed in the postoperative period and can lead to sepsis and thus higher mortality, reoperation, intestine blockage, chronic abdominal pain and others. In this study, the nanofibrous material based on biodegradable poly-e-caprolactone (PCL) serves as a fortification of gastrointestinal anastomoses and thus prevents leakage and formation of fibrin adhesions. The nanofibrous layers were fabricated via needle-less electrospinning technique called NanospiderTM, that is available at the Department of Nonwovens and Nanofibrous Materials at the Technical University of Liberec. The materials were further functionalized by using the plasma enhanced chemical vapour deposition (PA CVD) with hydrophobic hexamethyldisiloxane to enhance the anti-adhesive properties. The morphology of the prepared layers was assessed via scanning electron microscopy, the cytocompatibility was proved according to ČSN EN ISO 10993-5. The contact angle measurement testified the successful increasement of hydrophobic behaviour after PA CVD treatment. The materials were examined during 6 months period via FTIR spectroscopy to prove the stability of the treatment. Currently, the materials are tested in vivo to verify the prevention of anastomotic leakage and peritoneal adhesions.
ACKNOWLEDGEMENTS: The research was supported by the project MZ ČR AZV NU20J-08-00009 Prevention of intestinal anastomotic leakage and postoperative adhesions by using nanofibrous biodegradable materials.
Recently, nanofibres-based wound dressings and drug delivery (DD) systems have attracted attention for their unique properties. Nanofibres-based wound dressings possess exceptional properties including high specific surface with small pore size enabling drug delivery and release, while keeping the wound site moisturized and opened to an air exchange at the same time. This leads to stimulation of the wound site simply by the morphology and released material of the nanofibres. Therefore, the chemical composition of the nanofibres plays an important role being able to affect their degradation behaviour, biocompatibility and bioactivity. One of the extensively studied materials for wound dressing intended nanofibres construction is chitosan. It is characterised by variety of sources and deacetylation degrees available, good biocompatibility and natural antibacterial activity related to its overall positively charged molecule, which can be beneficial for difficult-to-heal wounds treatment.
The aim of this study is to evaluate natural antibacterial properties of chitosan-based nanofibres and their biocompatibility in vitro and impact of the thermal treatment, necessary for nanofibres stabilisation, on these properties. Antimicrobial activity of untreated and treated nanofibres is tested on model bacterial strains S. gallinarum and E. coli. Biocompatibility is tested on 3T3 fibroblast cell line and Hacat keratinocytes in vitro.
Recently discovered two-dimensional (2D) transition metal carbides and nitrides (MXenes) have received tremendous attention because of their unique electrical, optical and chemical properties. These exceptional properties make them a suitable candidate for a variety of applications including multimodal tumor therapy by photothermal effect. In this work, we demonstrate how to reduce the size of 1 5 µm large Ti3C2 monolayer MXene sheets into ultrasmall 120-160 nm nanosheets by applying consecutive ultrasonication processes. Different microscopic techniques have been used to visualize the formation of ultrasmall single layer Ti3C2 nanosheets. The as-prepared MXene nanosheets have shown good solubility in water and ethanol. Further, (3-aminopropyl) triethoxysilane (APTES) and poly(3,4-ethylene dioxythiophene) polystyrene sulfonate (PEDOT:PSS) were utilized for surface modification of the MXene nanosheets to open the possibility of subsequent antibody bio-conjugation. Moreover, PEDOT:PSS improved the photothermal conversion performance of the nanosheets as documented by increasing their temperature from 48.60 ºC to 58.10 ºC on irradiation by 808 nm wavelength laser. Further in vivo and in vitro studies will be necessary to optimize photothermal properties of Ti3C2 nanosheets. ACKNOWLEDGEMENTS: Financial support of the project APVV-19-0465 is acknowledged.
Upconverting luminescent lanthanide-doped nanoparticles (UCNPs) belong to promising new materials that absorb infrared light able to penetrate in the deep tissue level, while they emit photons in the visible or ultraviolet region, which makes them favorable for bioimaging and cell labeling. However, for medical applications colloidal stability of the particles in biological media has to be ensured. The selection of the proper covalent binding by a hydrophilic polymer also affects mechanism of the particle engulfment by cells.
In this report, differently charged polymer-coated core-shell UCNPs, uniform in size with diameter of 28 nm, were synthesized by a high-temperature coprecipitation of lanthanide chlorides in a high-boiling organic solvent. The synthesized nanoparticles were characterized by transmission electron microscopy, dynamic light scattering, luminescence spectroscopy and energy-dispersive X-ray analysis.
UCNPs were coated with poly(N,N-dimethylacrylamide) (PDMA) copolymers that were synthesized by copolymerization of N,N-dimethylacrylamide (DMA) with functional comonomers, such as 2-(acryloylamino)-2-methylpropane-1-sulfonic acid (AMPS) or tert-butyl [2-(acryloylamino)ethyl]carbamate (AEC-Boc) by reversible addition-fragmentation chain-transfer polymerization. We investigated the effect of negatively charged sulfo groups-containing P(DMA-AMPS), positively charged P(DMA-AEC), and two electroneutral polymers, PDMA and poly(ethylene glycol) (control), on engulfment of the particles in human hepatocellular carcinoma HepG2, human cervical cancer HeLa, and rat insulinoma INS-1E cells. The polymers contained both bisphosphonate anchoring groups covalently bound to the surface of UCNPs and amino or sulfonate groups. Optionally, the polymers were labeled with DY-615 fluorophore to make fluorescent imaging of carcinoma cells possible, allowing at the same time to control the stability of both surface-coated nanoparticles in the cell medium.
During the last decades, a great deal of attention was devoted to novel lanthanide-based upconverting nanoparticles (UCNPs) due to their perspective applications in medicine, as both therapeutic and diagnostic imagining agents, as well as in energy conversion and security applications. However, to obtain high-quality UCNPs, it is needed to overcome some challenges during the synthesis and particle surface engineering.
This study represents the use of UCNPs in photodynamic therapy (PDT). For this purpose, UCNPs were modified with Rose Bengal (RB) as a photosensitizer, which can produce reactive oxygen spices (ROS) while being excited. ROS then damages target cells, which is basis of PDT of tumors.
To achieve this goal, the synthesis and surface engineering of NaYF4:Yb,Er UCNPs were established by conjugation with RB as an active compound and poly(ethylene glycol) as a stabilizer. To bind RB to the particle’s surface, it was modified with phosphonate group known as an effective anchoring moiety for UCNPs. The colloidal stability of particles was achieved by the immobilization of PEG-alendronate (PEG-Ale) that is highly hydrophilic to the particle surface. ROS generation was monitored by 9,10-diphenylanthracene fluorophore that can bind singlet oxygen species while reducing absorbance in uv-visible spectrum.
ACKNOWLEDGEMENTS: This work was supported by MŠMT LTAB 19011.
