The fabrication of biomaterials to be used in segmental bone defects, mimicking the bone's organic-inorganic architecture and mechanical properties to induce osteogenesis, persists as a key challenge. The purpose of this study was to elucidate the effect of a lightweight, morphologically graded, and multiphase self-standing scaffold structure prepared from a combination of gelatine (Gel), collagen type 1 (Col) and/or hydroxyapatite (HAP) nanoparticles by a unidirectional freeze-casting process at different temperatures (−20, −40, −60 °C), followed by carbodiimide induced cross-linking, on their in-vitro mechanical stability and bioactive properties. In addition, the rheological study of differently formulated Gel solutions has been performed to determine the effect of Col and HAP content on their microstructural arrangement, which, together with the freezing kinetic, affects Gel/Col orientation and cross-linking, and, thus, the scaffold's mechanical strength and stability.

A bone-like anisotropic, interconnected, and graded porosity (from 120 to a few μm) scaffold structure with up to 30% total porosity and ~61 μm average pores' diameter is obtained by using a higher Col content (Col: Gel = 2:5) and freezing temperature (−20 °C) while forming a few μm thick close-to-parallel lamellae, separated with a 10–100 μm space when prepared at −60 °C. Such a structure influenced in-vitro stability strongly (lower swelling without weight loss), being accompanied with a ~76% increase of compression strength (to 37 kPa) and ~67% decrease of elastic modulus (to 17 kPa) when prepared with HAP and incubated in HBSS for 7 days. On the other hand, a significant reduction of both strength (~78%, to 15 kPa) and elasticity (~95%, to 5 kPa) was noted for a scaffold prepared with HAP at −60 °C, being related to faster degradation and the formation of a highly opened structure on the bottom, required to stimulate the bone ingrowth, while a more closed network structure on the top to adhere with the surrounding soft tissue. None of the scaffolds induced cytotoxicity to human bone-derived osteoblasts, even after 19 days of incubation, but rather improved their viability while promoting cells' adhesions, proliferation, and differentiation, being supported with an increased alkaline phosphatase activity and rod-like CaP formation.

用于节段性骨缺损的生物材料的制造，模仿骨骼的有机-无机结构和机械特性以诱导成骨，仍然是一个关键挑战。本研究的目的是阐明由明胶 (Gel)、1 型胶原蛋白 (Col) 和/或羟基磷灰石 (HAP) 纳米粒子组合制备的轻质、形态分级和多相自立支架结构的效果。在不同温度（-20、-40、-60°C）下进行单向冷冻铸造工艺，然后是碳二亚胺诱导的交联，对其体外机械稳定性和生物活性特性进行了分析。此外，还对不同配方的凝胶溶液进行了流变学研究，以确定 Col 和 HAP 含量对其微观结构排列的影响，其中，

通过使用较高的 Col 含量（Col：Gel = 2），获得了具有高达 30% 的总孔隙率和 ~61 μm 平均孔径的骨状各向异性、互连和分级孔隙率（从 120 到几微米）支架结构:5) 和冷冻温度 (-20 °C)，同时形成几微米厚的接近平行的薄片，在-60 °C 下制备时以 10–100 μm 的空间分隔。这种结构强烈影响体外稳定性（较低的溶胀而不减轻重量），当用 HAP 制备时，伴随着压缩强度增加约 76%（至 37 kPa）和弹性模量降低约 67%（至 17 kPa）并在 HBSS 中培养 7 天。另一方面，在 -60 °C 下用 HAP 制备的支架的强度（~78%，至 15 kPa）和弹性（~95%，至 5 kPa）均显着降低，与更快的降解和底部高度开放结构的形成有关，需要刺激骨骼向内生长，而顶部更封闭的网络结构与周围的软组织粘附。即使在孵育 19 天后，这些支架都没有对人骨源性成骨细胞产生细胞毒性，而是在促进细胞粘附、增殖和分化的同时提高了它们的活力，并得到了增加的碱性磷酸酶活性和棒状 CaP 形成的支持.