Graduate School of Humanities and Sciences, Otsuka, Bunkyo-ku, Tokyo, Japan; Institute for Human Life Innovation, Ochanomizu University, Otsuka, Bunkyo-ku, Tokyo, Japan.
Graduate School of Humanities and Sciences, Otsuka, Bunkyo-ku, Tokyo, Japan; Institute for Human Life Innovation, Ochanomizu University, Otsuka, Bunkyo-ku, Tokyo, Japan.
Neurosci Lett. 2021 Feb 16;746:135648. doi: 10.1016/j.neulet.2021.135648. Epub 2021 Jan 11.
Vitronectin, an extracellular matrix protein, controls the differentiation of cerebellar granule cell precursors (CGCPs) via αvβ5 integrin, particularly in the initial stage of differentiation to granule cells. In this study, we determined whether vitronectin regulates axon specification in this initial differentiation stage of CGCPs. First, we analyzed whether vitronectin deficiency, β5 integrin knockdown (KD), and β5 integrin overexpression affect axon specification of primary cultured CGCPs. Vitronectin deficiency and β5 integrin KD inhibited axon formation, while vitronectin administrated- and β5 integrin overexpressed-neurons formed multiple axons. Moreover, KD of β5 integrin suppressed vitronectin-induced multiple axon formation. These findings indicate that vitronectin contributes to regulating axon specification via αvβ5 integrin in CGCPs. Next, we determined the signaling pathway involved in regulating vitronectin-induced axon specification. Wortmannin, an inhibitor of phosphatidylinositol 3-kinase (PI3K), inhibited vitronectin-induced multiple axon specification, and lithium chloride, an inhibitor of glyocogen synthase kinase 3 beta (GSK3β), attenuated the inhibitory effect of vitronectin-KO and β5 integrin KD on the specification of CGCPs. In addition, vitronectin induced the phosphorylation of protein kinase B (Akt) and GSK3β in neuroblastoma Neuro2a cells. Taken together, our results indicate that vitronectin plays an important factor in axon formation process in CGCPs via a β5 integrin/PI3K/GSK3β pathway.
纤连蛋白是一种细胞外基质蛋白，通过αvβ5 整联蛋白控制小脑颗粒细胞前体细胞（CGCP）的分化，特别是在向颗粒细胞分化的初始阶段。在本研究中，我们确定纤连蛋白是否在 CGCP 这个初始分化阶段调节轴突的特化。首先，我们分析了纤连蛋白缺陷、β5 整联蛋白敲低（KD）和β5 整联蛋白过表达是否影响原代培养 CGCP 的轴突特化。纤连蛋白缺陷和β5 整联蛋白 KD 抑制轴突形成，而纤连蛋白处理和β5 整联蛋白过表达的神经元形成多个轴突。此外，β5 整联蛋白的 KD 抑制了纤连蛋白诱导的多轴突形成。这些发现表明，纤连蛋白通过αvβ5 整联蛋白在 CGCP 中有助于调节轴突的特化。接下来，我们确定了参与调节纤连蛋白诱导的轴突特化的信号通路。PI3K 的抑制剂 wortmannin 抑制了纤连蛋白诱导的多个轴突特化，而糖原合酶激酶 3β（GSK3β）的抑制剂氯化锂减弱了纤连蛋白-KO 和β5 整联蛋白 KD 对 CGCP 特化的抑制作用。此外，纤连蛋白诱导神经母细胞瘤 Neuro2a 细胞中蛋白激酶 B（Akt）和 GSK3β的磷酸化。综上所述，我们的结果表明，纤连蛋白通过β5 整联蛋白/PI3K/GSK3β 途径在 CGCP 的轴突形成过程中发挥重要作用。
