Cold Atmospheric Pressure Plasma-Activated Medium Modulates Cellular Functions of Human Mesenchymal Stem/Stromal Cells In Vitro.

Cold atmospheric pressure plasma (CAP) offers a variety of therapeutic possibilities and induces the formation of reactive chemical species associated with oxidative stress. Mesenchymal stem/stromal cells (MSCs) play a central role in tissue regeneration, partly because of their antioxidant properties and ability to migrate into regenerating areas. During the therapeutic application, MSCs are directly exposed to the reactive species of CAP. Therefore, the investigation of CAP-induced effects on MSCs is essential. In this study, we quantified the amount of ROS due to the CAP activation of the culture medium. In addition, cell number, metabolic activity, stress signals, and migration were analyzed after the treatment of MSCs with a CAP-activated medium. CAP-activated media induced a significant increase in ROS but did not cause cytotoxic effects on MSCs when the treatment was singular and short-term (one day). This single treatment led to increased cell migration, an essential process in wound healing. In parallel, there was an increase in various cell stress proteins, indicating an adaptation to oxidative stress. Repeated treatments with the CAP-activated medium impaired the viability of the MSCs. The results shown here provide information on the influence of treatment frequency and intensity, which could be necessary for the therapeutic application of CAP.

The Impact of Ultrashort Pulse Laser Structuring of Metals on In-Vitro Cell Adhesion of Keratinocytes.

Besides the need for biomaterial surface modification to improve cellular attachment, laser-structuring is favorable for designing a new surface topography for external bone fixator pins or implants. The principle of this study was to observe how bioinspired (deer antler) laser-induced nano-microstructures influenced the adhesion and growth of skin cells. The goal was to create pins that allow the skin to attach to the biomaterial surface in a bacteria-proof manner. Therefore, typical fixator metals, steel, and titanium alloy were structured using ultrashort laser pulses, which resulted in periodical nano- and microstructures. Surface characteristics were investigated using a laser scanning microscope and static water contact angle measurements. In vitro studies with human HaCaT keratinocytes focused on cell adhesion, morphology, actin formation, and growth within 7 days. The study showed that surface functionalization influenced cell attachment, spreading, and proliferation. Micro-dimple clusters on polished bulk metals (DC20) will not hinder viability. Still, they will not promote the initial adhesion and spreading of HaCaTs. In contrast, additional nanostructuring with laser-induced periodic surface structures (LIPSS) promotes cell behavior. DC20 + LIPSS induced enhanced cell attachment with well-spread cell morphology. Thus, the bioinspired structures exhibited a benefit in initial cell adhesion. Laser surface functionalization opens up new possibilities for structuring, and is relevant to developing bioactive implants in regenerative medicine.

Discrimination between the effects of pulsed electrical stimulation and electrochemically conditioned medium on human osteoblasts.

BACKGROUND: Electrical stimulation is used for enhanced bone fracture healing. Electrochemical processes occur during the electrical stimulation at the electrodes and influence cellular reactions. Our approach aimed to distinguish between electrochemical and electric field effects on osteoblast-like MG-63 cells. We applied 20 Hz biphasic pulses via platinum electrodes for 2 h. The electrical stimulation of the cell culture medium and subsequent application to cells was compared to directly stimulated cells. The electric field distribution was predicted using a digital twin. RESULTS: Cyclic voltammetry and electrochemical impedance spectroscopy revealed partial electrolysis at the electrodes, which was confirmed by increased concentrations of hydrogen peroxide in the medium. While both direct stimulation and AC-conditioned medium decreased cell adhesion and spreading, only the direct stimulation enhanced the intracellular calcium ions and reactive oxygen species. CONCLUSION: The electrochemical by-product hydrogen peroxide is not the main contributor to the cellular effects of electrical stimulation. However, undesired effects like decreased adhesion are mediated through electrochemical products in stimulated medium. Detailed characterisation and monitoring of the stimulation set up and electrochemical reactions are necessary to find safe electrical stimulation protocols.

Response of Osteoblasts on Amine-Based Nanocoatings Correlates with the Amino Group Density.

