Joe I-Seul, Cho Goang-Won
1 Department of Biology, College of Natural Science, Chosun University , Gwangju, Korea.
2 Department of Life Science, BK21-Plus Research Team for Bioactive Control Technology, Chosun University , Gwangju, Korea.
Cell Reprogram. 2016 Aug;18(4):224-9. doi: 10.1089/cell.2015.0061.
Increased intracellular cyclic adenosine monophosphate (cAMP) can promote axonal elongation and facilitate neuronal repair, while decreased cAMP is associated with losses in neuronal regenerative capacity. Rolipram, which upregulates intracellular cAMP by blocking phosphodiesterase-4 (PDE4) enzyme activity, can mitigate diverse neurological disorders. In this study, we investigated whether rolipram induces neuronal differentiation of human bone marrow-mesenchymal stem cells (hBM-MSCs). Rolipram-treated MSCs (Roli-MSCs) had significantly increased expression of the neuroprogenitor proteins Nestin, Musashi, GFAP, and Sox-2. When Roli-MSCs were differentiated with neuronal induction media (Roli-dMSCs), they exhibited cell body and dendritic morphologies similar to those of neurons. The neurite number and length of Roli-dMSCs were significantly increased compared to those of differentiated MSCs (dMSCs). Compared with undifferentiated hBM-MSCs, the Roli-dMSCs and dMSCs showed significantly increased expression of the neuronal-specific marker genes Nestin, Musashi, CD133, GFAP, NF-M, MAP-2, KCNH1, KCNH5, SCN3A, and CACNA1A, and decreased expression of other lineage-specific markers Adiponectin, ALP, FABP4, and MMP13. The Roli-dMSCs also showed a higher expression of the neuronal markers Nestin, Musashi, Sox-2, NF-M, and Tuj-1 compared to those of the undifferentiated hBM-MSCs, measured by immunocytochemistry and immunoblotting assay. Thus, we have shown that rolipram ameliorates neuronal differentiation by the regulation of neuroprogenitor expression in hBM-MSCs, and rolipram treatment of MSCs may improve the therapeutic efficacy of stem cell therapy for neurodegenerative disorders.
细胞内环状单磷酸腺苷（cAMP）水平升高可促进轴突延长并有助于神经元修复，而cAMP水平降低则与神经元再生能力丧失有关。咯利普兰通过阻断磷酸二酯酶-4（PDE4）的酶活性来上调细胞内cAMP，可减轻多种神经系统疾病。在本研究中，我们调查了咯利普兰是否能诱导人骨髓间充质干细胞（hBM-MSCs）向神经元分化。经咯利普兰处理的间充质干细胞（Roli-MSCs）中神经祖细胞蛋白巢蛋白（Nestin）、神经干细胞蛋白（Musashi）、胶质纤维酸性蛋白（GFAP）和性别决定区Y框蛋白2（Sox-2）的表达显著增加。当用神经元诱导培养基诱导Roli-MSCs分化（Roli-dMSCs）时，它们呈现出与神经元相似的细胞体和树突形态。与分化的间充质干细胞（dMSCs）相比，Roli-dMSCs的神经突数量和长度显著增加。与未分化的hBM-MSCs相比，Roli-dMSCs和dMSCs中神经元特异性标记基因巢蛋白、神经干细胞蛋白、CD133、GFAP、神经丝中链（NF-M）、微管相关蛋白2（MAP-2）、钾通道亚家族H成员1（KCNH1）、钾通道亚家族H成员5（KCNH5）、钠通道蛋白3亚基A（SCN3A）和电压依赖性钙通道α1亚基A（CACNA1A）的表达显著增加，而其他谱系特异性标记脂联素、碱性磷酸酶（ALP）、脂肪酸结合蛋白4（FABP4）和基质金属蛋白酶13（MMP13）的表达降低。通过免疫细胞化学和免疫印迹分析检测，与未分化的hBM-MSCs相比，Roli-dMSCs中神经元标记物巢蛋白、神经干细胞蛋白、Sox-2、NF-M和βIII微管蛋白（Tuj-1）的表达也更高。因此，我们表明咯利普兰通过调节hBM-MSCs中神经祖细胞的表达来改善神经元分化，并且对间充质干细胞进行咯利普兰处理可能会提高干细胞疗法对神经退行性疾病的治疗效果。