Microcontainers are micrometer-sized cylindrical reservoir-based devices serving as drug carriers for oral administration systems with ensuring a high drug concentration at the desired site of absorption and providing a sustained resident time. These polymeric micro-cylinders consist of drug-filled cavities and can be functionalized by sealing with polymeric lids to protect the active pharmaceutical ingredient against degradation before reaching the absorption site. The present study aimed to develop biopolymer-based lids for microcontainers using polysaccharides for the prolonged release of an antibiotic. Amoxicillin was chosen as a model drug and loaded into microcontainers using the polydimethylsiloxane masking technique, followed by spray coating with chitosan (CS) and chondroitin sulfate (ChS) as polymeric lids onto the cavity of the devices. The structural and physicochemical properties of the microcontainers were investigated. Moreover, the effect of polysaccharide-based lids on the drug release behavior was further analysed and discussed. The thickness and morphology of the lids were characterized by profilometry and SEM before and after spray coating. The in vitro release profiles revealed a fast and an immediate release of amoxicillin from only ChS-coated microcontainers as in the uncoated ones, and slow, prolonged release from only CS-coated ones. However, the microcontainers with CS-ChS lids showed a biphasic release pattern that consists of a reduced burst release in the early stage of treatment, followed by a more sustained release. Therefore, the formation of polyelectrolytes between CS and ChS plays an important role in controlling the release process. The results suggest that the CS-ChS based lids for microcontainers can be useful in biodegradable, tailor-made controlled drug delivery systems.
This contribution is focused on the interaction between positively charged catanionic vesicles and oppositely charged hyaluronan. Catanionic vesicles are formed from so called ion pair amphiphiles (IPA). Charged catanionic vesicles can also bring another benefit in the interaction with oppositely charged (bio)polymers through electrostatic interactions. This interaction can also improve the higher stability and biocompatibility of vesicles [1]. Polysaccharide hyaluronan, which as a polymer present in the human body is fully biocompatible and fully biodegradable, appears to be very suitable for this purpose.
Vesicles were prepared from hexadecyltrimethylammonium bromide (HTMAB) and sodium dodecyl sulphate (SDS) and they were doped with positively charged dioctadecyldimethylammonium chloride (DODAC) and cholesterol. For interaction with them, hyaluronan of four different molecular weights (16, 418, 750, and 1670 kDa) was used. Size and ζ-potential titrations were used to characterize these interactions. The ζ-potential decreased with increasing addition of hyaluronan for all molecular weights. During higher additions, the formation of larger aggregates was observed. Around the point of charge equilibration, visible aggregates were made (larger than 1000 nm), which gradually disintegrated with continued addition of the polymer due to increased electrostatic repulsion or dilution.
ACKNOWLEDGEMENTS: This work was supported by the Czech Science Foundation, project No. 19-14024J (GACR), and Ministry of Science and Technology, Taiwan, project No. MOST108-2923-E006-MY3.
REFERENCE: [1] M. Havlíková, J. Szabová, A. Jugl et al. Study of cholesterol’s effect on the properties of catanionic vesicular systems: Comparison of light-scattering results with ultrasonic and fluorescence spectroscopy, Colloids Surfaces A Physicochem. Eng. Asp. 607 (2020) 125526. https://doi.org/10.1016/j.colsurfa.2020.125526.
Fibrous Polyacrylonitrile-based hierarchically structured nanocomposites have been produced by wet-spinning and decorated by microwave plasma-enhanced chemical vapour deposition (MPECVD). By varying CH4/H2 ratio, different fibrous composites by morphology and chemical composition have been realized and investigated as metal-free electrodes for the enhanced electrochemical detection of paracetamol. Results showed how the higher CH4 content in the gas mixture during the MPECVD process led to the production of carbon nanowall-decorated polyacrylonitrile fibers, characterized by a higher content of sp2 carbon and by the lowest peak-to-peak separation, suggesting a faster electron-transfer kinetics.
A pathological Hallmark of Alzheimer's Disease (AD) Is the significant deposition of amyloid beta (Aβ) plaques in the brain [1]. Several studies were reported in inhibit the Aβ fibrillation using nanoparticles, nanoclusters and bioconjugate Nanomaterials for Alzheimer's treatment [2]. In recent years, natural compound (photochemical) have been largely used to study and identified as promising agents for the prevention and treatment of neurodegenerative diseases including AD [3]. Gold nanoparticle, with different size and shapes enable their multifunctional properties used in biomedical apply applications. Gold nanoparticle with appropriate surface conjugates with higher affinity has been studied and acknowledges the interaction between Bio Nano interfaces [4].The surface covering or anchoring conjugates will play a major role in vitro and in Vivo.Herein, We reported Epigallocatechin gallate (EGCG stabilized gold nanoparticle (AuNps) were synthesized as a A Aβ inhibitor. The synthesized EGCG- AuNps characterized using UV-Vis Spectroscopy, DLS, XRD, SEM (EDAX) and TEM (SAED) and the biocompatibility were studied by using MTT and scratch assay. The synthesized EGCG-AuNps inhibiting Aβ aggregation, dissociation Aβ fibrils which lead to prevent plaque formation, which confirmed using Thioflavin T- assay and TEM studies. Thus, this work provides new insights into the synthesized a AuNps as functional therapeutic agent for AD.
The photothermal effect of transition metal carbides known as MXenes has been extensively studied since their discovery. However, the vast majority of applied research has only focused on anticancer therapies and drug delivery systems. In our work, we investigated applying of MXenes as an active part of enzymatic support, which allows to control the efficiency of biocatalysis via IR laser irradiation. The biocatalyst was formed by encapsulation of β-D-fructofuranosidase (invertase) with magnetic iron oxide nanoparticles (Fe3O4/γ-Fe2O3) and Ti3C2Tx MXene flakes in alginate hydrogel beads. The invertase is an enzyme that catalyses hydrolysis of sucrose to fructose and glucose. The proposed biocatalyst system offers the following advantages, namely: i) immobilization in alginate beads allow to control the area of the enzymatic reaction, ii) presence of the magnetic nanoparticles grants possibility of “on/off” switch by removing the biocatalyst beads from reactor via magnetofection, and iii) presence of the MXenes allow to control the optimal temperature of the enzymatic process via external laser irradiation (808 nm) of the hydrogel beads. All nanomaterials were characterized by SEM, XRD, Raman. Four different batches of biocatalysts were prepared by encapsulation of invertase, magnetic nanoparticles and MXene flakes in alginate beads: invertase as a reference, invertase with chosen material and with both nanomaterials. Hydrogel biocatalysts were analyzed via crio-SEM, while their xerogels by SEM, XRD and Raman. The biochemical analyses included estimation of protein concentration in biocatalysts and their enzymatic activity via estimation of glucose concentration in reactant solution.
The experiments in the present work describe the calorimetry-founded characterization of a potential surfactant-based colloidal drug delivery system. Besides other techniques, isothermal titration calorimetry (ITC) was used to determine the critical micelle concentration (cmc), Gibbs energy- and enthalpy change of micellization of mixed micelles [1]. These self-assembled and nanosized semi-particles composed of the hexadecyl(trimethyl)ammonium bromide (CTAB) and 2-[4-(2,4,4-trimethylpentan-2-yl)phenoxy]ethanol (TX100) at different molar ratios. Besides the mentioned thermodynamic parameters, entropy- and the heat capacity change of self-assembling process were evaluated using a spreadsheet-edited routine developed for handling ITC data. The thermodynamic parameters describing the, were obtained from results of temperature dependence of ITC results according to the integrated form of the van't Hoff equation.
Further, the value of thermodynamic parameters the uncertainty of these data is also important, in this way, a weighted resampling ‘‘jackknife’’ procedure was used for calculation of the standard deviations of the fitting parameters. Thanks to the spreadsheet implementation of ITC analysis, our evolution method does not require any user input other than the ITC data in integrated and normalized form and the thermodynamic parameters can be predicted using the estimation protocol developed by our researchers.
