Go solution

The titanium (iv) isopropoxide (C₁₂H₂₈O₄Ti, 98%), polyvinylpyrrolidone (PVP, M_w = 1,300,000), dimethylformamide (DMF, anhydrous, 99.8%), GO solution (2 ml mg⁻¹) and Poly(allylamine hydrochloride) (PAH, M_w = 900,000) were purchased from Sigma-Aldrich. Acetic acid (CH₃COOH, 99.9% (m/m)) was purchased from Junsei. We used all materials without further purification.

The technique for converting GO into rGO involved heating it in a solution containing EGCG (BMG Inc., Kyoto, Japan). The procedure began by diluting the commercial GO solution (Sigma-Aldrich, St. Louis, MO, USA) to a concentration of 1 mg mL⁻¹ in sterilized deionized (DI) water. Next, EGCG powder was added to the solution to achieve a final concentration of 10 mg mL⁻¹. As shown in Figure 1a, the resulting mixture was tightly sealed and heated at 80 °C for 8 h. After heating, the mixture was sonicated for 1 h and gradually cooled down in water over a period of 12 h. Atlantic bluefin tuna (Thunnus thynnus) bone (Dongwon, Seoul, Republic of Korea) was cleansed thoroughly by washing with DI water to remove organic components. The bones were then boiled for 20 min and treated with 0.1 N NaOH, followed by rinsing with DI water. To purify HAp, the bones were calcined at 800 °C (Figure 1b). For the control, commercially available water-soluble HAp powder (Dentis Co., Ltd., Daegu, Republic of Korea) was used.

The functionalization of RGD peptides into the electrospun fiber mats was accomplished by utilizing RGD-M13 phages as reported in our previous studies [3 (link), 30 (link)]. The RGD peptides were expressed on the side wall of M13 phage by genetic engineering. Briefly, an inverse polymerase chain reaction cloning method was conducted to express RGD peptides on major coat proteins of M13 phages, as describe elsewhere [5 (link), 23 (link), 30 (link)].
RGD-GO-PLGA mats were produced by electrospinning technique. The electrospinning solution was prepared by dissolving PLGA (lactide/glycolide molar ratio = 75/25, molecular weight = 70 000–110 000 Da, 200 mg/ml, Evonik Industries, Essen, Germany) and RGD-M13 phages (10 mg/ml) in 1, 1, 1, 3, 3, 3-hexafluoro-2-propanol (Sigma-Aldrich Co., St Louis, MO, USA), and then GO solution (2 mg/ml, Sigma-Aldrich Co.) in water was blended with RGD-M13 phage and PLGA blend solution. Electrospinning was conducted by loading the RGD-M13 phage, GO and PLGA blend solution in a syringe attached to a 21-gauge needle. A voltage of 14 kV was applied and the blend solution was injected at a feeding rate of 0.2 ml/h. A steel rotating wheel covered by aluminum foil was placed 11 cm from the needle tip to collect the nanofibers. After then, RGD-GO-PLGA mats were kept in vacuum for at least 8 h at room temperature in order to eliminate all remaining solvent.