Adrian Chlanda

Quantitative imaging of electrospun fibers by PeakForce Quantitative NanoMechanics atomic force microscopy using etched scanning probes

Electrospun polymeric submicron and nanofibers can be used as tissue engineering scaffolds in regenerative medicine. In physiological conditions fibers are subjected to stresses and strains from the surrounding biological environment. Such stresses can cause permanent deformation or even failure to their structure. Therefore, there is a growing necessity to characterize their mechanical properties, especially at the nanoscale. Atomic force microscopy is a powerful tool for the visualization and probing of selected mechanical properties of materials in biomedical sciences. Image resolution of atomic force microscopy techniques depends on the equipment quality and shape of the scanning probe. The probe radius and aspect ratio has huge impact on the quality of measurement. In the presented work the nanomechanical properties of four different polymer based electrospun fibers were tested using PeakForce Quantitative NanoMechanics atomic force microscopy, with standard and modified scanning probes. Standard, commercially available probes have been modified by etching using focused ion beam (FIB). Results have shown that modified probes can be used for mechanical properties mapping of biomaterial in the nanoscale, and generate nanomechanical information where conventional tips fail.

Influence of biodegradable polymer coatings on corrosion, cytocompatibility and cell functionality of Mg-2.0Zn-0.98Mn magnesium alloy.

Four kinds of biodegradable polymers were employed to prepare bioresorbable coatings on Mg-2.0Zn-0.98Mn (ZM21) alloy to understand the relationship between polymer characteristics, protective effects on substrate corrosion, cytocompatibility and cell functionality. Poly-l-lactide (PLLA), poly(3-hydroxybutyrate) (PHB), poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) or poly(lactic-co-glycolic) acid (PLGA) was spin-coated on ZM21, obtaining a smooth, non-porous coating less than 0.5μm in thickness. Polymer coating characterization, a degradation study, and biocompatibility evaluations were performed. After 4 w of immersion into cell culture medium, degradation of PLGA and PLLA coatings were confirmed by ATR-FTIR observation. The coatings of PLLA, PHB and PHBV, which have lower water permeability and slower degradation than PLGA, provide better suppression of initial ZM21 degradation and faster promotion of human osteosarcoma cell growth and differentiation.

Biodegradable fiducial markers for X-ray imaging – soft tissue integration and biocompatibility

This study aims at the development of materials for biodegradable fiducial markers for X-ray based medical imaging and their anchorage in soft tissue. Towards this goal a degradable polymer matrix of poly(l-lactide-co-ε-caprolactone) (P[LAcoCL]) was combined with barium sulfate (BaSO4) and hydroxyapatite (HAp) as radio-opaque fillers. Low pressure plasma treatment was applied to the composite materials to improve cell adhesion and subsequent tissue integration. In particular, the effects of oxygen and ammonia plasmas were evaluated and compared using X-ray photoelectron spectroscopy, atomic force microscopy and dynamic water contact angle measurements as well as in vitro studies using the murine fibroblast cell line L929. To exclude the cytotoxic effects of degradation products of P[LAcoCL] and released BaSO4 or HAp cytotoxicity assays with the degradation products of the composite materials were conducted. The results obtained by this broad range of analytical techniques suggest the application of composites of P[LAcoCL] with BaSO4 and HAp as promising material systems for innovative fiducial markers for soft tissue in X-ray based medical imaging.

Microstructure and nanomechanical properties of single stalks from diatom Didymosphenia geminata and their change due to adsorption of selected metal ions

We present topographical and nanomechanical characterization of single Didymosphenia geminata stalk. We compared the samples before and after adsorption of metal ions from freshwater samples. Transmission electron microscopy studies of single stalk cross‐sections have shown three distinct layers and an additional thin extra coat on the external layer (called “EL”). Using scanning electron microscopy and atomic force microscopy (AFM), we found that topography of single stalks after ionic adsorption differed significantly from topography of pristine stalks. AFM nanoindentation studies in ambient conditions yielded elastic moduli of 214 ± 170 MPa for pristine stalks and 294 ± 108 MPa for stalks after ionic adsorption. Statistical tests showed that those results were significantly different. We conducted only preliminary comparisons between ionic adsorption of several stalks in air and in water. While the stalks with ions were on average stiffer than the pristine stalks in air, they became more compliant than the pristine stalks in water. We also heated the stalks and detected EL softening at 50°C ± 15°C. AFM nanoindentation in air on the softened samples yielded elastic moduli of 26 ± 9 MPa for pristine samples and 43 ± 22 MPa for stalks with absorbed metal ions. Substantial decrease of the EL elastic moduli after heating was expected. Significantly different elastic moduli for the samples after ionic adsorption in both cases (i.e., for heated and nonheated samples), as well as behavior of the stalks immersed in water, point to permanent structural EL changes due to ions.

The influence of carbon-encapsulated iron nanoparticles on elastic modulus of living human mesenchymal stem cells examined by atomic force microscopy

Nanomaterials and nanoparticles are regarded as promising candidates for various biomedical applications due to their unique physicochemical properties. In this study, three types of carbon-encapsulated iron nanoparticles (CEINs) were synthesized and their impact on cellular changes was investigated by atomic force microscopy (AFM). The AFM experiment was additionally compared with conventional methods, such as colorimetric assay and other microscopic techniques. A significant difference of reduced Youngs modulus of the cells was revealed, even at low concentration of nanoparticles in the culture medium. The AFM measurement proved to be a useful tool not only for visualization, but also for identification of local cellular changes at the nanoscale after exposure to carbon-encapsulated iron nanoparticles.

