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Micro-aggregates have several unique properties that make them suitable for tissue engineering: they are associated with increased cell viability, they enhance multilineage differentiation and they secrete trophic factors with an anti-inflammatory effect. In addition, micro-aggregates have the ability to fuse into one larger structure when brought into close contact. Hence, they are of great interest in bottom-up tissue engineering, in which large heterotypic tissues that mimic the native organisational tissue structure are gradually built up by using small units as building blocks. An important step for the application of micro-aggregates in tissue engineering is a complete understanding of their formation. During this formation, a change in cell morphology, determined by cell contractility and cortical tension, is required. These factors are both determined by the structure and functionality of the cytoskeleton. Therefore, the importance of different components of the cytoskeleton during the aggregation of hPDCs was studied by adding different inhibitors to the medium. In this thesis, Y-27632, a ROCK-inhibitor, and (S)-nitroblebbistatin, an inhibitor of MYH ATPase, were evaluated for their effect on aggregate formation. Two different aggregate sizes were tested since cell number might influence cell aggregation by affecting contact opportunities between cells and cell survival. The dynamics of the aggregation behaviour were captured using time-lapse microscopy. Area and circularity of the aggregates were extracted from the images and analysed using a data-based modelling approach. It is demonstrated that ROCK-inhibitor reduces compaction and slows down the formation of micro-aggregates compared to control conditions. Moreover, the effect of Y-27632 is found to be depended on aggregate size. Experiments with (S)-nitroblebbistatin were not coherent. Consequently, no conclusions concerning the contribution of myosin II to cellular aggregation could be made. In addition to time-lapse microscopy, aggregates treated with ROCK-inhibitor were imaged in confocal microscopy in order to quantify cell viability and visualise the actin network as well as the nucleus. Although this analysis did not reveal any statistically significant results, some clear trends are observed. First of all, it is stated that cells at the boundary have a higher volume compared to central cells. Secondly, it is seen that boundary cells have a tendency to become more spherical as the concentration of Y-27632 is increased. It is believed that a more complete insight in aggregate formation is the first step towards a better process control and thus would enhance the performance of the final engineered tissue.