## 3 Discussion  
Given the inherent shortcomings of autologous materials, xenogeneic natural materials have been considered as alternatives. Exploring different natural materials within the tissue engineering field primarily seeks to bio-mimic the natural ECM environment more accurately and intends to facilitate genuine tissue regeneration [26]. In addition, the limited sources of hPDL-MSCs play a primary role in the regeneration of periodontium, particularly periodontal ligament, and therefore, understanding their interaction with various materials is crucial to estimating their potential clinical effectiveness. In this study, we investigated the attachment, growth, and expression of functional parameters in hPDL-MSCs grown on two xenogeneic-derived scaffolds developed for application in periodontal regenerative medicine. The healing process after periodontal surgery, like any wound healing, consists of four overlapping phases: hemostasis, inflammation, proliferation, and tissue remodeling. On the cellular level, the following events occur: cell migration to the surgery area and attachment to the scaffold, their proliferation, production of various biologically active proteins, and differentiation to the functional cells. The first prerequisite for any three-dimensional (3D) scaffold is an effective cell attachment and establishing a cellular network of interacting cells [27]. SEM and fluorescence  
Figure 7: VEGF protein production by cells and absorption by materials. **(a)**—hPDL-MSCs were cultured on different substrates for 3, 7, and 14 days, and the content of VEGF in conditioned media was measured by ELISA. **(b)**—the scaffolds were incubated in a medium containing 10 ng/mL recombinant VEGF protein for 6 h at 37 \({}^{\circ}\)C, and the content of VEGF in conditioned media was determined by ELISA at the end of the incubation. **(c)**—after incubation with recombinant VEGF, the scaffolds were incubated for an additional 24 h in a medium, and the amount of released VEGF was measured by ELISA. Data are presented as mean \(\pm\) SD of five independent experiments with hPDL-MSCs isolated from five different donors. * and **—significantly different between substrates with \(p\) < 0.05 and 0.01, respectively. microscopy observations revealed the continued growth of hPDL-MSCs on both scaffolds. Regarding the expansion area, cells on GPP performed better than those on VSCM. After 3 days, SEM showed that cells grown on GPP were tightly intertwined with surrounding nanofiber structures. SEM observation from day 14 indicated that hPDL-MSCs cultured on two types of materials encased by continuous sheet-like structures which tended to cover the material's surfaces, implying the possible formation of ECM-like structures. The presence of nanofibers in GPP seems to be an essential factor influencing hPDL-MSCs attachment, as they provide the attachment points and also influence cell morphology and orientation. Similarly, previous researchers discovered that the arrangement of PDL stem cells was determined by the orientation of PCL/GE fibers [28]. In contrast, visual assessments indicated fewer cells on VSCM at the early stage of culture. This can be explained by the pore size of VSCM, which was comparable with the size of single cells and thus allowed potential cell migration within a scaffold. Thus, the scaffold architecture and structural features are essential for the initial interaction with cells: nanofibers facilitate cell attachment and might be used for cell spatial orientation, whereas the presence of microscale pores allows cell migration inside the scaffold. After a 2-week prolonged culturing, both GPP and VSCM scaffolds were covered by cells, but some differences in the coverage degree and cell morphology were still observed. The simple presence of cells on materials does not reflect their metabolic integrity, and cell proliferation and metabolic activity serve as fundamental metrics for evaluating the biological performance of scaffolds. Over a 2-week culture period, hPDL-MSCs grew consistently on both scaffolds.