Study of regeneration of peripheral nervous system with adenoviral vectors

对于周围神经的再生，轴突的延长和髓鞘的形成是必不可少的。雪旺细胞是周围神经系统的髓鞘胶质细胞，其发育受多种生长因子的调节。然而，这些配体刺激后的细胞内信号通路在雪旺细胞分化中的机制仍然不清楚。在这里，我们证明，在培养的雪旺细胞，中性调节蛋白和PDGF抑制髓鞘相关蛋白标记物的表达，而IGF-I促进它的表达。虽然这些配体激活了常见的下游信号通路，即细胞外信号调节激酶(ERK)和磷脂酰肌醇-3-激酶(PI3K)/Akt通路，但不同配体的激活情况有所不同。为了阐明这些通路的功能和雪旺细胞分化的机制，我们使用腺病毒载体选择性地激活或失活这些通路。在大鼠嗜铬细胞瘤细胞系PC12细胞中，CA-MEK病毒感染诱导ERKs持续激活，并在没有神经营养因子的情况下刺激轴突生长。我们发现ERK通路的选择性激活抑制了雪旺细胞的分化，而PI3K通路的选择性激活促进了雪旺细胞的分化。通过基因转移选择性激活雪旺细胞中的PI3K通路，进一步证实了体外雪旺细胞-背根神经节神经元共培养和体内同种异体神经移植实验中髓鞘形成的增加。我们的结论是，PI3K/Akt介导的信号对髓鞘的启动至关重要，生长因子的作用在很大程度上依赖于Erk和PI3K/Akt激活之间的平衡。我们的结果还提出了根据未来的细胞疗法通过调节细胞内信号来增强雪旺细胞功能的可能性。

For regeneration of peripheral nerves, the elongation of axons and their myelination are essential. Schwann cells are the myelinating glia of the peripheral nervous system and their development is regulated by various growth factors. However, the mechanisms of intracellular signaling pathways following these ligands stimuli in Schwann cells differentiation remain elusive. Here, we demonstrated that, in cultured Schwann cells, neuregulin and PDGF suppressed the expression of myelin associated protein markers, while IGF-I promoted it. Although these ligands activated common downstream signaling pathways, i.e., extracellular signal-regulated kinase (Erk) and phosphatidylinositol-3-kinase (PI3K)/Akt pathways, the profiles of activation varied among ligands. To elucidate the function of these pathways and the mechanisms underlying Schwann cell differentiation, we used adenoviral vectors to selectively activate or inactivate these pathways. In rat pheocromocytoma cell line PC12 cells, CA-MEK virus infection induced sustained activation of ERKs and stimulated neurite outgrowth in the absence of neurotrophic factors. We found that the selective activation of Erk pathways suppressed Schwann cell differentiation, while that of PI3K pathways promoted it. Selective activation of PI3K pathways in Schwann cells by gene transfer further demonstrated increased myelination in in vitro Schwann cell-DRG neuron cocultures and in vivo allogenic nerve graft experiment. We conclude that signals mediated by PI3K/Akt are crucial for initiation of myelination and the effects of growth factors are largely dependent on the balance between Erk and PI3K/Akt activation. Our results also propose the possibilities to augment Schwann cell functions by modulating intracellular signals in the light of future cell therapies.