Klimovich Polina, Rubina Kseniya, Sysoeva Veronika, Semina Ekaterina
Laboratory of molecular endocrinology, National Cardiology Research Center Ministry of Health of the Russian Federation, Institute of Experimental Cardiology, Moscow 121552, Russia.
Faculty of Medicine, Lomonosov Moscow State University, Moscow 119991, Russia.
Biomedicines. 2020 Mar 3;8(3):49. doi: 10.3390/biomedicines8030049.
Neurotrophiс factors play a key role in the development, differentiation, and survival of neurons and nerve regeneration. In the present study, we evaluated the effect of certain neurotrophic factors (NGF, BDNF, and GDNF) on axon growth and migration of Nestin-green fluorescent protein (GFP)-positive cells using a 3D model of dorsal root ganglion (DRG) explant culture in Matrigel. Our method generally represents a convenient model for assessing the effects of soluble factors and therapeutic agents on axon growth and nerve regeneration in R&D studies. By analyzing the DRG explants in ex vivo culture for 21 days, one can evaluate the parameters of neurite outgrowth and the rate of cell migration from the DRG explants into the Matrigel. For the current study, we used Nestin-GFP-expressing mice in which neural precursors express Nestin and the green fluorescent protein (GFP) under the same promoter. We revealed that GDNF significantly (two fold) stimulated axon outgrowth ( < 0.05), but not BDNF or NGF. It is well-known that axon growth can be stimulated by activated glial cells that fulfill a trophic function for regenerating nerves. For this reason, we evaluated the number of Nestin-GFP-positive cells that migrated from the DRG into the Matrigel in our 3D ex vivo explant model. We found that NGF and GDNF, but not BDNF, stimulated the migration of Nestin-GFP cells compared to the control ( < 0.05). On the basis of the aforementioned finding, we concluded that GDNF had the greatest stimulating potential for axon regeneration, as it stimulated not only the axon outgrowth, but also glial cell migration. Although NGF significantly stimulated glial cell migration, its effect on axon growth was insufficient for axon regeneration.
神经营养因子在神经元的发育、分化、存活以及神经再生过程中发挥着关键作用。在本研究中，我们使用基质胶中背根神经节（DRG）外植体培养的三维模型，评估了某些神经营养因子（神经生长因子、脑源性神经营养因子和胶质细胞源性神经营养因子）对巢蛋白-绿色荧光蛋白（GFP）阳性细胞轴突生长和迁移的影响。我们的方法总体上为评估研发研究中可溶性因子和治疗药物对轴突生长和神经再生的影响提供了一个便捷的模型。通过对体外培养21天的DRG外植体进行分析，可以评估神经突生长参数以及细胞从DRG外植体迁移到基质胶中的速率。在本研究中，我们使用了表达巢蛋白-GFP的小鼠，其中神经前体细胞在同一启动子下表达巢蛋白和绿色荧光蛋白（GFP）。我们发现胶质细胞源性神经营养因子显著（两倍）刺激了轴突生长（P<0.05），但脑源性神经营养因子或神经生长因子则没有。众所周知，活化的神经胶质细胞可以刺激轴突生长，这些神经胶质细胞对再生神经具有营养作用。因此，我们在三维体外外植体模型中评估了从DRG迁移到基质胶中的巢蛋白-GFP阳性细胞的数量。我们发现，与对照组相比，神经生长因子和胶质细胞源性神经营养因子而非脑源性神经营养因子刺激了巢蛋白-GFP细胞的迁移（P<0.05）。基于上述发现，我们得出结论，胶质细胞源性神经营养因子对轴突再生具有最大的刺激潜力，因为它不仅刺激了轴突生长，还刺激了神经胶质细胞迁移。尽管神经生长因子显著刺激了神经胶质细胞迁移，但其对轴突生长的影响不足以实现轴突再生。