According to the calorimetric profiles of the dilution of aqueous CTAB and TX100 micelles and their mixtures, the cmc values were determined at different temperatures and compositions. In the investigated temperature range and whole micelle mole fractions interval, we found some states where the cmc is lower than the ideal mixing model calculated value. These equimolar binary surfactant mixtures showed higher solubilization capacity for poorly water-soluble drugs than their individual compounds. Thus, the rapid and fairly accurate calorimertric analysis of mixed micelles can lead to the successful design of a nanoscale drug carrier [2].
ACKNOWLEDGEMENTS: The research was supported by the National Research, Development and Innovation Office-NKFIH through FK 131446. The Ministry of Human Capacities, Hungary grant TUDFO/47138-1/2019-ITM is acknowledged.
REFERENCES: [1] Juhász Á, Tabajdi R, Dékány I, Csapó E. Thermodynamic Characterization of Temperature- and Composition-Dependent Mixed Micelle Formation in Aqueous Medium. J Surfactants Deterg 2017;20:1291-9. https://doi.org/10.1007/s11743-017-2025-x.; [2] Csapó E, Szokolai H, Juhász Á, Varga N, Janovák L, Dékány I. Cross-linked and hydrophobized hyaluronic acid-based controlled drug release systems. Carbohydr Polym 2018;195:99-106. https://doi.org/10.1016/j.carbpol.2018.04.073.
Among non-viral gene carriers with low toxicity and high transfection efficiency, the use of gold nanoparticles (AuNPs) is of particular interest due to their biocompatibility and special properties. This is the first time we attempted to manipulate the surface of biological AuNPs in order to conjugate them with ssRNA through electrostatic interactions. A fungal strain, Fusarium oxysporum, was cultured in Sabouraud Dextrose Broth (SDB), centrifuged, and the mycelium-free supernatant was chalenegd with 1 mmol final concentration of HAuCl4.3H2O and incubated for 24 h at 37°C in a shake flask. AuNPs were characterized by visible spectrophotometry, Transmission Electron Microscopy (TEM), Scanning Electron Microscopy (SEM), Energy-Dispersive X-ray spectroscopy (EDS), and a zetasizer. The washed and sterilized AuNPs were used for cytotoxicity and conjugation assays. Transferrin (Tf), PEI, and antimiR were conjugated to the AuNPs and their association was confirmed by visible spectrophotometry and electrophoresis. Confocal microscopy was used to investigate the internalization of the complex. The results proved the formation of AuNPs with a maximum absorption peak at 528 nm, round and oval shapes (15-20 nm), and average zeta potential of -21.02. The AuNPs- ssRNA- Tf- PEI showed delayed electrophoresis unlike ssRNA. Functionalized AuNPs did not cause any toxicity in cell culture and confocal microscopy showed successful transfection of ssRNA- Tf- PEI-AuNPs into the vast majority of 4T1 cells. We concluded that the biological AuNPs were non-toxic and they could carry ssRNA to enable gene silencing.
One of the biggest concerns of nanomedicine is the long-term fate of nanoparticles in the body. Each treatment substance should be safely eliminated from the organism after fulfilling their therapeutic purpose. The results from studies focused on nanoparticle-based treatment agents indicate their accumulation in some organs short after application [1][2], but little is known about longer time periods. Gold nanoparticles are wide studied as a promising tool for e.g. drug delivery, imaging or photothermal therapy, therefore it is important to investigate their long-term effect on the organism.
In our study, we applied the sphere shaped gold nanoparticles (AuNPs) with 10 nm in diameter and coated with bovine serum albumin (BSA) to C57BL/6 mice. After intravenous administration, mice were observed and regularly weighted for 120 days. At the end of the experiment, organs - liver and spleen were chirurgical extracted and analysed.
The health of mice was not affected by the AuNPs application for duration of the experiment, despite we were able to detect AuNPs in the liver and spleen tissue by atomic absorption spectrometry even after 4 months from AuNPs application. Interestingly, both organs were slightly heavier in comparison to control group and whether this could be a direct effect of accumulated AuNPs is the object of further investigation. Thus our results indicate, that AuNPs accumulate in sites of excretion - liver (the main excretory organ for particles above the renal filtration limit) and spleen (splenic clearance) and persist there at least 4 months.
ACKNOWLEDGEMENTS: This work was supported by the Slovak Research and Development Agency under the contract No. APVV-16-0579
This study was performed during the implementation of the project Buildingup Centre for advanced materials application of the Slovak Academy of Sciences, ITMS project code 313021T081 supported by Research & Innovation Operational Programme funded by the ERDF.
REFERENCES: [1] Bailly, A. L., Correard, F., Popov, A., Tselikov, G., Chaspoul, F., Appay, R., ... & Esteve, M. A. (2019). In vivo evaluation of safety, biodistribution and pharmacokinetics of laser-synthesized gold nanoparticles. Scientific reports, 9(1), 1-12.; [2] Brown, A. L., Kai, M. P., DuRoss, A. N., Sahay, G., & Sun, C. (2018). Biodistribution and toxicity of micellar platinum nanoparticles in mice via intravenous administration. Nanomaterials, 8(6), 410.
A novel 2D nanomaterial - few-layer black phosphorus (FLBP) - was modified by peptide bioconjugates towards enhancing its therapeutic efficiency by reducing its side effects and improving its selective binding ability towards a biological target. Two various strategies of BP modification using poly-L-lysine (PLL) and NH2-PEG-OH conjugates with functionalised peptides were conducted.
The application of covalent immobilization protocol to coupling PLL linker with peptide allow to stronger bond formation and more stable hybrid than physisorption. The biocompatible FLBP-PLL-peptides conjugates are synthesized according to proposed non-covalent strategy. The attachment of cationic linker with peptide to surface of black phosphorus is stabilized by electrostatic and hydrophobic interaction. The proposed non-covalent functionalization of BP surface by poly-L-lysine improved nanomaterial stability. Application of RP-HPLC analysis for preliminary screening of covalent conjugation of linker and peptide proved to be efficient and practical method to be used in further investigations.
PEG was chosen as a linker for two reasons, firstly it protects FLBP structure from outer water, oxygen and non-specific interior interactions and increase stability of FLBP in physiological conditions. Furthermore, the use of non-ionic and hydrophilic linker can overcome problem of nonspecific adsorption upon the surface, e.g., by proteins from blood serum or a bioanalyte sample. Secondly, a stable immobilization of peptides on PEG linker prolonged circulation in the blood and prevent drug release during its transport which may improve pharmaceutical properties of peptides.
In conclusion, we have successfully fabricated nanoparticles of BP-PEG-peptide and BP-PLL-peptide conjugations. Two types of linkers PEG and PLL were compared as a spacer molecule due to their different polymer mass, structure and biological activity. The effect of these parameters was demonstrated in the in vitro experiments.
In recent years, biocompatible and biodegradable drug carriers offering controlled and targeted drug release have a huge role in the fields of medical and pharmaceutical research. For such systems, it is not easy to monitor their route and possible accumulation in the human body. It is an important task to develop non-invasive examination procedures, such as the labeling of drug carriers with fluorescent markers. In this work we chose the fluorescent gold nanoclusters based on their advantageous characteristics compared to fluorescent dyes or quantum dots.
