## 4 Discussion
As compared with the literature, fiber thickness is less than that of collagen fibers in thin films created by spin coating methods [36; 37]. By this technique, thin films consisting of randomly-oriented collagen fibrils were obtained, with the diameter varying from 300 nm up to 700 nm. The surface topography may play an important role in cellular response to collagen [38; 39]. According to the literature, the Young modulus (average value) of a single collagen type 1 fiber is between 1.2-2 GPa [36], as the mechanical strength of a single collagen fiber is due to hydrogen bonds that stabilize the triple helix within the collagen molecules. In the case of a membrane, which is formed by a network of crosslinked collagen fibers, higher Young modulus values are expected. Different values of Young modulus, related to each manufacturer, can be correlated to the typically required chemical or physical pretreatment, aiming to preserve and sterilize the tissue. These treatments include chemical, enzymatic, or mechanical decellularization techniques in order to remove cellular components [26; 40]. The resulting acellular tissue may retain, to a certain extent, the natural mechanical properties and promote tissue remodeling by neovascularization and recellularization. These aspects could be responsible for the relatively high scattering of data in the case of Biocollagen(r) and Jason(r), compared with Cova(tm)Max, before PRGF treatment. On the other hand, upon PRGF treatment, the changes of Young modulus values could be correlated with the ultrastructural properties of each membrane type, especially the porosity. The higher the porosity, the lower the Young modulus value obtained, which is consistent with similar results previously reported in literature [25]. This is also the reason why, after PRGF treatment, the Young modulus values are displayed in a narrow interval (2.8-4 GPa), compared with the corresponding values before treatment. Also, according to the literature, it has been demonstrated that as the collagen concentration or density is increased, the tissue becomes stiffer, and because of this, cells behave differently at different collagen concentrations [41]. Therefore, because the Young modulus is correlated with collagen density modifications, at physiological rate, the cells can sense this difference. The enzymatic (trypsin) degradation test also emphasized different behaviors of the membranes with or without PRGF treatment. As a general behavior, PRGF-modified membranes exhibited a slower degradation compared with the native ones, in all the three cases. The differences observed for the native membranes can be explained by the cross-linking techniques used by the manufacturers, as any cross-linking technique may have uncertain effects on the structure of collagen membrane [42]. On the other hand, the trypsin treatment also caused a time-dependent release of water-soluble proteins; the disaggregation of collagen fibrils occurs as a consequence of the degradation of proteoglycan bridges between collagen fibrils. Previous results indicated that in the case of trypsin-treated collagen fibers (over 72 h), the collagen fibrils still kept their structural integrity by retaining their triple-helical conformation [43]. [PERSON] et al. [42] previously reported that higher collagen density and compact structure may be a reason of slower biodegradation. However, further studies are required in order to assess whether the results obtained in our study can be applied to humans, as the local environment in the oral cavity is a complex biochemical \"reactor\" difficult to mimic. The present study was only limited to structural and mechanical characterization of the commercial collagen membranes incorporating PRGF factors. Our next approach will be focused on the biological performance of these membranes, performing in vitro assays to evaluate the cellular adhesion, proliferation, and viability of human osteoblasts, fibroblasts, and keratocytes.