The complex meniscus tissue plays a critical role in the knee. The high susceptibility to injury has led to an intense pursuit for better tissue engineering regenerative strategies, where scaffolds play a major role. In this study, indirect printed hierarchical multilayered scaffolds composed by a silk fibroin (SF) upper layer and an 80/20 (w/w) ratio of SF/ionic-doped β-tricalcium phosphate (TCP) bottom layer were developed. Furthermore, a comparative analysis between two types of scaffolds produced using different SF concentrations, i.e., 8% (w/v) (Hi8) and 16% (w/v) (Hi16) was performed. In terms of architecture and morphology, the produced scaffolds presented homogeneous porosity in both layers and no differences were observed when comparing both scaffolds. A decrease in terms of mechanical performance of the scaffolds was observed when SF concentration decreased from 16 to 8% (w/v). Hi16 revealed a static compressive modulus of 0.66 ± 0.05 MPa and dynamical mechanical properties ranging from 2.17 ± 0.25 to 3.19 ± 0.38 MPa. By its turn, Hi8 presented a compressive modulus of 0.27 ± 0.08 MPa and dynamical mechanical properties ranging from 1.03 ± 0.08 MPa to 1.56 ± 0.13 MPa. In vitro bioactivity studies showed formation of apatite crystals onto the surface of Hi8 and Hi16 bottom layers. Human meniscus cells (hMCs) and human primary osteoblasts were cultured separately onto the top layer (SF8 and SF16) and bottom layer (SF8/TCP and SF16/TCP) of the hierarchical scaffolds Hi8 and Hi16, respectively. Both cell types showed good adhesion and proliferation as denoted by the live/dead staining, Alamar Blue assay and DNA quantification analysis. Subcutaneous implantation in mice revealed weak inflammation and scaffold’s integrity. The hierarchical indirect printed SF scaffolds can be promising candidate for meniscus TE scaffolding applications due their suitable mechanical properties, good biological performance and possibility of being applied in a patient-specific approach.

中文翻译：

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间接打印弯月面组织工程的特定于患者的分层支架

复杂的半月板组织在膝盖中起关键作用。对损伤的高度敏感性导致人们强烈寻求更好的组织工程再生策略，其中支架在其中起主要作用。在这项研究中，开发了由丝素蛋白（SF）上层和SF /离子掺杂的β-磷酸三钙（TCP）底层组成的比率为80/20（w / w）的间接印刷分层多层支架。此外，对使用不同SF浓度（即8％（w / v）（Hi8）和16％（w / v）（Hi16））生产的两种类型的支架进行了比较分析。就结构和形态而言，所产生的支架在两个层中均呈现均匀的孔隙率，并且当比较两个支架时未观察到差异。当SF浓度从16降低到8％（w / v）时，观察到支架的机械性能降低。Hi16的静态压缩模量为0.66±0.05 MPa，动态力学性能为2.17±0.25至3.19±0.38 MPa。同时，Hi8的压缩模量为0.27±0.08 MPa，动态力学性能为1.03±0.08 MPa至1.56±0.13 MPa。体外生物活性研究表明，磷灰石晶体在Hi8和Hi16底层的表面形成。将人类半月板细胞（hMCs）和人类原代成骨细胞分别培养在分层支架Hi8和Hi16的顶层（SF8和SF16）和底层（SF8 / TCP和SF16 / TCP）上。如活/死染色所示，两种细胞类型均显示出良好的粘附和增殖，Alamar Blue分析和DNA定量分析。小鼠皮下植入显示出较弱的炎症和支架的完整性。分层的间接印刷SF支架由于其合适的机械性能，良好的生物学性能以及适用于患者特定方法的可能性，因此有望成为弯月形TE支架应用的有希望的候选者。