Increased life expectancy in industrialized countries is causing an increased incidence of osteoporosis and the need for bioactive bone implants. The integration of implants can be improved physically, but mainly by chemical modifications of the material surface. It was recognized that amino-group-containing coatings improved cell attachment and intracellular signaling. The aim of this study was to determine the role of the amino group density in this positive cell behavior by developing controlled amino-rich nanolayers. This work used covalent grafting of polymer-based nanocoatings with different amino group densities. Titanium coated with the positively-charged trimethoxysilylpropyl modified poly(ethyleneimine) (Ti-TMS-PEI), which mostly improved cell area after 30 min, possessed the highest amino group density with an N/C of 32%. Interestingly, changes in adhesion-related genes on Ti-TMS-PEI could be seen after 4 h. The mRNA microarray data showed a premature transition of the MG-63 cells into the beginning differentiation phase after 24 h indicating Ti-TMS-PEI as a supportive factor for osseointegration. This amino-rich nanolayer also induced higher bovine serum albumin protein adsorption and caused the cells to migrate slower on the surface after a more extended period of cell settlement as an indication of a better surface anchorage. In conclusion, the cell spreading on amine-based nanocoatings correlated well with the amino group density (N/C).

Optimized Gingiva Cell Behavior on Dental Zirconia as a Result of Atmospheric Argon Plasma Activation.

Several physico-chemical modifications have been developed to improve cell contact with prosthetic oral implant surfaces. The activation with non-thermal plasmas was one option. Previous studies found that gingiva fibroblasts on laser-microstructured ceramics were hindered in their migration into cavities. However, after argon (Ar) plasma activation, the cells concentrated in and around the niches. The change in surface properties of zirconia and, subsequently, the effect on cell behavior is unclear. In this study, polished zirconia discs were activated by atmospheric pressure Ar plasma using the kINPen®09 jet for 1 min. Surfaces were characterized by scanning electron microscopy, X-ray photoelectron spectroscopy (XPS), and water contact angle. In vitro studies with human gingival fibroblasts (HGF-1) focused on spreading, actin cytoskeleton organization, and calcium ion signaling within 24 h. After Ar plasma activation, surfaces were more hydrophilic. XPS revealed decreased carbon and increased oxygen, zirconia, and yttrium content after Ar plasma. The Ar plasma activation boosted the spreading (2 h), and HGF-1 cells formed strong actin filaments with pronounced lamellipodia. Interestingly, the cells' calcium ion signaling was also promoted. Therefore, argon plasma activation of zirconia seems to be a valuable tool to bioactivate the surface for optimal surface occupation by cells and active cell signaling.

3D Spheroid Cultivation Alters the Extent and Progression of Osteogenic Differentiation of Mesenchymal Stem/Stromal Cells Compared to 2D Cultivation.

Mesenchymal stem/stromal cells (MSC) are capable of progenitor cell fraction renewal or tissue-specific differentiation. These properties are maintained during in vitro cultivation, making them an interesting model system for testing biological and pharmacological compounds. Cell cultivation in 2D is commonly used to study cellular responses, but the 2D environment does not reflect the structural situation of most cell types. Therefore, 3D culture systems have been developed to provide a more accurate physiological environment in terms of cell-cell interactions. Since knowledge about the effects of 3D culture on specific differentiation processes is limited, we studied the effects on osteogenic differentiation and the release of factors affecting bone metabolism for up to 35 days and compared them with the effects in 2D culture. We demonstrated that the selected 3D model allowed the rapid and reliable formation of spheroids that were stable over several weeks and both accelerated and enhanced osteogenic differentiation compared with the 2D culture. Thus, our experiments provide new insights into the effects of cell arrangement of MSC in 2D and 3D. However, due to the different culture dimensions, various detection methods had to be chosen, which in principle limits the explanatory power of the comparison between 2D and 3D cultures.

Pulsed Electrical Stimulation Affects Osteoblast Adhesion and Calcium Ion Signaling.

An extensive research field in regenerative medicine is electrical stimulation (ES) and its impact on tissue and cells. The mechanism of action of ES, particularly the role of electrical parameters like intensity, frequency, and duration of the electric field, is not yet fully understood. Human MG-63 osteoblasts were electrically stimulated for 10 min with a commercially available multi-channel system (IonOptix). We generated alternating current (AC) electrical fields with a voltage of 1 or 5 V and frequencies of 7.9 or 20 Hz, respectively. To exclude liquid-mediated effects, we characterized the AC-stimulated culture medium. AC stimulation did not change the medium's pH, temperature, and oxygen content. The H2O2 level was comparable with the unstimulated samples except at 5 V_7.9 Hz, where a significant increase in H2O2 was found within the first 30 min. Pulsed electrical stimulation was beneficial for the process of attachment and initial adhesion of suspended osteoblasts. At the same time, the intracellular Ca2+ level was enhanced and highest for 20 Hz stimulated cells with 1 and 5 V, respectively. In addition, increased Ca2+ mobilization after an additional trigger (ATP) was detected at these parameters. New knowledge was provided on why electrical stimulation contributes to cell activation in bone tissue regeneration.