The aim of our research was to prepare chitosan and hyaluronan polysaccharide based nanoparticles capable of encapsulating different drugs (pharmaceutical drugs, vitamins). We thoroughly investigated the interaction between the two polymers and the effect of different parameters. Based on the results, we prepared kinetically stable nanocarriers with well-defined size and surface charge. The drug loading capabilities of the carriers were tested for tocopherol. The drug carriers were subsequently labeled with fluorescent bovine serum albumin- or lysozyme-stabilized gold nanoclusters. To ensure the proper stability and fluorescence of the system, the interaction between the carrier and the marker, the effect of labeling on structure, size and surface charge were investigated. Finally, bimetallic silver/gold nanoclusters with stronger fluorescence were also tested.
ACKNOWLEDGEMENTS: The research was funded by the National Research, Development and Innovation Office -NKFIH through projects GINOP 2.3.2-15-2016-00034 and FK131446. E. Csapó thanks the financial support of the “Momentum” Program of the Hungarian Academy of Sciences (LP2021-5).
The properties of hydrogels must vary according to the specific needs of individual applications, e.g., for drug delivery systems, suitable transport properties are crucial to their application. One of the basic tools for modifying the material properties of hydrogels is a manipulation of the internal chemical and morphological architecture. Using semi-interpenetrating polymer networks (semi-IPN) is a suitable way to modify morphological architecture. Therefore, this work focused on monitoring the influence of an incorporated polyelectrolyte component on the resulting transport properties and internal architecture of a model hydrogel system.
Poly(hydroxyethyl methacrylate) was the first synthetically prepared hydrogel to be used for medical applications (contact lenses). Due to its application potential and its extensive use in medicine, it was chosen as the model hydrogel system for this work. Sodium polystyrene sulfonate was used as the interpenetrating linear polymer.
Scanning electron microscopy was chosen as a suitable method for structural analysis, where the samples were examined in a dried form. To investigate transport and barrier properties, solutions with dye Rhodamine 6G in various concentrations were used. Transport properties were monitored by diffusion tests (absorption of the dye into the internal structure of the gel, subsequent elution, and monitoring of diffusion from the gel). Furthermore, diffusion through the gel (barrier properties) was monitored using diffusion cells (Franz cells, horizontal cells). Finally, fluorescence correlation spectroscopy was used to determine the diffusion coefficient at the microscopic level.
The noble metal nanoclusters are one of the most interesting nanomaterials thanks to their unique optical, electronic, and magnetic properties, which explains their applications as potential biosensors and bio-labeling agents. In our research, we had to pay close attention development of several green chemistry synthesis methods to produce noble metal nanostructure having the tunable optical feature. These nanostructures can be synthesized using simple biomolecules as reducing and stabilizing agents at the same time. Depending on the molar ratio of the precursor metal ion and the ligand, their size and optical property can be tuned. Namely, the small ligand excess results in the formation of plasmonic nanoparticles with a diameter larger than 2 nm. In contrast, at high biomolecule excess sub-nanometer-sized nanoclusters (NCs) are formed (d < 2nm), which have well-defined photoluminescence [1].
During the presented simple biocompatible syntheses, adenosine monophosphate (AMP) molecule was used as stabilizing and partially reducing agent due to its great metal ion binding and reducing capability. We firstly demonstrated the dominant effect of the reaction conditions (e.g. bioligand/precursor ration, pH, temperature) on the optical properties of the formed gold-, copper- or silver-contained nanohybrids. The synthesized nanoproducts were analyzed by several techniques. The optical features were determined by UV-Vis spectrophotometry and fluorescence spectroscopy. The morphology and the size of the nano-object were characterized by HRTEM and DLS measurements, while the structural analysis was performed by using FT-IR, XRD, and MS techniques. The oxidation state of metal atoms was examined by XPS. The produced NCs are promising nanomaterials as optical biosensors for metal ions and biologically active vitamin molecules [2,3].
ACKNOWLEDGEMENTS: The research was supported by the National Research, Development, and Innovation Office-NKFIH through the GINOP-2.3.2-15-2016-00038, FK131446, and PD137938 projects.
REFERENCES: [1] D. Ungor et al. Nanomaterials 9 (2019) 1229-1247.; [2] D. Ungor et al. Colloids Surfaces B: Biointerfaces 155 (2017) 135-141.; [3] D. Ungor et al Journal of Molecular Liquids 338 (2021) 116695.
Graphene oxide is a nanomaterial that has been studied in recent years for practical use in a number of fields, such as electronics, chemical industry or medicine. Due to its properties such as hydrophilicity, strength and a large specific surface area with the possibility of functionalization, graphene oxide is a particularly attractive material for use in targeted drug delivery. This is because two-dimensional nanomaterials, such as graphene oxide, are capable of carrying drug molecules and, with appropriate modifications, transport them in a targeted manner to a site of action. Such targeted treatment is important, for example, in the field of cancer treatment, where drugs work on the principle of cell killing, and it is therefore very important that they act in a targeted manner in tumor cells and do not damage healthy cells.
Here we present the penetration of graphene oxide into the tumor cells of cervical cancer (HeLa) using MCR-Raman spectroscopic method. We apply MCR algorithm for the Raman spectra of GO in HeLa cells and quantitatively determine its composition by comparison to the Raman spectra of the individual pure forms. This MCR-Raman method offers a promising approach for the determination of graphene oxide in HeLa cells and advantages of Raman spectroscopy, which is a well-established tool in the research of cancer treatment and in the study of targeted drug delivery.
With the growing incidence of multi-resistant bacterial strains, it is necessary to seek new approaches for the treatment of complicated bacterial superinfections. Antimicrobial resistance and the slow development of alternative therapeutic agents are bringing society into the so-called post-antibiotic era. Phage therapy can be used as an alternative approach. The methods for detailed characterization of interactions of lytic agents with bacterial cells and monitoring the bacterial lysis in native conditions are required. Here, we present advanced methods for studying lysis of opportunistic pathogen Staphylococcus aureus by enzyme lysostaphin and Podoviridae phage P68. Surface plasmon resonance (SPR) biosensors allow sensitive monitoring of changes at the sensor surface in real-time at physiological conditions. We revealed that the disintegration of bacteria starts immediately after injection of lysostaphin, whereas there was a concentration-dependent delay in the case of phages. Furthermore, atomic force microscopy (AFM) was used for real-time visualization and monitoring of changes in mechanical properties of bacteria during lysis at high resolution. In the case of phage-mediated lysis, the lytic cycle was realized in the growth medium at 37°C. The changes in Young's modulus were also determined, providing a view into the changes in bacterial cell wall stiffness during the lytic processes. The combination of SPR and AFM has proven to be suitable for detailed studies of bacterial lysis while maintaining native conditions. These advanced methods will provide better insight into bacterial lysis and can help in the selection of lytic agents for targeted treatment in the fight against bacterial diseases [1].
REFERENCE: [1] Obořilová R., Šimečková H., Pastucha M., Klimovič Š., Víšová I., Přibyl J., Vaisocherová-Lísalová H., Pantůček R., Skládal P., Mašlaňová I., Farka Z. Nanoscale. 2021, accepted manuscript. DOI: 10.1039/D1NR02921E
Living cells sense and respond to various signals through the generation of traction force. In this work, we show “top-down” fabrication of a highly ordered array of SiO2/Parylene C micropillars standing on the Si substrate that can be used as a probe to quantify the force generated from cells. We developed micropillars with an aspect ratio of 6 and 3.5 and spring constant of 4.7 µN µm-1 and 28 µN µm-1, respectively. The upper part of individual micropillars is coated with a 250 nm layer of SiO2 for covalent binding of adhesive proteins while micropillars’ side-walls remain non-adhesive. The results show biocompatibility of micropillar-based sub-strates and a dependence on its stiffness. Stiffer micropillars enhanced cell adhesion and proliferation rate, and a stronger cellular force of ~ 25µN was obtained. The signifi-cance of the cellular force transducer here is the fabrication of high aspect ratio pillars, elimination of the micro-contact printing of proteins, given that the SiO2 thin film allows the covalent binding of proteins or peptides. Next, pillars from Parylene C as the highly rigid material with a Young’s modulus of ~4 GPa can, compared to commonly used PDMS polymer, broaden the range of material stiff-ness to be probed by living cells. The last advantage is that our micropillars stand on Si wafer and thus any warping of underlying polymer membrane does not have to be con-sidered as it has been reported for low aspect ratio PDMS pillars.
