## 3 Results and Discussion
Given the prominence of the object, its considerable size and sharp features, it is most likely that it rests on the surface of the cell, which may not be able to uptake this foreign object considering that it is only 5-10 times larger than it. Figure 2: Representative optical fluorescent micrograph of control hMSCs, not subjected to interaction with the nanoparticles. Cells were stained for nucleus using DAPI (blue) and for f-actin cytoskeletal microfilaments using phalloidin (red). Scale bar equals 50 \(\upmu\)m.  
Figure 4, however, shows two interesting scenarios derived from optical fluorescent imaging. In both cases, a somewhat smaller, but equally irregularly shaped HAp nanoparticle cluster as that in Figure 3a coincides with the cell nucleus. In one case, however, it does not directly interfere with the nucleus (Figure 4a), while in the other case (Figure 4b) it obviously indents the nucleus due to its localization in the direct vicinity of it. This is to say that very large objects do get internalized by the cells, without necessarily getting molded to softer, more curved geometries during the passage through the cytoplasm towards different posts in the cell. Nanoparticle agglomerates can preserve their sharp outlines and this will be an important premise for the analysis that will be reported in Section 3.2. Figure 4: Fluorescent optical images of stained hMSCs and HAp nanoparticles. Both cells in the foci of images (**a**,**b**) display the uptake of finer nanoparticle aggregates, but also a single large agglomerate on the edge of the cell nucleus. Arrow in (**a**) points at the intact cell nucleus area overlapping with the nanoparticle agglomerate signal, while the arrow in (**b**) points at the indented cell  
Figure 3: Magnified portions of Figure 1 (**a**–**d**), along with an SEM image of HAp nanoparticles (**e**), confirming that what is visible in Figure 1 are agglomerates of individual nanoparticles rather than nanoparticles per se. White arrows point at the key particles discussed in the text. The dashed rectangle in (**c**) denotes an area brimming with spherical entities, which are fine nanoparticle agglomerates resting on or inside the cell. nucleus area in contact with the apparently intracellular irregularly shaped and sizable nanoparticle agglomerate. DAPI was used to stain the cell nuclei in blue, phalloidin to stain f-actin microfilaments in red, and OsteolImage to stain HAp nanoparticles in green. Another interesting cluster of nanoparticles to pay attention to is the one magnified in Figure 3b, apparently resting tangled with either the cell membrane or the microvilli extensions at the interface between the cell and its microenvironment, and having a pronouncedly elongated shape. This particle adheres parallel to the cell membrane, in a configuration that imposes a comparatively large membrane deformation energy cost to the cell upon its uptake [47], for which reason its rotation by \(\sim\)90\({}^{\circ}\) is expected to occur prior to the uptake [48]. In contrast to it, the particles appearing to be localized inside another cell, magnified in Figure 3c, appear rounder. The edges of these seemingly internalized particles are soft and curved, unlike the sharper edges of a set of particles tangled within the cellular filopodia and apparently resting outside the cell boundaries (Figure 3d). Inspired by this brief discussion of features intrinsic to a randomly taken AFM image, one question emerges: may the overly large particles as well as those with overly sharp edges be less prone to cellular internalization than their smaller and more softly shaped counterparts? This question is divisible to two distinct hypotheses, which will be treated and discussed in Sections 3.1 and 3.2, respectively: (1) the uptake is reduced as the size of the nanoparticle clusters increases, and (2) the uptake is diminished for nanoparticle clusters whose shapes deviate from the perfect spherical symmetry.