A Comparative Study on the Adipogenic Differentiation of Mesenchymal Stem/Stromal Cells in 2D and 3D Culture.

Mesenchymal stem/stromal cells (MSC) are capable of renewing the progenitor cell fraction or differentiating in a tissue-specific manner. Adipogenic differentiation of adipose-tissue-derived MSC (adMSC) is important in various pathological processes. Adipocytes and their progenitors are metabolically active and secrete molecules (adipokines) that have both pro- and anti-inflammatory properties. Cell culturing in 2D is commonly used to study cellular responses, but the 2D environment does not reflect the structural situation for most cell types. Therefore, 3D culture systems have been developed to create an environment considered more physiological. Since knowledge about the effects of 3D cultivation on adipogenic differentiation is limited, we investigated its effects on adipogenic differentiation and adipokine release of adMSC (up to 28 days) and compared these with the effects in 2D. We demonstrated that cultivation conditions are crucial for cell behavior: in both 2D and 3D culture, adipogenic differentiation occurred only after specific stimulation. While the size and structure of adipogenically stimulated 3D spheroids remained stable during the experiment, the unstimulated spheroids showed signs of disintegration. Adipokine release was dependent on culture dimensionality; we found upregulated adiponectin and downregulated pro-inflammatory factors. Our findings are relevant for cell therapeutic applications of adMSC in complex, three-dimensionally arranged tissues.

Structure-based knowledge acquisition from electronic lab notebooks for research data provenance documentation.

BACKGROUND: Electronic Laboratory Notebooks (ELNs) are used to document experiments and investigations in the wet-lab. Protocols in ELNs contain a detailed description of the conducted steps including the necessary information to understand the procedure and the raised research data as well as to reproduce the research investigation. The purpose of this study is to investigate whether such ELN protocols can be used to create semantic documentation of the provenance of research data by the use of ontologies and linked data methodologies. METHODS: Based on an ELN protocol of a biomedical wet-lab experiment, a retrospective provenance model of the raised research data describing the details of the experiment in a machine-interpretable way is manually engineered. Furthermore, an automated approach for knowledge acquisition from ELN protocols is derived from these results. This structure-based approach exploits the structure in the experiment's description such as headings, tables, and links, to translate the ELN protocol into a semantic knowledge representation. To satisfy the Findable, Accessible, Interoperable, and Reuseable (FAIR) guiding principles, a ready-to-publish bundle is created that contains the research data together with their semantic documentation. RESULTS: While the manual modelling efforts serve as proof of concept by employing one protocol, the automated structure-based approach demonstrates the potential generalisation with seven ELN protocols. For each of those protocols, a ready-to-publish bundle is created and, by employing the SPARQL query language, it is illustrated that questions about the processes and the obtained research data can be answered. CONCLUSIONS: The semantic documentation of research data obtained from the ELN protocols allows for the representation of the retrospective provenance of research data in a machine-interpretable way. Research Object Crate (RO-Crate) bundles including these models enable researchers to easily share the research data including the corresponding documentation, but also to search and relate the experiment to each other.

Laser Structured Dental Zirconium for Soft Tissue Cell Occupation-Importance of Wettability Modulation.

Various approaches are being pursued to physico-chemically modify the zirconia neck region of dental implants to improve the integration into the surrounding soft tissue. In this study, polished zirconia discs were laser microstructured with periodic cavities and convex waves. These zirconia samples were additionally activated by argon plasma using the kINPen®09. The surface topography was characterized by scanning electron microscopy and the surface wettability by water contact angle. The in vitro study with human gingival fibroblasts (HGF-1) was focused on cell spreading, morphology, and actin cytoskeleton organization within the first 24 h. The laser-induced microstructures were originally hydrophobic (e.g., 60 µm cavities 138.4°), but after argon plasma activation, the surfaces switched to the hydrophilic state (60 µm cavities 13.7°). HGF-1 cells adhered flatly on the polished zirconia. Spreading is hampered on cavity structures, and cells avoid the holes. However, cells on laser-induced waves spread well. Interestingly, argon plasma activation for only 1 min promoted adhesion and spreading of HGF-1 cells even after 2 h cultivation. The cells crawl and grow into the depth of the cavities. Thus, a combination of both laser microstructuring and argon plasma activation of zirconia seems to be optimal for a strong gingival cell attachment.