Sensitive detection of clinically important analytes is critical for early disease diagnosis and initiation of treatment. Immunochemical assays are widely used to determine the concentration of analytes in complex samples due to the high specificity provided by antibodies. Lateral flow immunoassays (LFIAs) belong among the most popular forms of immunoassays. They are based on nitrocellulose membranes, and the most significant advantage of this approach is that there is no need for laboratory equipment to perform the assay. Most LFIAs are based on gold nanoparticles that allow naked-eye readout. However, the recent progress in nanotechnology provided various other nanomaterials that can be used as alternative labels.
Our work focused on photon-upconversion nanoparticles (UCNPs), which exhibit anti-Stokes luminescence, meaning they can be excited by the NIR laser and detected in the Vis region without optical background interference. In our proof-of-principle assay, we detected biotinylated bovine serum albumin using streptavidin-coated UCNPs; the readout was carried out using upconversion scanner UPCON-S Pro (Labrox, Finland). The testing of different nitrocellulose membranes revealed that FF120HP Plus Thick provides the highest signal-to-background ratio, making it the most suitable for the development of the final LFIA kit. We have also highlighted the necessity of suitable blocking conditions to prevent non-specific binding; the best results were achieved with 50% SuperBlock with 0,05% Tween 20. The signal-to-background ratio of the optimized assay reached up to 290, demonstrating the potential of UCNPs as labels in LFIAs.
This study aims to compare the parameters used to describe vesicular membranes obtained by fluorescence spectroscopy techniques. The most frequently mentioned parameters in the literature are the fluidity of the membrane core and the mobility of molecules in the outer layer of the membrane, which is expressed by the parameter Generalized Polarization. The 1,6-diphenyl-1,3,5-hexatriene (DPH) fluorescence probe is used to determine the membrane fluidity, in which the value of the fluorescence anisotropy parameter is inversely proportional to the membrane fluidity. The technique of fluorescence anisotropy can be used for each fluorescent probe - this value then expresses the degree of freedom of its rotation in a given microenvironment. Generalized polarization is then one of the possible interpretations of the degree of relaxation of the polar environment of the probe, which has a localized fluorophore near the interfacial interface. The probe used in this study is Laurdan. Data obtained using fluorescent probes are supplemented by results from DLS and ELS techniques.
In this work, model systems are vesicles formed from ion pairs of amphiphiles that contain different amounts of cholesterol. This work proposes the interpretation of the obtained parameters in the form of temperature vs. composition, where the obtained parameters are expressed by a color scale. These 2D maps are supplemented by various parameters, which come from the fitting of the dependences of the obtained parameters on temperature or composition.
ACKNOWLEDGEMENTS: This work was supported by the Czech Science Foundation, project No. 19-14024J (GACR), and Ministry of Science and Technology, Taiwan, project No. MOST108-2923-E006-MY3.
The usage of enantiomeric compounds in medicine and pharmacology is an important issue. The precise detection of enantiomeric excess is not a simple task: it is time-consuming, requires sophisticated equipment, and can be carried out only in a specialized lab. Because of that, research for a new (bio)analytical approaches and methods for simplification of enantiomers detection and recognition is an actual scientific challenge. The main goal of this work was to create the specific plasmonic nanostructures, as an alternative background for enantioselective detection. The plasmon-active nanoparticles were prepared with utilization or recently proposed template-assisted synthesis and analyzed by several techniques, allowing the investigation of their structure and properties. The next step created structures were implemented in the analysis of dopamine enantiomers concentration.
African Swine Fever Virus (ASFV) is a DNA virus of the Asfivirus genus of the Asfarviridae family that is found in blood, body fluids, and internal organs. ASFV was described more than 40 years ago. This virus spreads pandemically and the mortality rate of the virus-related disease ranges from 90 to 100 %. The aim of this study was to propose the detection of the specific nucleic acid of ASFV using electrochemical detection. Determination of DNA was conducted by chronopotentiometric stripping analysis (time of accumulation 120 s, stripping current 20, 10, 5, 1 µA, measurement time 240 s). The volume of the analyzed sample was 10 µL. Sufficient analysis duration (200‒600 s) is required to fully record the H peak signal of DNA at a given applied current. Extending the DNA analysis time at a given current significantly improved the dt/dE records of the H peak and the reproducibility of the analysis (RSD up to 10%). It was found that the suitable time at which the H peak is plotted reliably is around 240 s. The detected dsDNA concentration dependence (0 to 60 µg/mL) on the catalytic signal of the H peak was described according to the equation y = 260.4 + 366.7x, r = 0.961, RSD 23.1%. Subsequently, the SPION particle was modified with dsDNA. After the thermal release of DNA from SPION, DNA was immediately determined by chronopotentiometric stripping analysis method.
Soft foams with micro-macro porous hierarchical structure have been prepared by freeze-drying of solutions of polylactide (PLA, Ingeo 4043D, Mn 160.000 g.mol-1) and polycaprolactone (PCL, Aldrich, Mn 80.000 g.mol-1) mixtures in 1,4-dioxane with or without addition of sucrose porogens. The foams prepared by freeze-drying had open-porous aerogel characteristics. Pore sizes decreased with increasing the concentration of the polymers in solution in the range from several micrometers to several hundreds of micrometers. The hierarchical structure with bimodal micro-macro pore size distribution were obtained after addition of sucrose porogens with defined crystal size distributions to the solutions before freeze-drying and leaching the sucrose crystals from the freeze-dried foams with demineralised water. Polyethyleneimine was chemically conjugated to the alkali-treated foams followed by conjugation of citric acid using carbodiimide (EDC-NHS) chemistry. Finally, they were mineralised by immersing and incubating in a simulated body fluid with ionic concentration similar to that of human blood plasma, to obtain tissue engineering scaffolds. To verify their biocompatibility, the scaffolds were incubated with adipose-derived stem cells (ASC). Morphology of the cells attached to the scaffolds was evaluated and their viability was verified by a metabolic test.
ACKNOWLEDGEMENTS: This work was supported by the Czech health research council project NU20-08-00208 and by the Institutional Development Fund of FRIP provided by the Ministry of Agriculture of the Czech Republic.
Wet electrospinning is type of electrospinning technology where liquid is used instead of solid or other type of the collector to gather produced material. The experimental laboratory device for wet electrospinning will be described, parts for the device are mostly 3D printed by Fused Filament Fabrication (FFF). The different morphologies of biodegradable electrospun materials produced by wet electrospinning are compared with classic needle electrospinning by SEM pictures analysis. The comparison in morphology, degree of crystallinity measured by Differential Scanning calorimetry (DSC) and in-vitro viability, between samples produced by classic electrospinning; wet electrospun method dried in laboratory conditions and wet electrospun dried by freeze-drying will be introduced. The wet electrospinning with freeze-drying method brings high porosity and bulky samples, in-vitro viability tests shows the highest fibroblast proliferation of all the samples.
