Figure 7

Effects of titania nanosurfaces on adhesion behavior of macrophages Representative confocal laser microscopic images of (A) F-actin (red), nuclear (blue), and paxillin (green) staining and (B) the intensity of paxillin-labeled signals in J774A.1 cells cultured on a polystyrene culture plate or smooth (SM), micro-roughened (MR), or nano-roughened (NR) 1 or 2 surfaces on day 1. (C) Gene expression of paxillin (Pxn) relative to glyceraldehyde 3-phosphate dehydrogenase (GAPDH), as analyzed by reverse transcription-polymerase chain reaction (RT-PCR), in the cells under the corresponding culture conditions on days 1 or 3. (D) Representative scanning electron microscopy (SEM) images and the area ratio of shining spots per unit area within the peripheral region of the cell body in the cells cultured on each surface on day 1. (E) Scatter plots indicating a positive linear relationship between the area ratio of shining spots per unit area or the intensity of paxillin-labeled signals and the spike density or (F) the gene expression levels of toll-like receptor (TLR) 4 and TLR2 relative to GAPDH, as analyzed by RT-PCR, in macrophages on the MR, NR1, and NR2 surfaces. Data represented as box plots (N = 15–52 in B) or mean ± standard deviation (SD; N = 3 in C and N = 5–8 in D). Different letters indicate statistically significant differences (P < 0.05; Tukey’s honestly significant difference [HSD] test in B, Bonferroni multiple comparisons test in C, and Games–Howell test in D). CTCF, Corrected total cell fluorescence. Note (A) the strong paxillin signals in macrophages on the NR1 or NR2 titania surfaces (double arrows) in contrast with fewer signals on the other surfaces (arrows), and (D) the shining spots (yellow arrows) corresponding to the vertices of surface spikes digging into the peripheral parts of the macrophage cell body (yellow dashed rectangular within the white dashed circle) are increased on the NR1 and NR2 titania surfaces compared to that on the MR titanium surface.