Structure and physico-mechanical properties of low temperature plasma treated electrospun nanofibrous scaffolds examined with atomic force microscopy

Electrospun nanofibrous scaffolds are willingly used in tissue engineering applications due to their tunable mechanical, chemical and physical properties. Additionally, their complex openworked architecture is similar to the native extracellular matrix of living tissue. After implantation such scaffolds should provide sufficient mechanical support for cells. Moreover, it is of crucial importance to ensure sterility and hydrophilicity of the scaffold. For this purpose, a low temperature surface plasma treatment can be applied. In this paper, we report physico-mechanical evaluation of stiffness and adhesive properties of electrospun mats after their exposition to low temperature plasma. Complex morphological and mechanical studies performed with an atomic force microscope were followed by scanning electron microscope imaging and a wettability assessment. The results suggest that plasma treatment can be a useful method for the modification of the surface of polymeric scaffolds in a desirable manner. Plasma treatment improves wettability of the polymeric mats without changing their morphology.

Microscopic Methods for Characterization of Selected Surface Properties of Biodegradable, Nanofibrous Tissue Engineering Scaffolds

Biodegradable polymeric fibers with nanoand submicron diameters, produced by electrospinning can be used as scaffolds in tissue engineering. It is necessary to characterize their mechanical properties especially at the nanoscale. The Force Spectroscopy is suitable atomic force microscopy mode, which allows to probe mechanical properties of the material, such as: reduced Youngs modulus, deformation, adhesion, and dissipation. If combined with standard operating mode: contact or semicontact, it will also provide advanced topographical analysis. In this paper we are presenting results of Force Spectroscopy characterization of two kinds of electrospun fibers: polycaprolactone and polycaprolactone with hydroxyapatite addition. The average calculated from Johnson-Kendall-Roberts theory Youngs modulus was 4 ± 1 MPa for pure polymer mesh and 20 ± 3 MPa for composite mesh.

Fabrication, multi-scale characterization and in-vitro evaluation of porous hybrid bioactive glass polymer-coated scaffolds for bone tissue engineering

Bioactive glass-based scaffolds are commonly used in bone tissue engineering due to their biocompatibility, mechanical strength and adequate porous structure. However, their hydrophobicity and brittleness limits their practical application. In this study, to improve nanomechanical properties of such scaffolds, pure bioactive hybrid glass and two bioactive hybrid glass-polymer coated composites were fabricated. A complementary micro and nanoscale characterization techniques (SEM, AFM, μCT, FTIR, compressive test, goniometer) were implemented for detailed description of architecture and physicochemical properties of hybrid bioactive glass-based scaffolds with emphasis on nano-mechanics. The final step was in-vitro evaluation of three dimensional macroporous structures. Our findings show that after polymer addition, architecture, topography and surface properties of the scaffolds were changed and promoted favoured behaviour of the cells.

Multi-scale characterization and biological evaluation of composite surface layers produced under glow discharge conditions on NiTi shape memory alloy for potential cardiological application

NiTi shape memory alloys are characterized by relatively good biocompatibility primarily thanks to their ability to self-passivate. However, before they can be used as medical implants for long term use, they need to undergo treatment aimed at producing layers on their surface that are superior to spontaneously formed oxide layers and that would increase their resistance to corrosion, limit nickel ion release from the surface (metallosis) and have the capability to shape their biological properties depending on the application. Furthermore, cardiac implants require addressing the issue of blood clotting on the surface. Treatment in glow-discharge low temperature plasma makes it possible to produce titanium layers with a structure and properties that are controlled via process parameters. In addition, antithrombogenic properties can be improved by depositing a carbon coating via the RFCVD process. The aim of the study was to investigate the structure, surface topography, adhesive properties, wettability, surface free energy and evaluate metallosis after producing TiO2 and a-C:N:H + TiO2 composite layers on NiTi alloy. The capabilities of AFM microscopes in studying the adhesive properties of a surface were also highlighted in the study. The study shows that the produced surface layers are capable of significantly reducing metallosis. Furthermore, in contrast to NiTi in its initial state, layers of nanocrystalline TiO2 titanium oxide (rutile) with a homogeneous structure demonstrate greater adhesion strength and more developed surface in the microscale, which facilitates the formation of an a-C:N:H coating. Therefore the formation of a coating of a-C:N:H amorphous carbon on NiTi alloy that has previously been oxidised in low-temperature plasma may prove to be a favourable solution in terms of using NiTi alloy to produce cardiac implants.

Micro and nanoscale characterization of poly(DL-lactic-co-glycolic acid) films subjected to the L929 cells and the cyclic mechanical load

In this paper, the effect of the presence of L929 fibroblast cells and a cyclic load application on the kinetics of the degradation of amorphous PLGA films was examined. Complex micro and nano morphological, mechanical and physico-chemical studies were performed to assess the degradation of the tested material. For this purpose, molecular weight, glass transition temperature, specimen morphology (SEM, μCT) and topography (AFM) as well as the stiffness of the material were measured. The study showed that the presence of living cells along with a mechanical load accelerates the PLGA degradation in comparison to the degradation occurring in acellular media: PBS and DMEM. The drop in molecular weight observed was accompanied by a distinct increase in the tensile modulus and surface roughness, especially in the case of the film degradation in the presence of cells. The suspected cause of the rise in stiffness during the degradation of PLGA films is a reduction in the molecular mobility of the distinctive superficial layer resulting from severe structural changes caused by the surface degradation. In conclusion, all the micro and nanoscale properties of amorphous PLGA considered in the study are sensitive to the presence of L929 cells, as well as to a cyclic load applied during the degradation process.