Figure 5: The blood clot matrix is composed of fibrin fibres, which colocalize to fibronectin, whereas newly assembled matrix is made of fibronectin only.
(a) Immunofluorescent micrograph of blood and subsequent seeded fibroblasts on alkali-treated Ti surface represented as maximum intensity projections of a confocal z-stack (original 9.8 μm z-thickness) shown separate for fibrin, Fn, nuclei (DAPI) and as merged image. (b) Higher magnification image of a region of interest specified in (a) (merged image, white rectangle) showing merged maximum intensity projection of a substack of the original z-stack of 2 μm thickness. (c) Maximum intensity projection image as in (b) with Fn-Fibrin colocalized voxels depicted in white. (d) Quantification of Fn fraction delocalized with fibrin, which represents the remaining green voxels depicted in (c). Calculated fractions are shown as datapoints (left) and boxplots (right) for all conditions. Boxplots represent mean (middle square), median (central line), 25th to 75th percentile (box) and standard deviation (whisker) of 7 fields of view of duplicate Ti surfaces per condition repeated for 5 donors. Statistically significant differences between blood and blood & fibroblasts conditions are indicated by (***) for p < 0.001. Fibroblasts were shown to produce new Fn ECM and therefore significantly increase Fn fibrils delocalized with fibrin.
