Abstract：
During neurodevelopment, differentiation of neural stem/progenitor cells (NSPCs) into neurons are regulated by many factors including Notch signaling pathway. Herein, we report the effect of a Notch signaling blocker, i.e. γ -secretase inhibitor (GSI), on this differentiating process, especially on the morphological development. NSPCs were cultured and induced to differentiate with or without GSI. The neurite outgrowth was impeded by GSI application and the expression of a Notch signaling downstream effector miR-342-5p increased with the downregulated expression of Notch effectors Hes1 and Hes5. Upregulated expression of miR-342-5p in differentiating NSPCs could shorten the neurite length of progeny neurons, which was similar to the effect of GSI. To avoid the possible influence from astrocytes into neurons, we directly applied cultured neurons, on which GSI could shorten the processes and RBP-J knockdown could also reduce the neurite length. Similarly, transfection of miR-342-5p mimics or inhibitors into PC12 cells led to shorter or longer processes of cells compared with control ones. Furthermore, in differentiating NSPCs, GSI-induced shorter neurites could be partially rescued by miR-342-5p inhibitors, and STAT3 was one of the possible targets of miR-342-5p during this differentiating process as indicated by results of Western Blot test, luciferase reporter assay and GFP reporter assay. To further demonstrate the role of STAT3, it was introduced into GSI-treated neurons and the GSI-affected neurites could also be partially rescued. In conclusion, GSI could influence the morphological development of neurons and the possible mechanism involved Notch/miR-342-5p and STAT3. These results would be informative for future therapeutic research.
摘要：
在神经发育过程中,神经干/祖细胞（NSPCs）向神经元的分化受Notch信号通路等多种因素的调控。在这里，我们报道了Notch信号阻断剂的作用,即γ-分泌酶抑制剂（GSI），在这个分化过程中，特别是在形态发育方面。在有或没有GSI的情况下培养NSPC并诱导分化。神经突生长受到GSI应用的阻碍,并且Notch信号传导下游效应子miR-342-5p的表达随着Notch效应子Hes1和Hes5的表达下调而增加。miR-342-5p在分化NSPCs中的上调表达可以缩短子代神经元的神经突长度，这与GSI的效果相似。为了避免星形胶质细胞对神经元的可能影响，我们直接应用培养的神经元，GSI可以缩短过程，RBP-J敲低也可以减少神经突长度。同样，与对照组相比,将miR-342-5p模拟物或抑制剂转染到PC12细胞中导致细胞过程更短或更长.此外，在区分NSPC方面，GSI诱导的较短的神经突可以通过miR-342-5p抑制剂部分挽救，并且STAT3是miR-342-5p在这个分化过程中可能的靶标之一，如WesternBlot测试的结果表明，荧光素酶报告测定和GFP报告测定。为了进一步证明STAT3的作用,将其引入GSI处理的神经元中,并且还可以部分挽救受GSI影响的神经突。总之,GSI可影响神经元的形态发育，其可能机制涉及Notch/miR-342-5p和STAT3。这些结果将为未来的治疗研究提供信息。