The nanomorphology of cell surfaces of adhered osteoblasts.

The functionality of living cells is inherently linked to subunits with dimensions ranging from several micrometers down to the nanometer scale. The cell surface plays a particularly important role. Electric signaling, including information processing, takes place at the membrane, as well as adhesion and contact. For osteoblasts, adhesion and spreading are crucial processes with regard to bone implants. Here we present a comprehensive characterization of the 3D nanomorphology of living, as well as fixed, osteoblastic cells using scanning ion conductance microscopy (SICM), which is a nanoprobing method that largely avoids mechanical perturbations. Dynamic ruffles are observed, manifesting themselves in characteristic membrane protrusions. They contribute to the overall surface corrugation, which we systematically study by introducing the relative 3D excess area as a function of the projected adhesion area. A clear anticorrelation between the two parameters is found upon analysis of ca. 40 different cells on glass and on amine-covered surfaces. At the rim of lamellipodia, characteristic edge heights between 100 and 300 nm are observed. Power spectral densities of membrane fluctuations show frequency-dependent decay exponents with absolute values greater than 2 on living osteoblasts. We discuss the capability of apical membrane features and fluctuation dynamics in aiding the assessment of adhesion and migration properties on a single-cell basis.

The Anchorage of Bone Cells onto an Yttria-Stabilized Zirconia Surface with Mild Nano-Micro Curved Profiles.

The high biocompatibility, good mechanical properties, and perfect esthetics of ceramic dental materials motivate investigation into their suitability as an endosseous implant. Osseointegration at the interface between bone and implant surface, which is a criterion for dental implant success, is dependent on surface chemistry and topography. We found out earlier that osteoblasts on sharp-edged micro-topographies revealed an impaired cell phenotype and function and the cells attempted to phagocytize these spiky elevations in vitro. Therefore, micro-structured implants used in dental surgery should avoid any spiky topography on their surface. The sandblasted, acid-etched, and heat-treated yttria-stabilized zirconia (cer.face®14) surface was characterized by scanning electron microscopy and energy dispersive X-ray. In vitro studies with human MG-63 osteoblasts focused on cell attachment and intracellular stress level. The cer.face 14 surface featured a landscape with nano-micro hills that was most sinusoidal-shaped. The mildly curved profile proved to be a suitable material for cell anchorage. MG-63 cells on cer.face 14 showed a very low reactive oxygen species (ROS) generation similar to that on the extracellular matrix protein collagen I (Col). Intracellular adenosine triphosphate (ATP) levels were comparable to Col. Ceramic cer.face 14, with its sinusoidal-shaped surface structure, facilitates cell anchorage and prevents cell stress.

Enhancement of Intracellular Calcium Ion Mobilization by Moderately but Not Highly Positive Material Surface Charges.

Electrostatic forces at the cell interface affect the nature of cell adhesion and function; but there is still limited knowledge about the impact of positive or negative surface charges on cell-material interactions in regenerative medicine. Titanium surfaces with a variety of zeta potentials between -90 mV and +50 mV were generated by functionalizing them with amino polymers, extracellular matrix proteins/peptide motifs and polyelectrolyte multilayers. A significant enhancement of intracellular calcium mobilization was achieved on surfaces with a moderately positive (+1 to +10 mV) compared with a negative zeta potential (-90 to -3 mV). Dramatic losses of cell activity (membrane integrity, viability, proliferation, calcium mobilization) were observed on surfaces with a highly positive zeta potential (+50 mV). This systematic study indicates that cells do not prefer positive charges in general, merely moderately positive ones. The cell behavior of MG-63s could be correlated with the materials' zeta potential; but not with water contact angle or surface free energy. Our findings present new insights and provide an essential knowledge for future applications in dental and orthopedic surgery.

ROS Dependent Wnt/ß-Catenin Pathway and Its Regulation on Defined Micro-Pillars-A Combined In Vitro and In Silico Study.

