**3. Results**

#### **3.1. Accuracy of iliac bone resection by computer-aided surgery**

In all animals, navigation-guided osteotomy could be accurately performed on the virtual bone tumor exactly as planned on the CAD system. The CAD-designed, 3D printer-produced

**Figure 3.** Representative mid-axial plane images of computed tomography (CT) sequentially taken over the experi‐ mental period. (a) In the BMP-treated group, radiopaque images were observed on the IP-CHA surface 3 weeks post‐ operatively. The radiologically dense area around the implant increased in a time-dependent manner. The implant was completely covered by new bone at 12 weeks postoperatively. (b) No significant radiopaque images were observed in the control group up to 12 weeks postoperatively.

implant fitted into the bone defect, and the IP-CHA implant prepared by CAD and CAM fitted well in all experimental and control animals. Over the 12-week study period, all animals survived without any complication.

#### **3.2. Evaluation of bone defect repair using reconstructed CT images**

**Figure 3** shows representative mid-axial plane images sequentially taken over the experimen‐ tal period in the BMP-treated group and no-BMP control group. In the BMP-treated group, in which the IP-CHA surface was coated with the BMP-retaining dough material, radiopaque images were observed on the implant surface 3 weeks postoperatively. Thereafter, the radiologically dense area around the implant increased in a time-dependent manner, and the implant was completely covered by radiopaque images at 12 weeks postoperatively. In contrast, no significant radiopaque images were observed in the control group images up to 12 weeks postoperatively.

#### **3.3. Macroscopic and histological findings**

two or three 0.6 mm wires (Synthes Co., Ltd., Tokyo, Japan). The implant was carefully sutured in place to ensure contact with the surrounding muscle. A prophylactic antibiotic agent (10 mg/kg body weight) was administered to animals before and after the surgery. This animal

CT slice images of the iliac bone were obtained immediately and 3, 6, 9, and 12 weeks post‐ operatively to observe the bone repair reaction at the bone defect site and reconstructed into slice and 3D images as desired using the reconstruction software (Aze, Tokyo, Japan). CT images of the mid-axial slice of the implant and the next slice 1 mm from the center were analyzed. A high-density area surrounding the implant was considered to be a new bone area and quantified with ImageJ software (Wayne Rasband, National Institutes of Health, Bethesda, MD). Results were statistically analyzed by the Mann-Whitney U test with a significance level of P < 0.05. The ilium was harvested 12 weeks postoperatively and fixed in 70% ethanol. From un-decalcified specimens, 10 μm sections were prepared and subjected to Villanueva bone

In all animals, navigation-guided osteotomy could be accurately performed on the virtual bone tumor exactly as planned on the CAD system. The CAD-designed, 3D printer-produced

**Figure 3.** Representative mid-axial plane images of computed tomography (CT) sequentially taken over the experi‐ mental period. (a) In the BMP-treated group, radiopaque images were observed on the IP-CHA surface 3 weeks post‐ operatively. The radiologically dense area around the implant increased in a time-dependent manner. The implant was completely covered by new bone at 12 weeks postoperatively. (b) No significant radiopaque images were observed in

experiment was performed after institutional approval was obtained.

**2.3. Evaluation of efficacy of computer-assisted bone defect repairing system**

staining for histological evaluation. Specimens were macroscopically evaluated.

**3.1. Accuracy of iliac bone resection by computer-aided surgery**

the control group up to 12 weeks postoperatively.

**3. Results**

148 New Trends in 3D Printing

Macroscopic findings for the BMP-treated group of animals indicated that the whole implant was completely encased by a new bone, and the new bone was continuous with the original ilium, suggesting successful repair of the bone defect to the normal anatomical morphology

**Figure 4.** (a) BMP-treated group. The whole implant was completely encased by a new bone continuous with original ilium on macroscopic findings. (b) No bone tissue was observed on the implant surface in the control group.

(**Figure 4a**). In contrast, no bone tissue was observed on the implant surface in the control group of animals (**Figure 4b**). Representative histological specimens of un-decalcified tissues with Villanueva bone staining are shown (**Figure 5a–d**). In the BMP-treated group, the implant was covered by a new bone that was continuous with the original ilium, and the new bone was found to have entered pores inside the implant (**Figure 5c**). The dough material pasted on the implant surface was not observed in histological specimens obtained 12 weeks postoper‐ atively. This indicates biodegradation during the experimental period. No inflammatory response was observed around the implant over the experimental period.

**Figure 5.** Histological sections at 12 weeks after surgery [lower (\*20) and higher magnification (\*200), *scale bar* indicates 100μm, 8μm in each magnification]. (a, b) Boxed area of reconstructed axial CT image shows a histological picture of (c) (BMP-treated group) and (d) (control group), respectively. (c) In the BMP-treated group, the implant was covered by a new bone that was continuous with the original ilium, and the new bone was observed in the pore of the implant. d. In the control group, no bone tissue was observed. (arrowhead: new bone, asterisk: IP-CHA implant, cross: ilium, arrow: fibrous tissue, circle: osteoid, box: calcified bone).

#### **3.4. Changes over time in the amount of the new bone formed**

In the BMP-treated group, the volume of the calcified new bone tissue around the implant on CT slice images of the pelvis peaked 3 weeks postoperatively, then decreased over time, and finally resulted in the anatomical shape of the original ilium in 12 weeks postoperatively (**Figure 6**). In contrast, significant new bone formation was not observed in the control group during the observation period.

**Figure 6.** The amount of new bone formation of the BMP-treated group was significantly larger than that of the control group in reconstructed mid-axial CT planes at all experimental periods (statistical significant difference was set at *p*<0.05, values showed mean ± standard deviation).
