Hydroxyapatite ha nanoparticles

Agarose (Fisher Scientific, Loughborough, UK) was mixed with water to a final concentration of 0.75% (w/v) with different concentrations of hydroxyapatite (HA) nanoparticles (Sigma-Aldrich, Gillingham, UK), namely 0% (v/v) for the muscle area, 0.2% (v/v) for the tendon area, and 40% (v/v) for the bone area. Type I rat tail collagen (Corning, Gillingham, UK) was prepared following the company’s instruction to a final concentration of 3 mg/mL. Therefore, the required volumes of sterile 10x phosphate buffer saline (PBS), sterile 1 N sodium hydroxide (NaOH) (Fisher Scientific, UK) in distilled water (dH₂O), and Agarose/HA solutions were calculated and mixed for each section in individual tubes. To avoid very rapid polymerization of Agarose, the solutions were kept in a water bath at 37 °C. Additionally, collagen was kept at 4 °C and added last to the mixture. After polymerization_, bone and tendon triphasic gels were crosslinked with 10% (v/v) oligomeric proanthocyanidins (OPC) in 1x PBS for 60 min at 37 °C and 5% CO₂. The resultant hydrogels are listed in Table 1.

All the reagents used in this study were of HPLC grade. Gellan gum (GG) (GelzanTM, Sigma-Aldrich Co., St. Louis, MI, USA) with an average molecular weight of 1,000,000 g/mole was purchased. PEG of [Poly (ethylene glycol)] (mw 6000, Sigma-Aldrich) and Trizol (Invitrogen, Life Technologies Co, Groningen, The Netherlands) were used. Hydroxyapatite (HA) nanoparticles were purchased from Sigma Aldrich (St Louis, MO, USA). The main characteristics of this HA powder were characterized by producer (mean diameter of about 70 nm, surface area of 27.6 m²/g, purity of 98.9%). Compound K solution was purchased commercially from general bio-Co. Ltd. (Gyeongcheon-myeon, Jeollabuk-do, Republic of Korea) and was completely characterized by the company in a previous research work [71 (link)].
All the scaffold components were previously used in tissue engineering and proved to be non-toxic [40 (link),47 (link),72 (link),73 (link)].