School of Chemical & Biomedical Engineering, Nanyang Technological University, 637459 Singapore. sychew.ntu.edu.sg.
Lee Kong Chian School of Medicine, Nanyang Technological University, 308232 Singapore.
Biomater Sci. 2018 Nov 1;6(11):3019-3029. doi: 10.1039/c8bm01034j. Epub 2018 Oct 2.
The use of human induced pluripotent stem cell-derived neural progenitor cells (hiPSC-NPCs) is an attractive therapeutic option for damaged nerve tissues. To direct neuronal differentiation of stem cells, we have previously developed an electrospun polycaprolactone nanofiber scaffold that was functionalized with siRNA targeting Re-1 silencing transcription factor (REST), by mussel-inspired bioadhesive coating. However, the efficacy of nanofiber-mediated RNA interference on hiPSC-NPCs differentiation remains unknown. Furthermore, interaction between such cell-seeded scaffolds with injured tissues has not been tested. In this study, scaffolds were optimized for REST knockdown in hiPSC-NPCs to enhance neuronal differentiation. Specifically, the effects of two different mussel-inspired bioadhesives and transfection reagents were analyzed. Scaffolds functionalized with RNAiMAX Lipofectamine-siREST complexes enhanced the differentiation of hiPSC-NPCs into TUJ1 cells (60% as compared to 22% in controls with scrambled siNEG after 9 days) without inducing high cytotoxicity. When cell-seeded scaffolds were transplanted to transected spinal cord organotypic slices, similar efficiency in neuronal differentiation was observed. The scaffolds also supported the migration of cells and neurite outgrowth from the spinal cord slices. Taken together, the results suggest that this scaffold can be effective in enhancing hiPSC-NPC neuronal commitment by gene-silencing for the treatment of injured spinal cords.
利用人诱导多能干细胞衍生的神经祖细胞（hiPSC-NPCs）是受损神经组织的一种有吸引力的治疗选择。为了指导干细胞向神经元分化，我们之前开发了一种电纺聚己内酯纳米纤维支架，通过贻贝启发的生物粘附涂层功能化，靶向 Re-1 沉默转录因子（REST）的 siRNA。然而，纳米纤维介导的 RNA 干扰对 hiPSC-NPCs 分化的效果尚不清楚。此外，尚未测试这种细胞接种支架与受损组织之间的相互作用。在这项研究中，优化了支架以实现 hiPSC-NPCs 中 REST 的敲低，从而增强神经元分化。具体来说，分析了两种不同贻贝启发的生物粘合剂和转染试剂的效果。用 RNAiMAX Lipofectamine-siREST 复合物功能化的支架增强了 hiPSC-NPC 向 TUJ1 细胞的分化（9 天后，与对照相比，siNEG 中的 siREST 复合物增加了 60%，而对照组中只有 22%），而不会引起高细胞毒性。当细胞接种支架被移植到横切的脊髓器官切片时，观察到类似的神经元分化效率。支架还支持细胞从脊髓切片中的迁移和突起生长。总之，这些结果表明，该支架通过基因沉默有效增强 hiPSC-NPC 神经元的承诺，可用于治疗受伤的脊髓。