Osteoporosis is sometimes asymptomatic until the patient, often women over the age of 50, develops disastrous late-stage musculoskeletal complications, such as fractures [1]. In normal conditions, bone can regenerate and repair itself, but in this case, the regenerative process is compromised and requires support. The repair or replacements of the damaged or traumatized bone tissue can be aaccomplished by different grafting methods like autografts, allografts or bone substitutes [2]. Another promising technique for bone repair and regeneration is tissue engineering, which is based on: scaffolds, cells, and signalling molecules [3]. The aim of this study was to prepare and characterize composite beads as scaffolds for bone regeneration in case of osteoporotic fractures. The beads from sodium alginate, calcium phosphate tribasic and carboxymethyl guar gum nanoparticles (previously prepared [4], incorporated in order to be functionalized with drugs that sustain bone regeneration), were prepared using ionic crosslinking. The beads morphology (Scanning Electron Microscopy - EDX), their chemical structure (Fourier Transform Infrared Spectroscopy and X-Ray Diffraction) and their in vitro behaviour (retention of simulated body) indicates that the obtained beads can be considered for the proposed application.
ACKNOWLEDGMENTS: This work was supported by a grant of the Ministry of Research, Innovation and Digitization, CNCS/CCCDI - UEFISCDI, project number PN-III-P1-1.1-PD-2019-0937, within PNCDI III.
REFERENCES: [1] Liu J. et al. J Endocrinol Invest. 2019; 42(10):1149-1164.; [2] Kashte S. et al. Tissue Eng. Regen. Med. 2017; 14(1):1-14.; [3] Zhong Li et al. Handb Exp Pharmacol. 2021; 265:235-268.; [4] Mater. Sci. Eng. C 2016; 63:628-636.
Promising material solution, with a number of properties desirable for bone implants, is calcium doped titanium coating, which combines biogenic nature of calcium and good mechanical stability of biocompatible titanium matrix. In this work we present the results of material research focused on the effect of dopant concentration on the morphology of calcium doped titanium (TiCa) layer. TiCa layers were deposited on the Ti6Al4V substrates by dual electron beam evaporation technique. An analysis of the chemical composition and chemical bonds showed that the layers are composed of a calcium and titanium in their higher oxidation states i.e. partial formation of titanium and calcium oxides. Surface roughness and grain size increase with the concentration of Ca. Contact angle characterizing the wettability shows increasing trend, which follows the increase in surface roughness and concentration of calcium.
In this study, the potential of one scaffold nanocomposite three-phase of BCP/gelatin/bioglass45 s was in- vestigated. The use of composites and ceramics are suitable for bone regeneration. BCP included hydroxyapatite (HA) and tricalcium phosphate (β-TCP) for the first phase. The second phase used gelatin for the mechanical nature while the third phase 45S5 glass (SG) served the purpose of biocompatibility. Loading the drug vanco- mycin enabled the scaffold to be antibacterial in character and resist Gram-positive bacteria. Structural evolution during the synthesis of hydroxyapatite and 45S5 glass nanocomposite, in vitro and in vivo and loading of the vancomycin were investigated different spectroscopy methods. The theoretical one is the software response surface methodology. In best condition for concentration level 5 mg l−1 was the best loading and indicated 80 % efficiency. This work has practical and theoretical contributions to make. In this research, for the first time the practical formation of a three-phase bone scaffold included BCP for the mechanism of HA. SG has the ability to chemically and physically band with the bone and biocompatibility helped to change the carbon nanotubes, which is useful for future research.
One of the many interesting properties of silver nanoparticles is their antimicrobial activity. Since these properties are related to the morphology of the particles, it is interesting to pay attention to the antimicrobial effects of various metal nanostructures, which differ in the method of synthesis, size, or shape. One of the very interesting methods of synthesis of metal nanoparticles, which seems to be very advantageous because of its low cost, efficiency, and ecology, is the synthesis using plant extracts. For this work, woody parts of the Vitis vinifera were selected as material for the preparation of the extract. The canes of this plant are an agricultural waste containing many bioactive substances, which are not used for other significant purposes, making it a promising material for the preparation of metal nanoparticles with antimicrobial potential. The biosynthesized nanoparticles were detected and characterized using UV Vis spectrophotometry, transmission electron microscopy (TEM) and dynamic light scattering (DLS). TEM showed that the obtained nanoparticles have a heterogeneous shape with a size 4-51 nm. The DLS analysis supported that the Z-average was 113.8 nm and 0.165 PDI value. The use of biosynthesized nanoparticles showed an excellent growth inhibition property against two strains of the opportunistic pathogenic bacteria Pseudomonas aeruginosa (ATCC 10145 and ATCC 15442). Furthermore, a bactericidal concentration (BC) 40% v/v of AgNPs was found for P. aeruginosa ATCC 15442, while no BC was found in the range of tested concentrations for P. aeruginosa ATCC 10145.
The oxidative stress of bacteria when interacting with reactive oxygen species (ROS) can lead to further death of bacteria. This mechanism underlies the photodynamic therapy of bacterial infections to help fight against antibiotic resistance in bacteria. Nanostructured materials, in particular titanium dioxide (TiO2) nanoparticles, are an effective source of ROS generation. A significant drawback of Titania NPs is the wide band gap, and as a consequence, the generation of ROS is only exposed to UV radiation. The formation of hybrid nanostructures based on semiconductor quantum dots (QDs) and Titania NPs can overcome this problem. Due to the mutual arrangement of the conduction bands of QDs with titania NPs, effective photoinduced electron transfer from QDs to NPs can be realized in such structures. Thus, Titania NP/QD hybrid structures will combine the optical properties of QDs with the ability of NPs to efficiently generate ROS, resulting in bactericidal activity under visible radiation.
In this work, several approaches to the formation of hybrid structures were used, multilayer structures and nanocomposites of the core/shell type were formed. Multilayered hybrid structures were formed using the modified Langmuir-Blodgett technique, layers of structure components were sequentially deposited on a dielectric substrate. During the formation of core/shell nanocomposites, a Titania shell was grown on the QD core. The formation of hybrid structures directly at the stage of synthesis makes it possible to exclude all the negative factors associated with the presence of molecular shells on nanoparticles surface. In this work, we report the results of the comprehensive study of the photophysical and functional properties of hybrid nanostructures based on Titania nanoparticles and quantum dots and their activity against various bacterial strains. The efficiency of electron transfer was estimated by two independent methods from the donor site (from QDs luminescence) and the acceptor side (from the ROS generation by Titania). The antibacterial activity of the structures was assessed by the survival rate of bacteria after interacting with the structures under irradiation with visible radiation. Our comprehensive approach will make it possible to determine the factors limiting the effectiveness of the formed structures as antibacterial systems. Our results clearly demonstrate the high potential of the Titania NPs/QDs hybrid structures should exhibit a pronounced effect even against bacterial infections
A significant attention has been paid to polymer nanofibers, due to their beneficial properties such as a high specific surface area and high porosity. They can be used in various applications including separation processes, wound healing, tissue engineering etc. Besides others, the nanofiber-based materials have great ability to aerosol filtration and trapping microorganisms that has become extremely relevant duo to the COVID-19 occurrence. Nanofibrous membranes doped with an active antibacterial component directly in the mass of nanofibers are a next step towards products that actively eliminate microorganisms and thus protect human health.
