**4. Discussion**

UV photofunctionalization, a method of modifying titanium surfaces after UV treatment that includes altering the physicochemical properties and enhancing biocompatibilities, has been proposed to reset the biological aging of titanium [7]. After treatment with UV radiation, the TiO2 layer of a titanium surface incorporated with hydrocarbons became remarkably hydrophilic or superhydrophilic. The amount of surface carbon is known to vary depending on the age of the surface and reportedly can increase to approximately 60% to 70% of surface atomic components. UV treatment cleans such carbon-contaminated titanium surfaces, reducing the carbon percentage to less than 20% and concurrently increasing the level of osseointegration [7]. In the UV spectrum, both UV-A and UV-C convert biologically aged titanium surfaces from hydrophobic to superhydrophilic, but only UV-C (200–280 nm) is known to effectively reduce surface carbon to a level equivalent to a new surface and enhance bioactivity [17]. UV-C is capable of removing hydrocarbon from a TiO2 layer of titanium by direct photodecomposition, which is more effective than photocatalysis by UV-A [18]. In our study, all implant fixtures were photofunctionalized by UV-C and VUV (100–200 nm), and the radical removal of hydrocarbon from TiO2 layer of titanium could provide more superhydrophilicity [15,19]. VUV is rapidly absorbed by water in the atmosphere and is therefore capable of generating various reactive oxygen species by breaking hydrogen bonds in water molecules via hydrolysis [19]. VUV-initiated hydrolysis is an efficient method of obtaining hydroxide or hydroxyl groups on a TiO2 layer that provides persistent superhydrophilicity [20,21]. Since VUV treatment tends to generate more ozone in the atmosphere and hydroxyl radicals in water [19], it should be strictly limited in only laboratory or factory, not clinical, settings.

To accelerate bone healing and improve bone anchorage to an implant, the bone/implant interface can be improved topographically and biochemically by incorporating inorganic phases, such as calcium phosphate, and organic molecules, such as proteins, enzymes or peptides, on or into a TiO2 layer [22–25]. Nanostructured implant surfaces, which have an extensive surface area, high surface free energy, and wettability, seem capable of modifying the host tissue response [9]. SA implant surfaces have demonstrated predictable clinical results and are regarded as standard implant surface [10,26–29]. The superhydrophilicity of SA + VUV + BS, which was previously found on the flat surface of disks [15], was confirmed in our test of static surface wettability. As the ability to attract blood near to the top of the implant fixture (approximately 4 mm above the horizontal plate in our study) is what most clinicians ultimately prefer to check in a clinical setting, we added a new dynamic test of surface wettability to compare the wetting velocities of SA + VUV and SA + VUV + BS in more detail. Since SA is a dry surface that has not been in contact with any liquid, the surface wettability of SA + VUV + BS might be significantly superior to that of SA solely by the effect of the pH-buffering agent of SA + VUV + BS itself. However, given that implant fixtures are placed with dry surfaces or without any additional hydration in real clinical situations, we chose a naïve SA, rather than an SA hydrated in solution, as a control [30]. A more hydrophilic surface was previously found to be closely related to superior and faster osseointegration [30,31]. Furthermore, surface wettability is known to alter the biological responses of implant surfaces with respect to the adhesion of proteins and other molecules, as well as cell interactions [32].

As blood clot formation signals the beginning of the healing process, the interaction between an implant and blood is considered important for the successful osseointegration of titanium implants after implantation [33]. Blood clot formation on rough titanium surfaces can induce cell recruitment and stimulate wound healing [34], and it has been revealed that both preosteoblasts and osteoblasts can attach to an implant surface covered by platelets and fibrin, where they differentiate under the stimulation of osteogenic factors and cytokines released from the peri-implant blood clot [35,36]. The formation of blood clots on the implant with various implant surfaces is believed to be a crucial factor in effective fibrin retention and may critically affect bone healing and osseointegration by influencing macromolecule transport, cell behavior, and contact/distant osteogenesis [34]. In a test of static blood clotting, the SA + VUV + BS showed superior blood absorption around the implant fixtures compared with SA + VUV, but not to statistically significant degree. This indicates that an SA surface photofunctionalized with VUV has at least an equal ability in blood clotting. We designed new experiments to confirm the blood clotting by hemostasis of continuous capillary bleedings to simulate real clinical situations as accurately as possible. In a test of in vitro dynamic blood clotting, the clots formed significantly faster, and the total volume of blood collecting through the gap between holes and implant fixture to hemostasis was significantly less in SA + VUV + BS than in SA + VUV. An in vivo test also showed a significant difference in the total weight of bleeding between the overprepared hole and an implant fixture among SA, SA + VUV, and SA + VUV + BS. This suggests that SA + VUV + BS can induce faster blood clot formation around the implant surface, leading to more effective interaction of the bone-to-implant interface for osseointegration. In a clinical respect, these features of SA + VUV + BS are important in visualizing the surgical site and simultaneous guided bone regeneration, which is frequently indicated for the adequate quantity and quality of peri-implant tissues for more aesthetic and functional results [37], because rapid blood clotting is closely associated with the stabilization of grafting material and the barrier membrane. Finally, SA + VUV could be an alternative to SA + VUV + BS to SA with respect to the potential for blood clot formation on implant surfaces.

During implant drilling in the bone, which produces a weakly acidic condition, a pH buffer may help keep the pH constant [38]. As a coating material, the pH-buffering agent appears to control the release of the inflammatory mediators [9] and enhance the conditions for osteoblast activity [15] by keeping the pH constant or at least preventing significant changes. The activity of platelets in blood clotting and both the activity of osteoblasts and the level of ALP for bone making are also inhibited by extracellular acidosis [11]. SA + VUV + BS could maximize the activity of platelets, thrombogenesis, the activity of osteoblasts, and the level of ALP in a bone-to-implant interface through a pH-buffering effect. Further studies will be necessary to investigate SA + VUV + BS with respect to its safety and effectiveness in clinical settings. Randomized controlled trials should also be followed to confirm its feasibility in various clinical conditions, such as implant placement immediately after tooth extraction or with simultaneous bone augmentation.

**Author Contributions:** Conceptualization, C.-J.P. and H.-K.P.; methodology, I.-S.J. and J.-D.S.; software, A.M.K. and H.-K.P.; validation, C.-J.P., K.-G.H., and H.-K.P.; formal analysis, J.H.L., M.T., K.-G.H., and A.M.K.; investigation, I.-S.J. and J.-D.S.; resources, M.T., K.-G.H., and J.H.L.; data curation, H.C., G.-J.C., and C.K.; writing—original draft preparation, C.-J.P. and J.H.L.; writing—review and editing, M.T., H.C., and S.H.J.; visualization, G.-J.C., C.K., and S.H.J.; supervision, H.-K.P. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was supported by the research fund of Medicine-Engineering-Bio (MEB) Global Center for Developmental Disorders, Hanyang University (HY-2020-000-0000-2809).

**Conflicts of Interest:** The authors declare no conflict of interest.