The physico-chemical surface design of implants influences the surrounding cells. Osteoblasts on sharp-edged micro-topographies revealed an impaired cell phenotype, function and Ca2+ mobilization. The influence of edges and ridges on the Wnt/ß-catenin pathway in combination with the cells' stress response has not been clear. Therefore, MG-63 osteoblasts were studied on defined titanium-coated micro-pillars (5 × 5 × 5 µm) in vitro and in silico. MG-63s on micro-pillars indicated an activated state of the Wnt/ß-catenin pathway. The ß-catenin protein accumulated in the cytosol and translocated into the nucleus. Gene profiling indicated an antagonism mechanism of the transcriptional activity of ß-catenin due to an increased expression of inhibitors like ICAT (inhibitor of ß-catenin and transcription factor-4). Cells on pillars produced a significant reactive oxygen species (ROS) amount after 1 and 24 h. In silico analyses provided a detailed view on how transcriptional activity of Wnt signaling is coordinated in response to the oxidative stress induced by the micro-topography. Based on a coordinated expression of regulatory elements of the Wnt/ß-catenin pathway, MG-63s are able to cope with an increased accumulation of ß-catenin on micro-pillars and suppress an unintended target gene expression. Further, ß-catenin may be diverted into other signaling pathways to support defense mechanisms against ROS.

Femtosecond Laser Nano/Micro Textured Ti6Al4V Surfaces-Effect on Wetting and MG-63 Cell Adhesion.

Nano- and microstructured titanium surfaces have recently attracted attention in the field of regenerative medicine because of the influence which surface characteristics such as roughness and wettability can have on cellular processes. This study focuses on the correlation of surface properties (wettability and nano/micro texture) of laser-structured Ti6Al4V samples with pronounced cell adhesion. Samples were structured with multiple laser parameters in order to create a range of surface properties. Surface characterization was performed by contact angle measurements 1 and 7 days after laser processing. The arithmetic mean roughness of the material surface in an area (Sa) was determined by means of confocal laser scanning microscopy (CLSM). Immediately after wettability tests of the laser-structured surfaces, in vitro experiments with human MG-63 osteoblasts were carried out. For this purpose, the cell morphology and actin cytoskeleton organization were analyzed using CLSM and scanning electron microscopy. On rough microstructures with deep cavities, the cell growth and spreading were inhibited. An improved cellular adhesion and growth on nanostructured and sinusoidal microstructured surfaces could be demonstrated, regardless of hydrophilicity of the surfaces.

Plasma Polymerized Allylamine-The Unique Cell-Attractive Nanolayer for Dental Implant Materials.

Biomaterials should be bioactive in stimulating the surrounding tissue to accelerate the ingrowth of permanent implants. Chemical and topographical features of the biomaterial surface affect cell physiology at the interface. A frequently asked question is whether the chemistry or the topography dominates the cell-material interaction. Recently, we demonstrated that a plasma-chemical modification using allylamine as a precursor was able to boost not only cell attachment and cell migration, but also intracellular signaling in vital cells. This microwave plasma process generated a homogenous nanolayer with randomly distributed, positively charged amino groups. In contrast, the surface of the human osteoblast is negatively charged at -15 mV due to its hyaluronan coat. As a consequence, we assumed that positive charges at the material surface-provoking electrostatic interaction forces-are attractive for the first cell encounter. This plasma-chemical nanocoating can be used for several biomaterials in orthopedic and dental implantology like titanium, titanium alloys, calcium phosphate scaffolds, and polylactide fiber meshes produced by electrospinning. In this regard, we wanted to ascertain whether plasma polymerized allylamine (PPAAm) is also suitable for increasing the attractiveness of a ceramic surface for dental implants using Yttria-stabilized tetragonal zirconia.

Terminal chemical functions of polyamidoamine dendrimer surfaces and its impact on bone cell growth.

Besides their use for drug and gene delivery, dendrimer molecules are also favorable for the design of new surface coatings for orthopedic and dental implants due to the wide variety of functional terminal groups and their multivalent character. The purpose of this work was to observe how covalently immobilized polyamidoamine (PAMAM) dendrimer molecules with different terminal chemical groups influenced serum protein adsorption and osteoblast behavior. To this end, fifth-generation PAMAM dendrimers were immobilized on silicon surfaces with an anhydride-containing silane coupling agent which results in positively charged terminal NH2-groups. Coatings with a net negative charge were generated by introduction of terminal CO2H- or CH3-groups. Surface characterization was performed by static and dynamic contact angle and zeta potential. The in vitro studies with human MG-63 osteoblastic cells focused on cell adhesion, morphology, cell cycle, apoptosis and actin formation within 24 h. This work demonstrated that cell growth was dependent on surface chemistry and correlated strongly with the surface free energy and charge of the material. The positively charged NH2 surface induced tight cell attachment with well-organized actin stress fibers and a well spread morphology. In contrast, CO2H- and CH3-functional groups provoked a decrease in cell adhesion and spreading and indicated higher apoptotic potential, although both were hydrophilic. The knowledge about the cell-material dialogue is of relevance for the development of bioactive implants in regenerative medicine.

