School of Pharmacy, Hangzhou Normal University, Hangzhou, 311121, Zhejiang, China.
Medtronic Technology Center, Shanghai, 201114, China.
J Nanobiotechnology. 2021 Dec 2;19(1):400. doi: 10.1186/s12951-021-01141-7.
The poor regenerative capability and structural complexity make the reconstruction of meniscus particularly challenging in clinic. 3D printing of polymer scaffolds holds the promise of precisely constructing complex tissue architecture, however the resultant scaffolds usually lack of sufficient bioactivity to effectively generate new tissue.
Herein, 3D printing-based strategy via the cryo-printing technology was employed to fabricate customized polyurethane (PU) porous scaffolds that mimic native meniscus. In order to enhance scaffold bioactivity for human mesenchymal stem cells (hMSCs) culture, scaffold surface modification through the physical absorption of collagen I and fibronectin (FN) were investigated by cell live/dead staining and cell viability assays. The results indicated that coating with fibronectin outperformed coating with collagen I in promoting multiple-aspect stem cell functions, and fibronectin favors long-term culture required for chondrogenesis on scaffolds. In situ chondrogenic differentiation of hMSCs resulted in a time-dependent upregulation of SOX9 and extracellular matrix (ECM) assessed by qRT-PCR analysis, and enhanced deposition of collagen II and aggrecan confirmed by immunostaining and western blot analysis. Gene expression data also revealed 3D porous scaffolds coupled with surface functionalization greatly facilitated chondrogenesis of hMSCs. In addition, the subcutaneous implantation of 3D porous PU scaffolds on SD rats did not induce local inflammation and integrated well with surrounding tissues, suggesting good in vivo biocompatibility.
Overall, this study presents an approach to fabricate biocompatible meniscus constructs that not only recapitulate the architecture and mechanical property of native meniscus, but also have desired bioactivity for hMSCs culture and cartilage regeneration. The generated 3D meniscus-mimicking scaffolds incorporated with hMSCs offer great promise in tissue engineering strategies for meniscus regeneration.
半月板的再生能力差且结构复杂，这使得其在临床上的重建极具挑战性。聚合物支架的 3D 打印技术有望精确构建复杂的组织架构，但由此产生的支架通常缺乏足够的生物活性，无法有效地生成新组织。
本研究通过冷冻打印技术构建了基于 3D 打印的策略，以制造出模仿天然半月板的定制型聚氨酯（PU）多孔支架。为了提高支架对人骨髓间充质干细胞（hMSCs）培养的生物活性，通过细胞死活染色和细胞活力测定研究了支架表面通过物理吸附胶原蛋白 I 和纤维连接蛋白（FN）进行改性。结果表明，FN 涂层在促进多方面干细胞功能方面优于胶原蛋白 I 涂层，并且 FN 有利于在支架上进行软骨形成所需的长期培养。通过 qRT-PCR 分析评估，hMSCs 的原位软骨分化导致 SOX9 和细胞外基质（ECM）的时间依赖性上调，免疫染色和 Western blot 分析证实了胶原 II 和聚集蛋白聚糖的沉积增加。基因表达数据还表明，3D 多孔支架与表面功能化相结合，极大地促进了 hMSCs 的软骨形成。此外，3D 多孔 PU 支架在 SD 大鼠皮下植入后不会引起局部炎症，并且与周围组织很好地整合，表明具有良好的体内生物相容性。
总体而言，本研究提出了一种制造生物相容性半月板构建体的方法，该构建体不仅可以再现天然半月板的结构和机械性能，而且还具有促进 hMSCs 培养和软骨再生的所需生物活性。与 hMSCs 结合生成的 3D 半月板模拟支架在半月板再生的组织工程策略中具有广阔的应用前景。
