## 4 Discussion  
In this study, we analyzed the physical property of a 3D SF scaffold for achievement in MSC cultivation. Morphometry and specifically porosity represent relevant aspects for the success of scaffolds in tissue engineering. Adequate values of the main 3D morphometric outcomes [34], Obj.S/Obj.V and St.Sp, allow and facilitate entry and migration within the newly formed bone tissue scaffold [21]. A high porosity improves the invasion of the graft by progenitor cells, the migration of mesenchymal cells and osteoblasts to the recipient site. A great amount of porosity mainly characterized by open porosity, the presence of many and small pores, and high contact surface-volume ratio promote colonization, cell adhesion, and degradation of the scaffolds [21, 35].  
The mentioned aspects were detected in the investigated samples within the main indices. Appropriate values of Obj.S/Obj.V and St.Sp were recorded (Table 1), in particular for the initially manufactured scaffold--SSF, as well as for Obj.V/TV, St.Th, and Conn.D, that suggest good mechanical properties. The total porosity with the not-relevant presence of closed porosity resulted in 86% for SSF (Table 2) and higher than 80% for all the other samples, representing a favorable amount for tissue engineering applications. Micro-CT analyses were carried out with a resolution of about 9.7 \(\upmu\)m for SSF, in line with other studies [36], and with a higher one of about 6.5 \(\upmu\)m for the samples with cells; therefore, due to the intrinsic technology limitation, smaller pores than these values could be present, incrementing the overall amount. The porous structures are easily visualizable from the 3D rendering (Figure 1), the 3D model section reconstructed (Figure 2), the internal cross-sectional images of Figure 3, and FE-SEM micrographs in Figure 4.  
High connectivity was identified with the presence of interconnected pores (Table 2), showing a relatively uniform distribution of cells within the scaffolds, confirmed by FE-SEM (Figure 4) and CLSM observations (Figure 5).  
Furthermore, the study allowed examining the effect of time on the usability of the scaffold structure (sample SSFtime) after the usual time of storage, demonstrating the changes in the internal distribution of the material in terms of porosity and morphology, which was mostly preserved, although not entirely. Quantitatively, variation in the structural thickness and separation were recorded as well as in a major amount of closed porosity and object surface, suggesting an initial process of degradation, hardening, and thickening among the biomaterial surfaces. Qualitatively, the effect of the process was also evident from the 3D rendering of the external surfaces at higher density with respect to the SSF sample (Figure 1B) and from the internal cross-sectional images (Figure 3B). Considering the preliminary results, at this time, the focus of the study was kept on the SSF, and the interaction of the scaffold SSFtime with cells was not investigated. Similarly, cells' viability, growth rate, and MSC differentiation were not investigated. Nevertheless, CLSM and flow cytometry analysis showed that MSC, over a three-week period, colonized and distributed throughout the scaffold.  
Therefore, further investigations are required to say if and how any modifications in time may affect the cell adhesion and the cell growth.  
Developments will include further evaluations starting from the successful results achieved for the porous scaffold designed, manufactured and assessed here and the comparison with other advanced and innovative manufacturing technique such as 3D bioprinting.  
Moreover, technical assessment for selected applications will be performed including the setting up of custom experimental mechanical testing for addressing specific biomechanical and functional requirements.