Zhuo Yi, Yuan Ting, Duan Da, Wang Lei, Ge Lite, Wu Pei, Wang Hao, Lu Ming
Department of Neurosurgery, 163 Hospital of PLA/ Second Affiliated Hospital, Hunan Normal University, Changsha 410003, China.
Zhong Nan Da Xue Xue Bao Yi Xue Ban. 2016 Dec 28;41(12):1252-1259. doi: 10.11817/j.issn.1672-7347.2016.12.002.
To explore whether hypoxic condition could promote the olfactory mucosa mesenchymal stem cells (OM-MSCs) to differentiate into neurons with the olfactory ensheathing cells (OECs) supernatant and the potential mechanisms. Methods: The OM-MSCs and OECs were isolated and cultured, and they were identified by flow cytometry and immunofluorescence. The OM-MSCs were divided into three groups: a 3%O2+ HIF-1α inhibitors (lificiguat: YC-1) + OECs supernatant group (Group A) , a 3%O2 + OECs supernatant group (Group B) and a 21%O2 + OECs supernatant group (Control group). The neurons, which were differentiated from OM-MSCs, were assessed by immunofluorescence test. The mRNA and protein expression of hypoxia-inducible factor-1α (HIF-1α), βIII-tubulin and glial fibrillary acidic portein (GFAP) were detected by quantitative polymerase chain reaction (Q-PCR) and Western blot. The potassium channels were analyzed by patch clamp. Results: The neurons differentiated from OM-MSCs expressed the most amount of βIII-tubulin, and the result of Q-PCR showed that HIF-1α expression in the Group B was significantly higher than that in the other groups (all P<0.05). Western blot result showed that the βIII-tubulin protein expression was significantly higher and GFAP protein expression was obviously decreased in the Group B (both P<0.05). The patch clamp test confirmed that the potassium channels in the neurons were activated. Conclusion: Hypoxic condition can significantly increase the neuronal differentiation of OM-MSCs by the OECs supernatant and decrease the production of neuroglia cells, which is associated with the activation of HIF-1 signal pathway.
探讨低氧条件下嗅黏膜间充质干细胞（OM-MSCs）能否在嗅鞘细胞（OECs）上清作用下分化为神经元及其潜在机制。方法：分离培养OM-MSCs和OECs，通过流式细胞术和免疫荧光进行鉴定。将OM-MSCs分为三组：3%O₂ + HIF-1α抑制剂（利西胍：YC-1）+ OECs上清组（A组）、3%O₂ + OECs上清组（B组）和21%O₂ + OECs上清组（对照组）。通过免疫荧光试验评估由OM-MSCs分化而来的神经元。采用定量聚合酶链反应（Q-PCR）和蛋白质免疫印迹法检测缺氧诱导因子-1α（HIF-1α）、βIII-微管蛋白和胶质纤维酸性蛋白（GFAP）的mRNA及蛋白表达。采用膜片钳技术分析钾通道。结果：由OM-MSCs分化而来的神经元中βIII-微管蛋白表达量最高，Q-PCR结果显示B组HIF-1α表达显著高于其他组（均P<0.05）。蛋白质免疫印迹结果显示B组βIII-微管蛋白表达显著升高，GFAP蛋白表达明显降低（均P<0.05）。膜片钳试验证实神经元中的钾通道被激活。结论：低氧条件下OECs上清可显著增加OM-MSCs向神经元的分化，减少神经胶质细胞的产生，这与HIF-1信号通路的激活有关。