In our study, nanofibers of polymers doped with ferrous sulphate (FeSO4) or quaternary ammonium salt (QAS) were prepared by electrospinning. Three types of polymers (polyvinylidenfluoride/PVDF, polylactic acid/PLA and polyurethane/PU) with good electrospinning processability and good mechanical properties of nanofibers were chosen. The prepared nanofibrous membranes were characterized in terms of materials by FTIR, the morphology of the fibers was assessed by SEM, and the pore size was determined by porometry. The leaching test showed a firm anchoring of the additive in the nanofiber structure. Antimicrobial activity was monitored after 0 h, 0.5 h, 4 h and 24 h using Staphylococcus aureus (CCM 4516) and Klebsiella pneumoniae (CCM 4415) strains.
The membrane prepared from PU doped with FeSO4 showed the best antibacterial efficiency. The porosity and morphology of the nanofibrous membrane effectively contribute to the trapping of microorganisms. This system was also evaluated as the most suitable from the electrospinning process effectivity point of view.
Despite benefits nanotherapeutics offer to healthcare strategies, there are still concerns about their biological safety. Kidney is responsible for elimination of toxic wastes from the blood, but nanoparticles independently of filtration can accumulate here, mostly in the glomerular mesangium. Magnetic nanoparticles (MNPs) are promising candidates for modern biomedicine. As prolonged residency of nanoparticles has been associated with the nephrotoxicity, we investigated the impact of MNPs on renal mesangial cells controlling glomerular hemodynamics and filtration process.
Primary murine glomerular mesangial cells were exposed to MNPs with two different coatings - polyethylene glycol (PEG) or bovine serum albumin (BSA).
PEGylated MNPs activated stronger inflammatory response compared to BSA coated MNPs. Strong induction of iNOS by PEG MNPs after 24 hours of exposure was in contrast to low-to-no induction of inflammatory factors (TNFa, IL-6, Mip2) induced by BSA MNPs. This was confirmed on mRNA as well as protein levels.
Both types of MNPs activated slight actin fiber remodeling associated with weak reduction in cell adhesion properties. Mesangial cells also displayed certain signs of damage after MNPs internalization, concerning mostly endoplasmic reticulum and mitochondria.
Taking together, the results indicate a crucial role of coating in MNPs-induced mesangial cell toxicity.
ACKNOWLEDGMENTS: This work was supported by the Slovak Research and Development Agency under the contract No. APVV-16-0579
This study was performed during the implementation of the project Buildingup Centre for advanced materials application of the Slovak Academy of Sciences, ITMS project code 313021T081 supported by Research & Innovation Operational Programme funded by the ERDF.
Modern medicine is now unimaginable without the use of various assistive materials and devices such as catheters, implants, and others, which come into contact with body fluids and tissues. In this context, the potential danger of bacterial colonization on the device surfaces and the subsequent formation of biofilms can cause serious infections and diseases. In an effort to prevent bacterial colonization, the smart self-activating antibacterial coating on a medically relevant surface was proposed in this work. The polypropylene film surface was grafted with a 3D periodic structure of Cu-based MOFs, which is capable of effectively catalyzing NO radicals from its endogenous donors and thus inhibit bacteria growth. NO is known for its antibacterial and anti-inflammatory properties and is not only effective in killing bacteria but also potent in the prevention of bacterial biofilms formation. Unlike materials requiring an external stimulus to activate its antibacterial properties, the proposed coating responds to an increase in the concentration of RSNOs associated with bacterial metabolites in body fluids and is therefore suitable for long-term use as well as for self-activation in bacteria presence.
Bioartificial heart valves and vessels grafts prepared from decellularized tissues could be recellularized with bone marrow-derived mesenchymal stem cells (MSCs). For this purposes we cultivated the stem cells on fibrin assemblies with immobilized growth factors. The cell differentiation will be facilitated by sustained release of growth factors
In our study assemblies based on fibrin, fibrin with heparin, fibrin with adsorbed or covalently-immobilized vascular endothelial growth factor A165 (VEGF) or fibroblast growth factor (FGF-2) via binding to heparin attached to fibrin have been prepared and evaluated for their stimulation of MSCs differentiation. We estimated the mRNA expression of endothelial marker CD31 (PECAM1), smooth muscle marker α-actin (ACTA2), osteoblast marker osteocalcin (BGLAP) and alkaline phosphatase (ALP). The gene expression was estimated using RT-PCR on days 1, 7 and 21 after seeding. The cell morphology and viability was evaluated by LIVE/DEAD staining. VEGF, both adsorbed and covalently bound, increased significantly the expression of smooth muscle marker α-actin. The mRNA expression of ACTA2 on day 7 and 21 raised more than 200 times in comparison to control samples (undifferentiated cells before seeding). Expression of the endothelial marker CD31 was low at the beginning. On day 21 the expressions of CD31 and ACTA2 were similar. The expression of osteocalcin was minimal. We conclude that MSCs on fibrin containing VEGF should be appropriate for the bioartificial heart valve replacements. ACKNOWLEDGEMENTS: Supported by the Czech Health Research Council, the Ministry of Health of the Czech Republic (grant No. NV19-02-00068).
Nanotechnology is a scientific field for the characterization, production, and application of nanoscale particles of 1-100 nm size. Due to their large surface area and small size, nanoparticles have different properties compared to bulk materials such as mechanical, electrical, magnetic, and chemical properties. They can be synthesized by physical, chemical and biological methods. The biosynthesis of nanoparticles is the synthesis of nanoparticles using microorganisms or plants with biomedical applications. This synthesis approach stands out as a cost-effective, environmentally friendly, bio-compatible, safe, and green approach.
The present study aims to synthesize silver nanoparticles by green chemistry using Trachystemon orientalis L. extract and to investigate anti-bacterial and anti-fungal effects properties.
UV-Visible absorption, fourier-transform infrared spectroscopy, and scanning electron microscopy analyses confirmed the synthesis. The results showed that silver nanoparticles have inhibitory effect on both gram-negative (E. coli, Klebsiella pneumonia and Pseudomonas aeruginosa) and gram-positive (Staphylococcus aureus and Enterococcus faecalis) bacterial strains. Furthermore, they showed anti-fungal activities against Candida utilis and Candida albicans.
Silver nanoparticles can affect the cell walls of the microorganism, changing the structure of cell membranes and even causing cell death. They can also increase the permeability of cell membranes, generate reactive oxygen species, and interrupt the replication of deoxyribonucleic acid.
In conclusion, green synthesis of silver nanoparticles using Trachystemon orientalis L. extract can be an effective method for biomedical purposes.
Nanoparticles are widely used in industry as well as biomedicine. An interesting group are aluminium oxide nanaoparticles (alumina nanoparticles) belonging to the family of metal oxide nanoparticles, which can be prepared by various methods (the sol-gel, precipitation, micro-emulsion and hydrothermal methods). In this work, alumina nanoparticles were prepared by biological synthesis using plant extract from fresh neem (Azadirachta indica) leaves. The particles were confirmed by the SEM analysis. The distribution diagram showed a size from 60 to 1,500, with a maximum around 200-400 nm. Alumina nanoparticles possess many potential applications in biomedicine such as drug delivery, therapy of cancer and other diseases, and immunotherapy.
Camomilla has been used for thousands of years in traditional medicine to treat different diseases. The dried flowers are one of the most popular single ingredients herbal teas that have potential health benefits. Common sources for lecithin extraction to subsequently form liposomes are soya bean, egg yolk and sunflower that are widely used in pharmaceutical, food and cosmetic industries.
