Soft substrates promote direct chemical reprogramming of fibroblasts into neurons.
Soft substrates promote direct chemical reprogramming of fibroblasts into neurons.
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DOI：
10.1016/j.actbio.2022.08.049
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发表时间：
2022-08
影响因子：
9.7
通讯作者：
Ziran Xu;Yan Li;Pengdong Li;Yingying Sun;Shuang Lv;Yin Wang;Xia He;Jinying Xu;Zhixiang Xu;Lisha Li;Yulin Li
中科院分区：
文献类型：
--
作者：
Ziran Xu;Yan Li;Pengdong Li;Yingying Sun;Shuang Lv;Yin Wang;Xia He;Jinying Xu;Zhixiang Xu;Lisha Li;Yulin Li
成纤维细胞可以通过小分子的组合直接重编程以产生诱导神经元（iN），绕过中间阶段。这种方法为再生医学带来了巨大的希望;然而，它仍然效率低下。最近的研究表明，物理因素可能会改善成纤维细胞直接重编程为神经元，但其潜在的机制仍有待进一步探讨，迄今报道的物理因素并没有表现出细胞外基质（ECM）的全部特性。以前的体外研究主要使用刚性聚苯乙烯培养皿，而神经元原生体内环境的特征之一是大脑ECM的柔软性。所报道的脑组织硬度非常软，范围在100 Pa和3 kPa之间，并且尚未探索基底硬度对直接神经元重编程的影响。在这里，我们第一次表明，软基板大大提高了生产效率和质量的iNs，而不需要在重编程过程中与神经胶质细胞共培养，产生更多的电生理功能的神经元在更短的时间。转录组测序表明，软底物可能通过整合素、肌动蛋白细胞骨架、Hippo信号通路和间充质-上皮转化的调节促进多巴胺能神经元重编程，竞争性内源性RNA网络分析为神经元重编程提供了新的靶点。我们证明了软基质可以通过抑制microRNA-615- 3 p靶向整合素β亚基4来促进神经元重编程。我们的研究结果可以帮助再生疗法的发展，并有助于提高我们对神经元重编程的理解。重要性声明首先，我们已经表明，低刚度促进小分子组合的基础上直接重编程。第二，我们发现microRNA（miR）-615-3p可能与整合素β亚基4（ITGB 4）相互作用，软底物可能通过抑制miR-615- 3 p靶向ITGB 4而促进神经重编程。我们是第一个报道这一机制的，我们的发现将为后续神经再生医学的基础和临床研究提供更多的功能神经元，并将有助于提高对神经重编程的整体认识。本工作也为神经再生医用生物材料的设计提供了新的思路。
Fibroblasts can be directly reprogrammed via a combination of small molecules to generate induced neurons (iNs), bypassing intermediate stages. This method holds great promise for regenerative medicine; however, it remains inefficient. Recently, studies have suggested that physical cues may improve the direct reprogramming of fibroblasts into neurons, but the underlying mechanisms remain to be further explored, and the physical factors reported to date do not exhibit the full properties of the extracellular matrix (ECM). Previousin vitrostudies mainly used rigid polystyrene dishes, while one of the characteristics of the nativein-vivoenvironment of neurons is the soft nature of brain ECM. The reported stiffness of brain tissue is very soft ranging between 100 Pa and 3 kPa, and the effect of substrate stiffness on direct neuronal reprogramming has not been explored. Here, we show for the first time that soft substrates substantially improved the production efficiency and quality of iNs, without needing to co-culture with glial cells during reprogramming, producing more glutamatergic neurons with electrophysiological functions in a shorter time. Transcriptome sequencing indicated that soft substrates might promote glutamatergic neuron reprogramming through integrins, actin cytoskeleton, Hippo signalling pathway, and regulation of mesenchymal-to-epithelial transition, and competing endogenous RNA network analysis provided new targets for neuronal reprogramming. We demonstrated that soft substrates may promote neuronal reprogramming by inhibiting microRNA-615-3p-targeting integrin subunit beta 4. Our findings can aid the development of regenerative therapies and help improve our understanding of neuronal reprogramming.Statement of significanceFirst, we have shown that low stiffness promotes direct reprogramming on the basis of small molecule combinations. To the best of our knowledge, this is the first report on this type of method, which may greatly promote the progress of neural reprogramming.Second, we found that microRNA (miR)-615-3p may interact with integrin subunit beta 4 (ITGB4), and the soft substrates may promote neural reprogramming by inhibiting miR-615-3p targeting ITGB4. We are the first to report on this mechanism.Our findings will provide more functional neurons for subsequent basic and clinical research in neurological regenerative medicine, and will help to improve the overall understanding of neural reprogramming. This work also provides new ideas for the design of medical biomaterials for nerve regeneration.