## 3 Results
### Actin Cytoskeleton and Quantification of Filaments  
We focused on the quantification of the cellular actin filament organization on the three differently charged surfaces: Ti (\(-\)), PPAAm (\(+\)), and PDADMA (\(++\)). Confocal microscopy was applied to visualize the actin cytoskeleton after 1 and 24 h and the quantification was carried out by the software FilaQuant (Figure 4).  
When regarding actin distribution after 1 h, considerable rearrangements were revealed, in particular on PPAAm. Here, the cells are extremely well developed with numerous, long, filopodial protrusions in comparison with uncoated Ti. Compared with PPAAm, filopodia on PDADMA surfaces are considerably shorter. In detail, the resulting data quantification via FilaQuant revealed a significantly shorter mean filament length on PDADMA (4.3 \(\upmu\)m, compared with 6.2 and 6.6 \(\upmu\)m on Ti and PPAAm, respectively), whereas the total filament length on PPAAm surfaces (1050.0 \(\upmu\)m) tripled as a result of the threefold increase in the actin filament number (148.3) after 1 h in contrast to Ti (filament number: 62.3, total filament length: 386.5 \(\upmu\)m) and PDADMA (filament number: 51.4, total filament length: 241.6 \(\upmu\)m) (Figure 5, Table 2). The advantage of this moderately positively charged surface for cell occupation is clearly evident.  
Figure 3: High-resolution SEM images showing the MG-63 cell morphology after 1 h on plasma polymer (PPAAm)-coated Ti. Note the multiple elongated filopodia protrusions (arrow) in the cells’ growth zone in order to probe the environment and facilitating cell adhesion (magnification from left to right 2000 and 5000\(\times\), scale bars 2 \(\upmu\)m; FE-SEM Merlin VP compact, Carl Zeiss).  
After 24 h, actin is organized typically as a network of well-defined stress fibers, spanning the entire cell body on negatively charged Ti and moderately positively charged PPAAm surfaces (Figure 4). On highly positively charged PDADMA surfaces, the actin formation is still confined to the outer cell edge and is presented in a clustered form. Here, the cells could neither spread nor form stress fibers, remaining round in shape, and having also shrunk in size. The total filament length shortened by about 6 times with respect to cells on Ti and PPAAm (Figure 5, Table 2). Similarly, the mean and maximum filament length decreased significantly by approximately 4.5 and 6 times, respectively.  
Figure 4: Actin cytoskeleton in MG-63s after 1 h (left) and 24 h (right) on Ti (negative surface charge), PPAAm (moderately positive surface charge), and PDADMA (highly positive surface charge). Shown are confocal microscopic images as a precondition for the actin quantification (bars = 10 μm; LSM780, [PERSON]) and the resulting FilaQuant filament curve graphs of the actin filaments and stress fibers. Note the multiple elongated actin filament protrusions in cells’ filopodia after 1 h on PPAAm in contrast to shorter actin filament extensions on PDADMA. After 24 h, cells on Ti and PPAAm are equally well spread with long actin filaments throughout the cell. Cells on PDADMA have shrunk and remain round shaped. The actin filaments are short and clustered toward the leading edge.