Automatic Actin Filament Quantification of Osteoblasts and Their Morphometric Analysis on Microtextured Silicon-Titanium Arrays

Abstract

Microtexturing of implant surfaces is of major relevance in the endeavor to improve biorelevant implant designs. In order to elucidate the role of biomaterial’s topography on cell physiology, obtaining quantitative correlations between cellular behavior and distinct microarchitectural properties is in great demand. Until now, the microscopically observed reorganization of the cytoskeleton on structured biomaterials has been difficult to convert into data. We used geometrically microtextured silicon-titanium arrays as a model system. Samples were prepared by deep reactive-ion etching of silicon wafers, resulting in rectangular grooves (width and height: 2 µm) and cubic pillars (pillar dimensions: 2 × 2 × 5 and 5 × 5 × 5 µm); finally sputter-coated with 100 nm titanium. We focused on the morphometric analysis of MG-63 osteoblasts, including a quantification of the actin cytoskeleton. By means of our novel software FilaQuant, especially developed for automatic actin filament recognition, we were first able to quantify the alterations of the actin network dependent on the microtexture of a material surface. The cells’ actin fibers were significantly reduced in length on the pillared surfaces versus the grooved array (4–5 fold) and completely reorganized on the micropillars, but without altering the orientation of cells. Our morpho-functional approach opens new possibilities for the data correlation of cell-material interactions.

Osteoblasts with impaired spreading capacity benefit from the positive charges of plasma polymerised allylamine

“…Taking the unaltered level of the number and length of actin filaments for Ti and TiPPAAm into account, these results indicate that the enhancement of cell area elicited by PPAAm is carried out by cells without a distinct increase in the number or length of actin filaments. This assumption can be affirmed by in vitro data from (Matschegewski et al, 2010). Herein, the authors describe the behaviour of MG-63 osteoblasts on defined micropillar-structured surfaces, where cell orientation and spreading occurred despite the complete lack of organised actin fibres, but cells showed short filamentous actin solely on top of the pillars similar to what we see in the presence of CD.…”

“…The microscopic images were processed in three steps according to (Matschegewski et al, 2012): pre-processing, feature detection and quantification. Before feature detection, the main sources of errors were reduced, namely noise, by variational methods (Birkholz, 2011b) , 2008).…”

Cell-material interaction

“…For example the lateral spacing of the nanotube system was varied by altering the tube diameter between 30 and 100 nm. On such topographies, cells alter not only the phenotype but also their internal architecture, e.g., the actin cytoskeleton, in a surface geometrydependent manner [14,15] (Figure 1).…”

Osteogenic activity of titanium surfaces with hierarchical micro/nano-structures obtained by hydrofluoric acid treatment

“…16,20,21 Micropits and micropillars are typically arranged in an organized manner (isotropic) rather than an unorganized manner (anisotropic) on the microgrooves. 22 It has been reported that the microscopic bone properties of cancellous and cortical bone enable them to be more suitable for use with the anisotropic surface for interlocking. 23 Nanoleaves may provide a larger contact area for adherent cells compared to nanoplates; however, the production process is complicated.…”