**1. Introduction**

Titanium has been widely used for dental and orthopedic restoration and reconstruction due to its biocompatibility, resistance to corrosion, and mechanical properties. Titanium oxidizes easily, forming a thin (1–5 nm), stable, and passive layer that is self-limiting and protects the surface of the metal from further oxidation [1]. This titanium dioxide (TiO2) surface layer is considered to be responsible for its effective biological performance due to the transfer of calcium and phosphorus ions from the bone matrix within the TiO2 layer [2]. However, significant reductions in osseointegration and other biological capabilities of titanium occur over time as surface carbon increases because of an unavoidable deposition of carbon from the atmosphere on the TiO2 layer in a form of hydrocarbon [3]. This phenomenon is defined as the biological aging of titanium, and the ability of titanium surfaces to attract proteins and osteogenic cells decreases in a time-dependent manner [4]. Another notable change in titanium surfaces with time is the disappearance of hydrophilicity. Immediately after processing, titanium surfaces exhibit a contact angle of water of 0 or less than 5 degrees, and such surfaces are called superhydrophilic [4–7]. This feature gradually attenuates and becomes hydrophobic in 2 and 4 weeks, with a contact angle of greater than 40 and 60 degrees, respectively.

Surface treatment is used to modify dental implant surface topography and energy, resulting in improved wettability, increased cell proliferation and growth, and accelerated osseointegration [1,8,9]. Surface treatment can be achieved by an additive or subtractive technique [9]. The subtractive technique either removes or roughens a layer of core material, as typified by a sand-blasted and acid-etched (SA) surface. In the addictive technique, other materials or chemical agents are added superficially to the surface of the titanium through coating, such as titanium plasma spraying, hydroxyapatite coating, calcium phosphate coating, and other biomimetic coating. Drilling prior to implant placement causes bone tissue to undergo trauma similar to a fracture. The site becomes relatively hypoxic, and the extracellular pH becomes acidic. In such conditions, bone marrow stromal cells exhibit reduced alkaline phosphatase (ALP) activity and collagen synthesis, both of which are important in bone formation and osseointegration [10]. Glycolysis and DNA synthesis of osteoblasts are also found to be affected by acidic conditions [11]. Platelet aggregation, which is a critical step in blood clot formation or thrombogenesis, is also reduced by extracellular acidosis, as mediated by the calcium ion entry pathway [12]. Formation of a sufficient blood clot offers a direct and stable link at the bone-to-implant interface and plays an important role in thrombogenic responses and osseointegration [13]. Moreover, a relationship was found between various implant surface and the extent of the fibrin clot [14].

In our previous study, a novel SA surface coated with a pH-buffering agent after vacuum-UV (VUV) treatment was introduced [15,16]. This surface was closely associated with greater affinity for proteins, cells, and platelets, which promoted rapid and stable blood clotting, thrombogenesis, and osseointegration. The purpose of the present study was to evaluate and compare the surface wettability and blood clotting abilities of various implant surfaces, including a conventional SA surface (SA), an SA surface with VUV treatment (SA + VUV), and an SA surface coated with a pH-buffering agent after VUV treatment (SA + VUV + BS), by in vitro and in vivo analyses.