The aim of this work is to encapsulate extract of Matricaria chamomilla that is rich in total phenolic content, antioxidants and shows inhibition against pathogenic strains. Various types of lecithin extracted from soya bean, egg yolk and sunflower have been used to evaluate encapsulation efficiency and long-term stability.
Four types of solvents were used for preparation of camomilla extracts - distilled water, 20 % ethanol, acetone and hexane. Polar solvents were added to dry herb Matricaria chamomilla at concentration 0,1 g dry weight/mL and the samples were incubated at 37 °C while shaking for 24 hours. After incubation, centrifugation was performed and supernatants were used for characterization. Oils were prepared by hexane extraction with automatic Soxtherm equipment. Subsequently extracts were characterized for their total phenolic and flavonoid content, antioxidant and antimicrobial activity against selected grampositive and gramnegative bacterias. Eventually ethanol extracts were encapsulated into various liposome particles and their size and stability was analysed.
Ethanol extract showed the highest amount of bioactive compounds, especially high total content of phenolic compounds. All liposome particles were stable and obtained high encapsulation efficiency with good ability to slowly release bioactive compounds. Prepared particles can be considered as safe and used in food and cosmetic industry.
ACKNOWLEDGEMENTS: This work was supported by the project Nr. FCH-S-21-7483 of the Faculty of Chemistry, Brno University of Technology.
Thermal treatments offer numerous opportunities in control of the shape, size, and purity of nanodiamonds (NDs) which is important for their technological use. This work brings a comparison between several thermal treatments of high pressure high temperature (HPHT) NDs, namely annealing in air, molten oxidizing salt (LiNO3) and solid non-oxidizing salt (NaCl) thermal treatments. The residual ND mass and the treatment time were fixed to 25 wt. %, and 30 min, respectively. The suitable temperatures to achieve these parameters were found experimentally. The annealing in air exhibits the highest temperature of processing (560°C) and the treatment with molten LiNO3 the lowest (430°C). Surprisingly, the presence of non-oxidizing NaCl in solid state also reduces the temperature to 490°C compared to the annealing in air. The obtained NDs were washed and dispersed in water and the obtained colloids were characterized by dynamic light scattering (DLS) and zeta potential measurements. Fourier transform infrared spectroscopy revealed minor changes among the samples, especially in the carbonyl signal position. Raman spectroscopy evidenced great suppression of the non-diamond sp2-C in all the samples. Transmission electron microscopy (TEM) was used to verify the shape of the treated NDs. The results point out that the use of salts in both cases generate rounded NDs, while the air oxidation preserve its originally irregular shape. This is supported by the thermal analysis using simultaneous differential scanning calorimetry, thermogravimetry, and mass spectrometry, which proved influence of NaCl on the acceleration of the combustion effect.
The dependence of biological activity of nanoparticles on their morphology is well known [1]. The key aspect is the number of {111} surface facets that exhibit the highest atomic density [2]. The nanoparticles containing the highest number of {111} facets are formed with decahedral or icosahedral shapes, described as 5 or 20 tetrahedra joined by twin boundaries. Because the tetrahedra do not fill the space completely, grain-boundaries or elastic strains are required to complete the nanoparticle shape [3]. The nanoparticle morphologies described above are known as multiply-twinned nanoparticles (MTPs) with fivefold symmetry axes.
In our study, we have synthesized nanoparticles with the spark erosion method [4]. We prepared a composite of silver nanoparticles embedded in a carbon black matrix to prevent the coalescence of the smallest nanoparticles. We have found that the resulting morphology contained nanoparticles of single crystalline, polycrystalline, and multiply-twinned shapes with sizes smaller than 50 nm. Complementarily to our experimental research, that provided evidence of MTPs with fivefold symmetry axes in our samples, we have used quantum-mechanical calculations to test stability of MPTs with other distributions of internal defects. Building upon our previously used methodology [5], we computed structural, electronic and elastic properties of grain boundaries and their junctions. Interestingly, the studied quadruple junction has been found close to limits of mechanical stability with respect to shearing processes.
ACKNOWLEDGEMENTS: The authors acknowledge the Czech Science Foundation for the financial support received under the Project No. 21-31852J. Computational resources were made available by the Ministry of Education, Youth and Sports of the Czech Republic under the Project of the IT4Innovations National Supercomputer Center (Project “e-Infrastructure CZ-LM2018140”) within the program Projects of Large Research, Development and Innovations Infrastructures and via the CESNET (Project No. LM2015042) and CERIT-Scientific Cloud (Project No. LM2015085).
REFERENCES: [1] Z. L. Wang (ed.), Characterization of Nanophase Materials, WILEY-VCH Verlag GmbH, Weinheim, 2000, ISBN 3-527-29837-1.; [2] M. Song et al., Colloids and Surfaces A: Physicochem. Eng. Aspects 408 (2012), 114.; [3] L. D. Marks and A. Howie, Multiply-twinned particles in silver catalysts, Nature 282 (1979), 196.; [4] Berkowitz A. E., Walter J. L., Spark erosion: A method for producing rapidly quenched fine powders, J. Mater. Res. 2 (1987), 277.; [5] S. Polsterová, M. Friák, M. Všianská, M. Šob: Quantum-Mechanical Assessment of the Energetics of Silver Decahedron Nanoparticles. Nanomaterials 10 (2020) 767.
This contribution is focused on the influence of liposomes composition on their stability during nebulization process by vibrating mesh nebulizer. Three different molecules (cholesterol, phosphatidic acid, and polyethylene glycol) were used for the stabilization of liposomes during nebulization. Physicochemical properties of liposomes before and after nebulization process were checked by dynamic and electrophoretic light scattering and membrane properties were measured by fluorescence spectroscopy - especially generalized polarization (Laurdan) and anisotropy (Diphenylhexatriene).
Cholesterol was capable of liposome stabilization because of increased membrane fluidity. The membrane properties of the outer and inner parts were not influenced by the nebulization process. Electrostatic stabilization was successful for the lowest concentration of phosphatidic acid, but after the nebulization process the hydration of the membrane outer part was changed. Higher amount of PEG needs to be added for successful stabilization of liposomes during nebulization. The nebulization process of the two lowest concentrations of PEG slightly influenced immobilized water and the rigidity of inner part of the membrane (especially around the phase transition temperature).
ACKNOWLEDGEMENTS: This work was supported by the University Specific Research, project No. FCH/FSI-J-21-7308.
Packaging plays important part of the visual communication and in consumer’s choice of purchasing goods. To enhance visual appearance, packaging material is often coated with some type of varnish or coating. Beside enhancement of visual appearance, additional coating often improves other packaging properties. The COVID-19 pandemic stressed the importance of the antimicrobial properties of goods that come in contact with consumers. During purchasing, consumer first comes into contact with packaging making it significant in the consumers’ protection. The aim of this research is to determine antimicrobial properties of nanocomposite coating which includes nanosized TiO2. For the purpose of the research a set of offset cardboard prints will be coated with nanocomposite coating composed of water based varnish and nanoscale TiO2 particles. The prepared samples will be characterized by determining CIE L*a*b* coordinates of primary colours, detecting colour fading in the accelerated ageing process and by determining inhibition of microorganisms’ growth by using XXX test.
It is expected that the research will enable determining influence of proposed nanocomposite coatings on the cardboard prints appearance, their resistance to accelerated ageing and effect of added nanoparticles on the antimicrobial behaviour of proposed coatings