## 4 Discussion  
The results show that alkali hydrothermal treatment of titanium implants enhanced bone integration in the cortical portion and bone formation in the bone marrow portion of rabbit femurs _in vivo_. Ooseointegration is achieved by bone formation on the implant surface, which is largely associated with cell migration, cell adhesion, and cell differentiation [23]. Modification of the implant surface topography provides a suitable matrix for osteoblast cell adhesion, proliferation, differentiation, and bone formation. A modified titanium surface structure can potentially be applied in biomaterials. Alkali-treated titanium surfaces with nanostructures have been of great interest [22]. Alkali hydrothermal treatment of titanium can create a super hydrophilic titanate layer with nano-network structures. In the present study, the alkali hydrothermally-treated implant showed a modified surface with uniform nanostructure pores on SEM observation. Furthermore, examination of the contact angle during the wettability test indicated a super hydrophilic surface. These characteristics are consistent with those of previous reports [24; 25]. There are other advantages of the alkali hydrothermal treatment method. Acid treatment and blasting processes produce a microscale roughened structure on the surface rather than the nanoscale surface produced by alkali hydrothermal treatment [6]. The blasting treatment may be difficult to process uniformly depending  
Figure 8: Bone implant contact ratio (%; BIC). The BIC values of the whole area and bone area were higher in the alkali group than in the control group. SD: standard deviation; * \(p\) < 0.01. Figure 9: Bone tissue area ratio (%; BTA). The BTA values of the whole area and marrow area were higher in the alkali group than in the control group. SD: standard deviation; * \(p\) < 0.01.  
on the composition of the target, such as those with a porous or fibrous structure. In acid treatment, there is concern that corrosion reduces the mechanical strength of metal objects. However, alkali hydrothermal treatment is a simple chemical treatment, and any object can be treated uniformly. In addition, since this process forms a sodium titanate layer on the surface, there is little influence on the structure. To evaluate osseointegration, we performed histological observation and histomorphometric measurements. Measurements were performed in two areas and combined to a third one for analysis: the cortical bone portion, the bone marrow portion, and the whole portion comprising the area the length of the implant. Analysis of the whole portion was undertaken to evaluate implant stability in the surrounding bone. Analysis of the cortical bone portion evaluated the integration between bone and implant, because the implant was in contact with surrounding bone at placement. Finally, analysis of the bone marrow portion was undertaken to evaluate osseoinduction, because this portion was originally without bone. Histological observations revealed osseointegration for both the alkali group and the control group. The BIC ratios in the whole portion and cortical bone portion were higher in the alkali group than in the control group. This result seems to be related to the higher bone integration at the cortical bone portion. Several studies have reported that the surface of alkali-treated titanium promotes osteoinduction by the nanoscale structure _in vitro_, the hydrophilicity of the surface promotes initial cell adhesion, and nutrient supply and is advantageous for bone regeneration at early stages [23; 26]. Additionally, alkali-treated titanium forms an apatite layer on the surface in SBF [17; 18; 19]. When titanium is immersed in a NaOH solution and heated, an amorphous layer of alkali titanate is formed and strongly bonds to the substrate. When this alkali-treated titanium is soaked in SBF, alkali ions are eluted from the surface layer, the surface gel induces nucleation of apatite, and an apatite layer is formed [27].