Phenotypic stability of the human MG-63 osteoblastic cell line at different passages.

One of the most popular cell lines in osteogenesis studies is the human osteoblastic line MG-63. For cell biological investigation, it is important that the cells remain stable in their phenotype over several passages in cell culture. MG-63 cells can be used to provide fundamental insights into cell--material interaction. The aim of this study is to present a systematic characterization of the physiological behavior of MG-63 cells in the range of passages 5-30. Significant cell physiology processes during the first 24 h, including cell morphology, availability of adhesion receptors, cell cycle phases, as well as the expression of the signaling proteins Akt, GSK3a/b, IkB-α, ERK1/2, p38-MAPK, and intracellular calcium ion mobilization, remained stable over the entire range of passages P5-P30. Due to these stable characteristics in a wide range of cell culture passages, MG-63 cells can be considered as a suitable in vitro model to analyze the biocompatibility and biofunctionality of implant materials.

Enhanced calcium ion mobilization in osteoblasts on amino group containing plasma polymer nanolayer.

BACKGROUND: Biomaterial modifications-chemical and topographical-are of particular importance for the integration of materials in biosystems. Cells are known to sense these biomaterial characteristics, but it has remained unclear which physiological processes bio modifications trigger. Hence, the question arises of whether the dynamic of intracellular calcium ions is important for the characterization of the cell-material interaction. In our prior research we could demonstrate that a defined geometrical surface topography affects the cell physiology; this was finally detectable in a reduced intracellular calcium mobilization after the addition of adenosine triphosphate (ATP). RESULTS: This new contribution examines the cell physiology of human osteoblasts concerning the relative cell viability and the calcium ion dynamic on different chemical modifications of silicon-titanium (Ti) substrates. Chemical modifications comprising the coating of Ti surfaces with a plasma polymerized allylamine (PPAAm)-layer or with a thin layer of collagen type-I were compared with a bare Ti substrate as well as tissue culture plastic. For this purpose, the human osteoblasts (MG-63 and primary osteoblasts) were seeded onto the surfaces for 24 h. The relative cell viability was determined by colorimetric measurements of the cell metabolism and relativized to the density of cells quantified using crystal violet staining. The calcium ion dynamic of osteoblasts was evaluated by the calcium imaging analysis of fluo-3 stained vital cells using a confocal laser scanning microscope. The positively charged nano PPAAm-layer resulted in enhanced intracellular calcium ion mobilization after ATP-stimulus and cell viability. This study underlines the importance of the calcium signaling for the manifestation of the cell physiology. CONCLUSIONS: Our current work provides new insights into the intracellular calcium dynamic caused by diverse chemical surface compositions. The calcium ion dynamic appears to be a sensitive parameter for the cell physiology and, thus, may represent a useful approach for evaluating a new biomaterial. In this regard, reliable in vitro-tests of cell behavior at the interface to a material are crucial steps in securing the success of a new biomaterial in medicine.

Restricted cell functions on micropillars are alleviated by surface-nanocoating with amino groups.

The topographical and chemical surface features of biomaterials are sensed by the cells, affecting their physiology at the interface. When placed on titanium, we recently discovered osteoblasts attempted caveolae-mediated phagocytosis of the sharp-edged microstructures. This active, energy-consuming process resulted in decreased osteoblastic cell functions (e.g. secretion of extracellular matrix proteins). However, chemical modification with plasma polymerized allylamine (PPAAm) was able to amplify osteoblast adhesion and spreading, resulting in better implant osseointegration in vivo In the present in vitro study, we analyzed whether this plasma polymer nanocoating is able to attenuate the microtopography-induced changes of osteoblast physiology. On PPAAm, we found cells showed a higher cell interaction with the geometrical micropillars by 30 min, and a less distinct reduction in the mRNA expression of collagen type I, osteocalcin and fibronectin after 24 h of cell growth. Interestingly, the cells were more active and sensitive on PPAAm-coated micropillars, and react with a substantial Ca2+ ion mobilization after stimulation with ATP. These results highlight that it is important for osteoblasts to establish cell surface contact for them to perform their functions.